Isolation and Characterization of Lactic Acid Bacteria from Raw Cow Milk and Evaluation of Their Probiotic Potential

Isolation and Characterization of Lactic Acid Bacteria from Raw Cow Milk and Evaluation of Their Probiotic Potential | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Isolation and Characterization of Lactic Acid Bacteria from Raw Cow Milk and Evaluation of Their Probiotic Potential Goiteom Senay Niguse, Desta Berhe Sbhatu, Gerbreselema Gebreyohannes This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8906693/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Lactic acid bacteria (LAB) have diverse applications in dairy, food, feed, and health, yet their probiotic potential from cow milk remains under-researched. This study aimed to identify and characterize LAB isolates from raw cow milk and evaluate their probiotic suitability. A total of 23 pure bacterial isolates were obtained from seven milk samples, with nine gram-positive, catalase-negative isolates selected for further analysis. These isolates were citrate-negative, non-motile, and indole-negative, with seven exhibiting homo-fermentative traits, and two (M5 and M9) identified as hetero-fermentative. Physicochemical analysis of milk samples showed notable variation in composition, including protein (2.2–3.2%), lactose (3.3–4.4%), fat (1.6–2.6%), ash (0.4–0.6%), salt (0.4–0.6%), titratable acidity (0.1–0.4%), pH (5.7–6.7%), moisture (83.9–87.5%), density (25.9–27.3), and freezing point (0.4–0.6). Antibacterial tests revealed strong inhibition of S. aureus by M1 (11 mm) and P. aeruginosa by M6 and M8 (9 mm and 13 mm), while E. coli showed complete resistance. Hemolysis tests confirmed LAB isolates as non-pathogenic, with no α- or β-hemolysis observed. Antibiotic susceptibility varied, with M2, M3, M4, and M8 completely resistant, while M1, M5, M6, M7, and M9 showed susceptibility to meropenem, ampicillin, penicillin G, and other antibiotics. Sensory evaluation indicated M1 and M7 had the highest overall acceptability, while M3 and M8 scored lower in color and taste. LAB isolates were tentatively classified under Lactobacillus, Enterococcus , and Leuconostoc , indicating their probiotic potential and viability as starter cultures in dairy and food industries. Further molecular identification and characterization are needed to determine specific strains and commercial applications. Lactic acid bacteria Milk Probiotics Starter culture Yogurt 1. INTRODUCTION Milk, a nutrient-dense secretion from mammary glands composed of water, fat, proteins, lactose, minerals, and vitamins, is essential for human nutrition. In Ethiopia, cattle contribute the largest portion of annual milk production (81.2%), followed by goats (7.9%), camels (6.3%), and sheep (4.6%) (Bereda et al., 2014 ). Milk and its derivatives are vital for human health and nutrition and also provide a conducive environment for the proliferation of lactic acid bacteria (LAB), which have been integral to food fermentation and preservation for centuries (Hatti-kaul et al., 2018 ). LAB's metabolic capability allows them to convert carbohydrate substrates into organic acids, predominantly lactic acid (LA), while also producing diverse metabolites with numerous health benefits (Rodriguez et al ., 2019). Historically, LAB have served as starter cultures for the fermentation of a wide variety of foods, including milk, vegetables, meat, fish, cereals, and animal feed, contributing to flavor, aroma, and extended shelf life, thus making them invaluable in food production (Kunene et al., 2000 ). Notably, fermented dairy products, valued at over 80 billion euros in 2011, held a particularly significant position in the global market (Hatti-kaul et al., 2018 ). LABs are used in food fermentation and functional starter cultures to produce beneficial compounds like Extracellular Polymeric Substance (EPS), organic acids, polyols, aromatic compounds, and bacteriocins, which improve food product texture, aroma, flavor, health benefits, and shelf life (Aritonang et al., 2022 ). LAB-derived lactic acid, known for its bio-preservative and antimicrobial properties, is highly valuable in various industries like food, cosmetics, pharmaceuticals, and chemicals. Its demand is driven by its role in poly-lactic acid production, biodegradable thermoplastic polyester used in packaging materials, films, fibers, and medical implants. Selected LAB serve as probiotics, enhancing human health by improving intestinal health, enhancing immune response, reducing cholesterol, and mitigating cancer risk. They also contribute to disease prevention, allergic response regulation, and gastrointestinal health maintenance (Fijan, 2014; Somashekaraiah et al ., 2019; Aritonang et al., 2022 ). The global market for LAB, valued at USD 2.08 billion in 2016, is expected to grow due to increasing demand in packaging, personal care, and textiles, driven by the isolation and characterization of LAB from traditional raw and fermented foods (Hatti-kaul et al., 2018 ). LAB, found in traditional foods like milk, is uncharacterized and requires further investigation. Recognized as "Generally Regarded as Safe" (GRAS), LAB produce enzymes that break down and synthesize food components like carbohydrates, proteins, and lipids, essential for survival and widely used in food processing and industrial applications (Amenu & Bacha, 2023 ). LAB are essential for fermented foods, providing benefits such as improved gut health, enhanced immune response, and potential anti-cancer properties. However, the growing challenge of antibiotic resistance has highlighted the need for probiotics, particularly LAB, to promote livestock health. Despite their potential, the identification and characterization of LAB in milk and dairy products remain insufficient due to environmental variables, dairy management practices, and microbial interactions. This study aims to characterize and evaluate LAB isolates from raw cow milk to expand the understanding of their application in dairy fermentation and probiotic development. 2. MATERIALS AND METHODS 2.1 Milk Sampling and Physicochemical & Compositional Analysis Cow milk samples were collected from producers and sellers in the sub-cities of Mekelle using sterilized, sealed bottles. The samples were stored in an ice box and transported to the microbiology laboratory at Mekelle University, where they were kept at 4°C until analysis. The physicochemical properties (pH, density, freezing point, and moisture content) and compositional attributes (ash content, protein, fat, lactose, and titratable acidity) were analyzed using a Lacto-Scan milk analyzer (Olugbuyiro & Oseh, 2011 ; Asefa & Teshome, 2019 and Sharma et al., 2021 ). 2.2 Isolation and Morphological Characterization of LAB LAB isolation followed the method of Ismail1 et al ., (2018), starting with serial dilutions of the samples. A 0.1 ml aliquot was spread onto de Man, Rogosa, and Sharpe (MRS) agar plates, which were incubated at 37°C under aerobic conditions for 48–72 hours (Taye et al., 2021 ). Morphological characterization of the isolates involved observing colony and cell morphology (shape, size, and color) under a microscope (Lingga et al., 2023 ). Gram staining of pure cultures was performed by applying a bacterial culture to a heat-fixed slide, followed by sequential treatment with crystal violet (1 minute, rinsed), iodine solution (1 minute, rinsed), 95% alcohol (30 seconds, rinsed), and safranin (30 seconds, rinsed). After a final rinse, the slide was observed under a 100x objective lens, with Gram-positive bacteria appearing purple and Gram-negative bacteria appearing red. 2.3 Biochemical Characterization and Identification of LAB Isolates The LAB's biochemical characterization involved tests to assess their metabolic capabilities, with the catalase test indicating the presence of the enzyme by producing gas bubbles when 3% hydrogen peroxide was added to a pure isolate (Asha et al., 2024 ). To assess their fermentative pathways, the gas production test utilized nutrient broth with an inverted Durham tube, where incubation at 37°C for 48 hours allowed for the differentiation of homo-fermentative LAB, which produce no gas, from hetero-fermentative LAB, which generate gas trapped in the Durham tube (Prastujati et al., 2022 ). Further carbohydrate fermentation was evaluated using Triple Sugar Iron Agar (TSIA), with a yellow color change after 24 hours at 37°C signifying the fermentation of glucose, sucrose, and/or lactose, and the absence of blackening indicating no hydrogen sulfide production (Tigistu et al., 2022 ). The ability of the isolates to utilize citrate as a sole carbon source was tested using Simmons citrate medium, where a color change from green to blue or yellow after 24 hours at 37°C indicated a positive result (Asha et al., 2024 ). Additional biochemical characteristics were assessed using SIM and MR tests. Motility was determined in SIM media by observing the spreading of bacterial colonies after 24 hours of incubation at 37°C. The production of indole was detected in the same incubated SIM media by the addition of Kovac's reagent, with the formation of a red layer at the top indicating a positive result (Ngene et al., 2019 ). Finally, the Methyl Red (MR) test, performed on isolates grown in MR-VP media for 24 hours at 37°C, identified acid producers by a change in the medium's color to red upon the addition of the MR indicator (Ngene et al., 2019 ). 2.4 Physiological Characterization of LAB isolates The physiological characteristics of the LAB isolates were evaluated through several tolerance tests. Temperature resistance was assessed by culturing the isolates in MRS broth and observing their growth after 48 hours of incubation at 15°C, 27°C, 37°C, and 45°C (Lingga et al., 2023 ). Sodium chloride (NaCl) tolerance was determined by initially culturing the isolates in MRS broth for 24 hours, followed by subsequent 24-hour incubation in MRS broth supplemented with 1%, 4%, and 6% NaCl. These cultures were then plated on MRS agar and incubated for 48 hours, after which colony growth was observed and recorded (Asha et al., 2024 ). To test survival in acidic conditions, the isolates were incubated in MRS broth at pH levels of 6.5, 6, 4, and 2 for 24 hours, then streaked onto MRS agar and incubated at 37°C for 48 hours to monitor colony growth (Safi et al., 2023). Finally, phenol resistance was assessed by culturing the LAB isolates in MRS broth for 24 hours and then transferring them onto MRS agar supplemented with 0.4% and 0.6% phenol solution, with growth observed after 48 hours of incubation at 37°C. 2.5 Safety Evaluation of LAB isolates The safety evaluation of LAB isolates involved hemolysis, antibacterial activity, and antibiotic susceptibility tests. The hemolysis test was conducted by growing LAB colonies on blood agar supplemented with sheep blood and incubating them at 37°C for 24 hours (Amenu & Bacha, 2023 ). Hemolytic activity was assessed by holding the plate up to a light source, with control strains including E. coli (ATCC 29922), S. aureus (ATCC 25923), and S. typhimurium (ATCC 13311) Meradji et al., ( 2023 ). Antibacterial activity was tested against S. aureus (ATCC 25923), E. coli (ATCC 25922), and P. aeruginosa (ATCC 27853) using the agar well diffusion method (Amenu & Bacha, 2023 ). Inhibition zones around the wells were categorized as resistant (R) (no zone), weak inhibition ( 8–12 mm) (sensitive) (Akabanda et al., 2014 ). Additionally, the antibiotic susceptibility of the isolates to penicillin G (10 µg), ampicillin (10 µg), cotrimoxazole (25 µg), vancomycin (30 µg), meropenem (10 µg), tetracycline (30 µg), ceftriaxone (30 µg), clindamycin (2 µg), ciprofloxacin (5 µg), erythromycin (15 µg), levofloxacin (5 µg), and amikacin (10 µg) was determined using the agar disc diffusion method (Bauer et al., 1966 ), with inhibition zones categorized as susceptible (S) (≥ 21 mm), intermediate (I) (16–20 mm), or resistant (R) (≤ 15 mm) (Bauer et al., 1966 ). 2.6 Evaluation of Potent isolate as a Starter Culture The evaluation of potent LAB isolates as a starter culture involved the formation of fermented milk (yogurt). Following (Ismail et al., 2016 ), fresh raw cow milk was procured from a local farm in Mekelle. The milk was pasteurized at 80–90°C for 15 minutes to eliminate existing microbes, then cooled to approximately 40°C (Lee & Lucey, 2010 ). It was then inoculated with one colony or 1% of the LAB strain, ensuring a cell count of 1 × 10⁶, and poured into sterilized bottles (Sharma et al., 2021 ). A total of 550 mL of milk was used for yogurt formation, with 50 mL allocated per sample. Fermentation occurred at room temperature for 20–24 hours (Ismail1 et al ., 2018). The physiochemical properties of yogurt, including pH, freezing point, titratable acidity, and density, were analyzed using a Lacto-Scan milk analyzer (Olugbuyiro & Oseh, 2011 ; Asefa & Teshome, 2019 ; Sharma et al., 2021 ). The analyzer was cleaned using Lacto Daily and Lacto Weekly solutions and rinsed three times with distilled water before analysis. A 25 mL sample was placed in a sample holder, activated via the start button, and measured, with results displayed digitally. The composition of yogurt, including ash content, protein, lactose, fat, and moisture, was also assessed using the Lacto-Scan milk analyzer (Olugbuyiro & Oseh, 2011 ; Sharma et al., 2021 ). 2.7 Sensory Evaluation of Yogurt Sensory evaluation of yogurt was conducted following a previous method (Ademosun et al., 2019 ). Twelve trained panelists—six women and six men, aged 22–45—participated in the evaluation, including undergraduate and graduate students, as well as teaching and non-teaching staff members of Mekelle University. All panelists were familiar with conventional natural cow milk yogurt. The yogurt samples were assessed for organoleptic characteristics and overall acceptability using a 7-point hedonic scale: like extremely (7), like moderately (6), like slightly (5), neither like nor dislike (4), dislike slightly (3), dislike moderately (2), and dislike extremely (1). Sensory properties were evaluated based on fermented color, fermented odor, and taste quality. To minimize bias, each sample group was coded before being served as cooled yogurt to the panelists. The results were generated based on two trials and recorded using a table with mean values. 2.8 Identification of LAB at Genus Level The identification and characterization of LAB isolates relied on their morphological, biochemical, and physiological properties. Morphological analysis included observing colony shape, size, and color. Biochemical tests assessed metabolic activities such as catalase production and carbohydrate fermentation profiles. Physiological characterization evaluated their adaptability and functional potential by determining their resistance to various temperatures, tolerance to sodium chloride, survival in acidic conditions, and resistance to phenol (Yelnetty et al., 2014 ; Hafsan and Mustami, 2015). 2.9 Data Analysis Data were recorded using Microsoft Excel, and differences between the means were determined through one-way analysis of variance (ANOVA). Quantitative data were further analyzed using the Fisher Least Significant Difference (LSD) method with Minitab software (Version 19). Statistical significance was established at a P-value of less than 0.05. 3. RESULTS AND DISCUSSION 3.1 Physicochemical Properties and Composition of Raw Cow Milk The study found significant variations in protein and lactose content in milk samples, with highest protein levels in S2 and lowest in S3. These average values differ from previous studies but fall within the ranges of other studies, likely influenced by factors like forage quality (Asefa & Teshome, 2019 ; Meskel & Sebho, 2018). The study found that lactose content in S5 was highest, while in S1 and S4, it was lower, indicating the significant impact of environmental conditions and animal breeding, with crossbred animals showing higher lactose levels (Elnour et al ., 2022; Ibrahim et al ., 2021), Ketema et al . (2018). The physical and chemical properties of the milk samples exhibited notable variations. Density was highest in S5 and lowest in S3 (Table 3 ), aligning with previous findings linking it to fat and solids-not-fat content (Meskel & Sebho, 2018). In contrast, the freezing point (FP) varied, with S2 displaying the highest value, while S3, S4, and S6 had the lowest (Table 3 ), a finding that underscores the influence of soluble milk constituents, particularly lactose and minerals, and contradicts Asefa & Teshome ( 2019 ). The pH values of the milk ranged from the highest in S6 to the lowest in S4 (Table 3 ), consistent with previously reported ranges (Elnour et al., 2022; Asefa & Teshome, 2019 ). Salt content was most elevated in S7, with S1, S2, and S4 exhibiting the lowest levels (Table 2 ), a finding influenced by genetic factors, feed composition, and environmental conditions. Likewise, fat content was highest in S2 and lowest in S1 (Table 2 ), and these variations align with findings from other studies (Zebib et al., 2023; Ibrahim et al., 2021; Asefa & Teshome, 2019 ; Meskel & Sebho, 2018), underscoring the significant impact of feed, genetics, breed, and lactation stage (Ketema et al., 2018). Titratable acidity (T.A), ash, and moisture content varied across the milk samples. T.A, a key indicator of freshness, peaked in S7 and was lowest in S6 (Table 2 ), aligning with the range reported by Asefa & Teshome ( 2019 ). Ash content, reflecting mineral composition, was highest in S7 and lowest in S1, S2, and S4 (Table 2 ), showing discrepancies with previous findings (Elnour et al., 2022; Asefa & Teshome, 2019 ), likely due to variations in mineral matter, as noted by Zebib et al. (2023). Moisture content, influenced by animal species and feed, was highest in S1 and S4, while S7 had the lowest value (Table 2 ), aligning with reported findings (Ibrahim et al., 2021; Elnour et al., 2022). Table 2 Physiochemical properties and Compositional Analysis of Milk Samples Samples Composition (%) Physiochemical properties P L F A S T.A pH M D F.P S1 2.4 ± 0.06 3.3 ± 0.22 1.6 ± 0.19 0.4 ± 0.02 0.4 ± 0.02 0.2 ± 0.03 6.5 ± 0.04 87.5 ± 0.03 27.0 ± 0.09 0.5 ± 0.03 S2 3.2 ± 0.07 3.6 ± 0.03 2.6 ± 0.13 0.4 ± 0.06 0.4 ± 0.06 0.3 ± 0.04 5.8 ± 0.22 86.7 ± 0.06 27.0 ± 0.07 0.6 ± 0.05 S3 2.2 ± 0.25 4.1 ± 0.04 2.1 ± 0.15 0.5 ± 0.04 0.5 ± 0.04 0.3 ± 0.06 6.1 ± 0.18 85.8 ± 0.10 25.9 ± 0.83 0.4 ± 0.07 S4 3.1 ± 0.15 3.3 ± 0.16 1.9 ± 0.13 0.4 ± 0.05 0.4 ± 0.05 0.3 ± 0.06 5.7 ± 0.22 87.5 ± 0.15 26.4 ± 0.17 0.4 ± 0.04 S5 2.8 ± 0.09 4.4 ± 0.15 1.8 ± 0.11 0.5 ± 0.01 0.5 ± 0.01 0.2 ± 0.03 6.5 ± 0.20 84.8 ± 0.10 27.3 ± 0.15 0.5 ± 0.01 S6 2.4 ± 0.08 3.6 ± 0.13 2.2 ± 0.29 0.5 ± 0.08 0.5 ± 0.08 0.1 ± 0.06 6.7 ± 0.04 86.1 ± 0.11 26.2 ± 0.03 0.4 ± 0.04 S7 2.6 ± 0.15 4.0 ± 0.01 2.1 ± 0.21 0.6 ± 0.06 0.6 ± 0.06 0.4 ± 0.05 6.2 ± 0.03 83.9 ± 0.25 26.8 ± 0.46 0.5 ± 0.04 Key : Values were given mean ± SD from triplicate experiment. S-sample, P-protein, L-lactose, F-fat, A- ash, S-salt & T.A- Taitrable acidity, D-density, F.P-freezing point and M-moisture 3.2 Morphological Characterization and Identification of LAB The study identified 23 pure bacterial isolates, with 78% showing gram-positive patterns and 22% showing gram-negative ones. Microscopic examination identified them as cocci (35%), rods or bacilli (26%), and coccobacilli (39%). Cocci and coccobacilli were observed in similar percentages within the gram-positive group (Hafsan & Mustami, 2015). Table 3 Cultural and Microscopic Characteristics of LAB Isolates Isolates G-staining Morphology Color Isolate 1 + Coccibacillus Purple Isolate 2 - Coccibacillus Red Isolate 3 + Coccibacillus Purple Isolate 4 + Cocci Purple Isolate 5 + Cocci Purple Isolate 6 + Bacillus Purple Isolate 7 + Cocci Purple Isolate 8 + Cocci Purple Isolate 9 - Coccibacillus Red Isolate 10 + Bacillus Purple Isolate 11 + Bacillus Purple Isolate 12 + Cocci Purple Isolate 13 - Coccibacillus Red Isolate 14 + Cocci Purple Isolate 15 + Coccibacillus Purple Isolate 16 + Coccibacillus Purple Isolate 17 + Coccibacillus Purple Isolate 18 - Bacillus Red Isolate 19 + Coccibacillus Purple Isolate 20 - Cocci Red Isolate 21 + Cocci Purple Isolate 22 + Bacillus Purple Isolate 23 + Bacillus Purple 3.3 Biochemical Characterization and Identification of LAB The study found that out of the total bacterial isolates, only nine isolates were catalase-negative, indicating that 61% were catalase-positive and 39% were catalase-negative. Therefore, only Gram-positive isolates without gas bubble formation were selected for further investigation (Kabir et al., 2022; Nandha & Shukla, 2023; Goicochea-Vargas et al., 2024). Nine selected isolates, numbered M1 through M9, were identified based on their Gram-positive, catalase-negative characteristics, which align with the defining traits of lactic acid bacteria (LAB), which are rod- or coccus-shaped and catalase-negative (Das et al., 2024; Nasr & Abd, 2024 ). Table 4 reveals that two isolates (M5 and M9) were classified as hetero-fermentative LAB due to gas production, while seven isolates (M1, M2, M3, M4, M6, M7, and M8) were categorized as homo-fermentative. Homo-fermentative LAB do not generate gas, while hetero-fermentative LAB do. These findings align with Kavitha & Jeevaratnam's (2016) report on homo-fermentative Lactobacillus isolates from piper beetle leaves. Gram-positive and catalase-negative isolates' fermentation type was determined by glucose CO₂ production (Kabir et al., 2022). LABs are classified as homo- or hetero-fermentative based on carbohydrate fermentation end products (Nasr & Abd, 2024 ). All tested isolates fermented TSIA media containing glucose, lactose, and sucrose, producing acid. Positive LAB isolates are identified by a yellow color change in these sugars (Asha et al., 2024 ). The citrate test confirmed that isolates M1–M9 could not utilize citrate as a carbon source, as indicated by the absence of a color change (Table 4 ). This suggests a deficiency in citrate-catabolizing enzymes (Asha et al., 2024 ), aligning with Hafsan and Mustami (2015), despite some LAB's ability to metabolize citrate (Castro & Poveda, 2024). Additionally, isolates M1–M9 were non-motile and indole-negative (Table 4 ), indicating the absence of flagella (Prastujati et al., 2022 ) and tryptophanase, preventing tryptophan breakdown (Hafsan & Mustami, 2015). Moreover, these isolates tested MR-negative (Table 4 ), confirming the lack of mixed acid fermentation. This finding is consistent with Ngene et al. ( 2019 ) but differs from some LAB strains found in fermented foods (Sirisha et al., 2021), suggesting that these isolates are unable to produce organic acids such as lactic, succinic, acetic, and formic acid from glucose. Table 4 Biochemical Characteristics of LAB Isolate Isolates Catalase test Citrate test MR test TSIA test SIM test Fermentative type S B Gas H 2 S M I 1 (M1) − − − Y Y − − − − Homo 4 (M2) − − − Y Y − − − − Homo 5 (M3) − − − Y Y − − − − Homo 7 (M4) − − − Y Y − − − − Homo 8 (M5) − − − YR Y + + − − Hetro 11 (M6) − − − R Y − − − − Homo 14 (M7) − − − Y Y − − − − Homo 15 (M8) − − − R Y − − − − Homo 17 (M9) − − − YR Y + + − − Hetro Key : “S” slant, “B” butt, “M” motility, “I” indole, “Y” yellow, “R” red, “−” negative, “+” positive 3.4 Physiological Characteristics of LAB Isolates Most LAB isolates (77.8%, M1-M3, M5-M7, M9) tolerated all tested temperatures (Table 5 ), consistent with prior LAB temperature tolerance research (Khalil & Anwar, 2016; Terpou et al., 2019). Temperature control is vital for maintaining probiotic potential in food applications (Sionek et al., 2024). All LAB isolates (M1-M9) grew at 1% and 4% NaCl, with 88.9% (M1-M7, M9) tolerating 6% NaCl (M8 failed at 6%). This broad salt tolerance suggests potential for gastrointestinal survival, crucial for probiotic efficacy (Asha et al., 2024 ; Tsega et al., 2023 ). Genetic analysis will confirm probiotic safety (Tsega et al., 2023 ). Isolates M3 and M4 survived all tested acidic conditions (pH 2-6.5, Table 5 ), with 22% overall tolerance to this range and 88.9% tolerance to pH 4-6.5, suggesting potential gastrointestinal survival (Chen et al., 2020; Iraporda et al., 2024; Hanifa et al., 2020). Acid tolerance, crucial for probiotic efficacy given stomach pH (Russell et al., 1993; Dressman et al., 1990; Sahadeva et al., 2011; El-Beksh et al., 2021), makes M3 and M4 promising candidates in Mekele, Tigray, Ethiopia. All isolates tolerated 0.4% phenol (Table 5 ), while 78% (M2-M3, M5-M9) resisted 0.6% (M1 and M4 failed). This phenol resistance, important due to inhibitory effects on LAB (Nandha & Shukla, 2023) and gut phenol presence, further supports their probiotic potential in the local context. Table 5 Physiological Characteristics of LAB Isolates Physiological characteristics Isolates M1 M2 M3 M4 M5 M6 M7 M8 M9 Temperature (℃) 15 + + + + + + + + + 27 + + + + + + + + + 37 + + + + + + + + + 45 + + + − + + + − + NaCl (%) 1 + + + + + + + + + 4 + + + + + + + + + 6 + + + + + + + − + pH level 2 − − + + − − − − − 4 + + + + + + − + + 6 + + + + + + + + + 6.5 + + + + + + + + + Phenol conc (%) 0.4 + + + + + + + + + 0.6 − + + − + + + + + Key : “+” indicates bacterial growth, while “−” indicates no bacterial growth 3.5 Safety Evaluation of LAB Isolates Hemolysis testing confirmed the safety of LAB isolates M1–M9, as they exhibited γ-hemolysis (non-hemolytic), unlike the β-hemolytic pathogenic controls (Table 6 ) (Nandha & Shukla, 2023; Meradji et al., 2023 ). The absence of hemolysin, likely due to the lack of cytolysin genes (Meradji et al., 2023 ), indicates non-virulence (Tsega et al., 2023 ). Given that hemolysis testing is a crucial criterion for probiotic selection, these isolates hold promising potential for applications in dairy, non-dairy, and medicinal products, pending further analysis. Table 6 Hemolysis Effect Hemolytic Effect Isolates Controls M1 M2 M3 M4 M5 M6 M7 M8 M9 E. coli S. aureus S. typhimurium α − − − − − − − − − − − − β − − − − − − − − − + + + γ + + + + + + + + + − − − Key : (−) indicates no hemolytic effect, (+) there is hemolytic effect 3.5.1 Antibacterial Activity Test In this study, all tested isolates (M1-M9) showed no zone of inhibition (resistance) against E. coli ATCC 25922 (a) (Table 7 ). This suggests these isolates lack the ability to inhibit E. coli growth, possibly due to their co-existence without antagonism in milk. Against S. aureus ATCC 25923 (b), four isolates (M2, M4, M7, and M9) showed no inhibition, four (M3, M5, M6, and M8) showed intermediate inhibition, and only M1 showed sensitivity. Against P. aeruginosa ATCC 27853 (c), six isolates (M1, M2, M3, M4, M7, and M9) showed no inhibition, M5 showed intermediate inhibition, and M6 and M8 showed sensitivity. These results demonstrate varying degrees of inhibition against selected indicator organisms, consistent with Akabanda et al., ( 2014 ). Consequently, no isolate inhibited E. coli ATCC 25922, only M1 inhibited S. aureus ATCC 25923, and only M6 and M8 inhibited P. aeruginosa ATCC 27853. Rattanachaikunsopon & Phumkhachorn (2010) suggest that LAB antimicrobial activity stems from organic acids and other compounds like ethanol, H 2 O 2 , diacetyl, reuterin, and bacteriocins. Akabanda et al. ( 2014 ) also demonstrated that LAB antimicrobial compounds can inhibit pathogenic bacteria growth. Table 7 Antibacterial Test Antibacterial effect Isolates M1 M2 M3 M4 M5 M6 M7 M8 M9 S. aureus (ATCC 25923) (11)S R (7)I R (6)I (4)I R (3)I R E. coli (ATCC 25922) R R R R R R R R R P. aeruginosa (ATCC 27853) R R R R (6)I (9)S R (13)S R Key Results mean ± SD from triplicate experiment. Findings were recorded as no detectable inhibition or resistance (R), weak inhibition zone ( 8–12mm) (Akabanda et al., 2014 ). 3.5.2 Antibiotic Susceptibility Test Isolate M1 showed varied antibiotic sensitivity. Isolates M2-M4 and M8 were resistant to all tested antibiotics. Isolates M5, M6, and M7 exhibited similar resistance patterns (clindamycin, ceftriaxone, amikacin, tetracycline), with M6 additionally resistant to erythromycin. Isolate M9 displayed broad resistance, with susceptibility only to penicillin and amikacin. These results align with previous findings: LAB sensitivity to ampicillin (Jadhav & Narayanaswamy, 2024 ), and varying sensitivity to ampicillin, erythromycin, tetracycline, and clindamycin (Ramu et al ., 2022). Clindamycin was the most prevalent resistance, followed by ceftriaxone, amikacin, and tetracycline. This is consistent with amikacin resistance reported by Szutowska & Gwiazdowska (2021), but differs from reports of kanamycin and vancomycin resistance (Marchwińska & Gwiazdowska, 2022). Ciprofloxacin resistance was also noted (Jadhav & Narayanaswamy, 2024 ). Antibiotic resistance may stem from genetic factors (Okoye et al., 2023 ). While antibiotic sensitive is desirable in probiotics, genetic characterization is crucial for safety assurance. Table 8 Antibiotic Susceptibility Test of Isolates Isolates Inhibition Zone of Each Antibiotics (mm) Cln Me E Amp Cro P A Van Cip Te Cot Liv M1 (6) R (21) S (13) R (29) S (18) I (27) S (15) R (18) I (23) S (6) R (16) I (22)S M2 R R R R R R R R R R R R M3 R R R R R R R R R R R R M4 R R R R R R R R R R R R M5 (10) R (22) S (22) S (32) S (6) R (29) S (6) R (19) I (23) S (6) R (23) I (24)S M6 (6) R (21) S (6) R (29) S (18) I (30) S (10)R (19) I (25) S (27)S (26) S (22)S M7 (8) R (23) S (22) S (30) S (6) R (30) S (30) R (20) I (25) S (6) R (16) I (24)S M8 R R R R R R R R R R R R M9 (13) R R R R R (31) S (21) S R R R (17) I R Key : Cln-Clindamycin, Me-Meropinium, E-Erythromycin, Amp-Ampicillin, Cro-Ceftriaxone, P-Penicillin G, A-Amikacin, Van-Vancomycin, Cip-Ciprofloxacin, Te-Tetracycline, Cot-Cotrimoxazole, Liv-Levofloxacillin, R-Resistance, I-Intermediate susceptible, S-Susceptible 3.6 Evaluation of Potent LAB as a Starter Culture 3.6.1 Formation of Yogurt In this study, 67% of LAB isolates (M1, M2, M3, M7, M8, and M9) successfully fermented pasteurized milk into yogurt, while 33% (M4, M5, and M6) failed, likely due to lactase deficiency. The positive control (Y7) fermented, whereas the negative control (Y8) remained raw. The six yogurt-producing isolates show promise as starter cultures, pending further research. LAB plays a vital role in milk fermentation, typically involving Streptococcus thermophilus and Lactobacillus bulgaricus , and has broad applications in dairy, food, and feed industries (Widyastuti & Febrisiantosa, 2014). 3.6.2 Physiochemical Properties of Yogurt As shown in Table 10 , M8 exhibited the highest pH, while M3 had the lowest. M7's pH of 4.6 aligns with Lee & Lucey ( 2010 ). Iranian yogurt pH ranges from 4.06 to 4.64 (Nemati et al ., 2023). Yogurt consumption benefits lactose-intolerant individuals due to lactose conversion to lactic acid during fermentation (Ademosun et al., 2019 ). During milk acidification, pH decreases from 6.7 to ≤ 4.6 (Lee & Lucey, 2010 ). The low pH in all yogurt products is significant, as it inhibits most pathogenic microorganisms (Madora et al ., 2019). However, excessively low pH can also hinder beneficial microorganisms, necessitating optimal pH maintenance. M2, M7, and M8 showed the highest freezing points (FP), while M9 and Y8 had the lowest. FP, indicating the solidification point, is influenced by sugars, additives, and starter cultures. Pasteurization-induced changes in calcium phosphate complexes and CO 2 pressure can also affect FP. M8 had the highest density, and M3 the lowest, demonstrating the starter culture's impact on yogurt density. Y8 had the highest titratable acidity (TA), and M8 the lowest. Iranian yogurt TA ranges from 0.91 to 1.57% (Nemati et al ., 2023). TA variations are attributed to starter culture differences. Increased TA is crucial for inhibiting undesirable microbial growth and ensuring proper fermentation (Madora et al ., 2019). Table 9 Physiochemical Properties of Yogurt Fermented by the Isolates Sample Physiochemical Properties pH FP TA D M1 4.4 ± 0.02 0.3 ± 0.02 5.4 ± 0.02 40.4 ± 0.03 M2 4.4 ± 0.01 0.5 ± 0.02 4.1 ± 0.12 30.6 ± 0.01 M3 4.3 ± 0.02 0.2 ± 0.02 5.7 ± 0.02 10.0 ± 0.10 M7 4.6 ± 0.02 0.5 ± 0.01 4.4 ± 0.01 40.1 ± 0.04 M8 4.8 ± 0.03 0.5 ± 0.02 3.6 ± 0.02 44.8 ± 0.02 M9 4.5 ± 0.01 -0.5 ± 0.01 5.1 ± 0.13 27.2 ± 0.04 Y7 ( + control) 4.5 ± 0.01 0.4 ± 0.02 4.8 ± 0.01 29.1 ± 0.03 Y8 (-control) 6.4 ± 0.02 -0.5 ± 0.01 7.2 ± 0.02 27.5 ± 0.03 Key : Values were given mean ± SD from triplicate experiment. Y-yogurt, D-density, FP-freezing point, TA-titratable acidity 3.6.3 C ompositions of Yogurt As shown in Table 10 , M8 had the highest protein content, while M3 had the lowest. Previous reports on cow milk yogurt protein ranged from 2.56 to 4.07 (Ibrahim et al ., 2021; Wulansari & Kusmayadi, 2016; Elnour et al ., 2022). This variability is attributed to the bacterial strains' ability to break down milk protein. M8 also exhibited the highest fat content and M9 the lowest. Fat content in previous studies ranged from 3.61 to 4.51 (Wulansari & Kusmayadi, 2016; Elnour et al ., 2022). Variations in fat are due to starter culture differences and milk composition. M8 had the highest lactose content and M3 the lowest. Elnour et al . (2022) reported yogurt lactose at 3.20, with differences attributed to starter culture activity (Ayivi et al ., 2020). M3 had the highest salt content and M1 the lowest. Starter cultures indirectly affect salt perception through flavor compound production. Y8 had the highest ash content and M9 the lowest, aligning with Olugbuyiro & Oseh ( 2011 ). Ash content in previous reports ranged from 0.39 to 0.70 (Elnour et al ., 2022; Ibrahim et al ., 2021). Ash represents the inorganic residue after incineration. M9 had the highest moisture content, and Y7 the lowest, consistent with reports of 87.27% to 87.77% moisture (Wulansari & Kusmayadi, 2016; Ibrahim et al ., 2021). Yogurt composition is influenced by various factors, including carbohydrate (lactose) conversion to lactic acid during fermentation, resulting in lower carbohydrate levels (Elnour et al ., 2022; Ademosun et al., 2019 ). Table 10 Compositions of Yogurts Fermented by the LAB Isolates Sample Composition in Percent (%) P L F A M S M1 4.6 ± 0.02 2.4 ± 0.02 9.4 ± 0.02 0.8 ± 0.01 88.4 ± 0.02 0.3 ± 0.02 M2 3.1 ± 0.06 4.7 ± 0.02 2.1 ± 0.05 0.4 ± 0.01 87.2 ± 0.15 1.0 ± 0.05 M3 1.4 ± 0.02 2.1 ± 0.06 5.2 ± 0.02 0.8 ± 0.02 87.7 ± 0.15 1.2 ± 0.06 M7 4.6 ± 0.02 6.7 ± 0.03 9.5 ± 0.01 0.6 ± 0.03 89.1 ± 0.11 0.6 ± 0.02 M8 5.1 ± 0.05 7.7 ± 0.02 10.1 ± 0.03 0.6 ± 0.02 88.7 ± 0.01 0.7 ± 0.01 M9 3.1 ± 0.58 4.2 ± 0.02 1.8 ± 0.01 0.1 ± 0.02 89.3 ± 0.01 0.7 ± 0.02 + ve control 3.0 ± 0.02 4.5 ± 0.01 2.0 ± 0.03 0.6 ± 0.01 86.9 ± 0.02 1.1 ± 0.06 -ve control 2.9 ± 0.02 4.4 ± 0.02 3.4 ± 0.03 2.4 ± 0.01 87.6 ± 0.02 1.0 ± 0.02 Key : Values were given mean ± SD from triplicate experiment. M-isolate, Y-yogurt P-protein, L-lactose, F-fat, A-ash, M-moisture, S-salt 3.7 Sensory Evaluation of Yogurt As shown in Table 11 , Y7's color was rated "like moderately" and was well-accepted, while Y8's color was "dislike extremely" and rejected. M7's odor was rated "like extremely" and was highly accepted. M1, M9, M3, M2, and M8's odors were rated "like moderately," compared to Y7. Y8's odor was unpleasant and rated "dislike extremely." M1's taste was rated "like extremely," while M8 was rated "like slightly." Y8's taste was unpleasant and rejected. M1, M2, M9, and Y7 had good shape/size, resembling normal yogurt. M3, M7, and M8 had slightly less ideal shapes/sizes. Y8 had an abnormal shape/size and was rated "dislike moderately. Overall acceptability (OA) was also evaluated. M1, M3, M8, and M9 were rated "like moderately," and Y7 was rated "like slightly." Y8 was rated "dislike extremely" in OA, reflecting its poor sensory scores due to the absence of a starter culture. LAB isolates significantly influence aroma, quality, and consistency through varying fermentation potentials (Zapasnik et al ., 2022). Sensory scores are closely linked to free fatty acid concentration and amino acid degradation in products like yogurt. They also play a crucial role in host health (Fijan, 2014). Table 11 Sensory Scores of the Yogurt Sample Sensory scores Color Odor Test Shape/size Overall acceptability M1 5.8 ± 0.28 6.5 ± 0.35 6.8 ± 0.14 6.2 ± 0.50 6.3 ± 0.35 M2 5.2 ± 0.21 5.9 ± 0.21 5.2 ± 0.28 6.2 ± 0.28 5.8 ± 0.28 M3 4.3 ± 0.35 6.2 ± 0.42 5.1 ± 0.28 5.4 ± 0.50 6.1 ± 0.14 M7 5.9 ± 0.21 6.8 ± 0.35 6.0 ± 0.28 4.8 ± 0.21 5.3 ± 0.42 M8 4.9 ± 0.21 5.2 ± 0.28 4.9 ± 0.21 5.2 ± 0.21 6.0 ± 0.07 M9 5.4 ± 0.50 6.4 ± 0.14 5.3 ± 0.42 6.2 ± 0.14 6.0 ± 0.28 + ve control 6.2 ± 0.28 4.4 ± 0.14 6.3 ± 0.35 5.5 ± 0.35 5.2 ± 0.28 -ve control) 1.1 ± 0.14 1.5 ± 0.71 1.3 ± 0.35 2.2 ± 0.21 1.3 ± 0.42 Key M-isolate, Y-yogurt, 7 = like extremely, 6 = like moderately, 5 = like slightly, 4 = neither like nor dislike, 3 = dislike slightly, 2 = dislike moderately and 1 = dislike extremely). Values mean ± SD from duplicate experiment. 3.8 Identification of LAB Probiotic at Genus level The LAB isolates in this study were tentatively grouped under the genera Lactobacillus , Enterococcus , and Leuconostoc . This classification was primarily based on morphology (rods, cocci, tetrads), glucose fermentation mode (homo- or hetero-fermentative), growth at 15°C and 45°C, and salt tolerance. Yelnetty et al., ( 2014 ) reported that Lactobacillus was the dominant genus in fermented cow and lamb milk. LAB is typically gram-positive, catalase-negative, non-motile, non-spore-forming rods or cocci, and can be homo- or hetero- fermentative. Lactobacillus species exhibit rod-shaped cells, ranging from short to long, with some species displaying coccobacilli. Lactobacillus and Enterococcus share similar characteristics, differing mainly in cell shape: rods for Lactobacillus and cocci for Enterococcus (Ismail et al., 2018 ). Leuconostoc species are hetero-fermentative cocci. 3.9 Conclusion This study analyzed cow milk composition in Mekele, Ethiopia, identifying and characterizing potential lactic acid bacteria (LAB) for probiotic and yogurt-producing applications. Milk samples showed significant compositional and physicochemical variations, influenced by genetic, environmental, and feed factors. Nine Gram-positive, catalase-negative LAB isolates (M1-M9) were selected from 23 due to their tolerance to temperature, salt, pH, and phenol, and their non-hemolytic activity, indicating safety. Antibacterial and antibiotic susceptibility tests revealed diverse inhibitory effects and resistance patterns. Six isolates successfully fermented pasteurized milk into yogurt, with M1, M3, M8, and M9 receiving favorable sensory evaluations. Biochemical tests classified seven isolates as homo-fermentative and two as hetero-fermentative, with preliminary identification linking them to Lactobacillus, Enterococcus , and Leuconostoc , common in Ethiopian fermented dairy. The study highlights promising LAB isolates for yogurt production, recommending genetic analysis to confirm safety, probiotic potential, and antibiotic resistance, alongside in vivo studies for efficacy. Standardized starter cultures are essential for industrial fermentation, with future research needed on probiotic mechanisms and their impact on gut microbiota and immune function. Declarations Ethics approval and consent to participate This study was approved by the Mekelle University Institutional Review Board (Approve No. MU-IRB 2305/2024). All methods were conducted according to relevant guidelines and regulations. The study was carried out ethically in accordance with the World Medical Association Declaration of Helsinki (2024 version). Written informed consent was also obtained from zonal and district administrations of dairy farms and milk collection to inform all participants in the research. Informed consent to participate was obtained from all participants in the study. Consent for publication Not applicable. Competing interest The authors declare no competing interests. Funding This work was supported by Mekele University (Ethiopia), Hawassa University (Ethiopia), Norwegian University of Life Sciences (Norway) collaboration program phase V (Grant No. MU-HU-NMBU RADO/EXTERNAL/SS003/2024). Authors’ contributions GS wrote and develop the manuscript. GG provided constructive comments. GS and GG edited and finalized the manuscript. DB reviewed the manuscript. All authors strongly contribute to enhance the quality of manuscript. Finally, all authors read and approved the final manuscript. Acknowledgements The authors thank the Mekelle University for granting M.Sc. to Goiteom Senay Niguse and thank the Scholars Rescue Fund of the International Institute of Education (SRF-IIF) for granting a fellowship to Dr Desta Berhe Sbhatu and the UiTM for hosting him during the preparation of this manuscript. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8906693","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":593139547,"identity":"89f6d5ae-f607-4a33-8ff2-9c53e7af8417","order_by":0,"name":"Goiteom Senay Niguse","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABDUlEQVRIiWNgGAWjYFACxgYEO6GiRg5EH3iAX0sjQs+DM8eMwVoSiLWG8WEbcyKYh0+LvHtz+4MPFdvs+aWbn0kksLGlzw87/BBoi52cbgN2LYZnDjY2zjhzO3HmnGNmEgk8Mrkbb6cZALUkG5sdwKFlRmJjM2/b7QSDGwlALRJsuRtnJ4C0HEjchk/L37bb9vY30r9JJBgwpxvOTv+AV4u8BFALY9ttxg0SOUBbEpgT5KVz8NtiwHOwcWYP0C8zbuQUWyQcOGa4QTqn4ECCAW6/yLcDA+xHxW17/hnpG2/+/FcjLz87ffOHDxV2cri0GCCJs0ggRAywKwfb0oBgM39AFxkFo2AUjIJRAAIAbRxrxQnDnRIAAAAASUVORK5CYII=","orcid":"","institution":"Mekelle University","correspondingAuthor":true,"prefix":"","firstName":"Goiteom","middleName":"Senay","lastName":"Niguse","suffix":""},{"id":593139548,"identity":"63b8a603-4c96-44a1-b98b-fd71f6f43f67","order_by":1,"name":"Desta Berhe Sbhatu","email":"","orcid":"","institution":"Mekelle University","correspondingAuthor":false,"prefix":"","firstName":"Desta","middleName":"Berhe","lastName":"Sbhatu","suffix":""},{"id":593139549,"identity":"d71ea08d-f601-42d1-8fcf-09d8d84f381c","order_by":2,"name":"Gerbreselema Gebreyohannes","email":"","orcid":"","institution":"Mekelle University","correspondingAuthor":false,"prefix":"","firstName":"Gerbreselema","middleName":"","lastName":"Gebreyohannes","suffix":""}],"badges":[],"createdAt":"2026-02-18 07:11:03","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":true,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":true},"doi":"10.21203/rs.3.rs-8906693/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8906693/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003e\u003cstrong\u003eIsolation and Characterization of Lactic Acid Bacteria from Raw Cow Milk and Evaluation of Their Probiotic Potential\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"1. INTRODUCTION","content":"\u003cp\u003eMilk, a nutrient-dense secretion from mammary glands composed of water, fat, proteins, lactose, minerals, and vitamins, is essential for human nutrition. In Ethiopia, cattle contribute the largest portion of annual milk production (81.2%), followed by goats (7.9%), camels (6.3%), and sheep (4.6%) (Bereda et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Milk and its derivatives are vital for human health and nutrition and also provide a conducive environment for the proliferation of lactic acid bacteria (LAB), which have been integral to food fermentation and preservation for centuries (Hatti-kaul et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eLAB's metabolic capability allows them to convert carbohydrate substrates into organic acids, predominantly lactic acid (LA), while also producing diverse metabolites with numerous health benefits (Rodriguez \u003cem\u003eet al\u003c/em\u003e., 2019). Historically, LAB have served as starter cultures for the fermentation of a wide variety of foods, including milk, vegetables, meat, fish, cereals, and animal feed, contributing to flavor, aroma, and extended shelf life, thus making them invaluable in food production (Kunene et al., \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Notably, fermented dairy products, valued at over 80\u0026nbsp;billion euros in 2011, held a particularly significant position in the global market (Hatti-kaul et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eLABs are used in food fermentation and functional starter cultures to produce beneficial compounds like Extracellular Polymeric Substance (EPS), organic acids, polyols, aromatic compounds, and bacteriocins, which improve food product texture, aroma, flavor, health benefits, and shelf life (Aritonang et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). LAB-derived lactic acid, known for its bio-preservative and antimicrobial properties, is highly valuable in various industries like food, cosmetics, pharmaceuticals, and chemicals. Its demand is driven by its role in poly-lactic acid production, biodegradable thermoplastic polyester used in packaging materials, films, fibers, and medical implants. Selected LAB serve as probiotics, enhancing human health by improving intestinal health, enhancing immune response, reducing cholesterol, and mitigating cancer risk. They also contribute to disease prevention, allergic response regulation, and gastrointestinal health maintenance (Fijan, 2014; Somashekaraiah \u003cem\u003eet al\u003c/em\u003e., 2019; Aritonang et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe global market for LAB, valued at USD 2.08\u0026nbsp;billion in 2016, is expected to grow due to increasing demand in packaging, personal care, and textiles, driven by the isolation and characterization of LAB from traditional raw and fermented foods (Hatti-kaul et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). LAB, found in traditional foods like milk, is uncharacterized and requires further investigation. Recognized as \"Generally Regarded as Safe\" (GRAS), LAB produce enzymes that break down and synthesize food components like carbohydrates, proteins, and lipids, essential for survival and widely used in food processing and industrial applications (Amenu \u0026amp; Bacha, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2023\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eLAB are essential for fermented foods, providing benefits such as improved gut health, enhanced immune response, and potential anti-cancer properties. However, the growing challenge of antibiotic resistance has highlighted the need for probiotics, particularly LAB, to promote livestock health. Despite their potential, the identification and characterization of LAB in milk and dairy products remain insufficient due to environmental variables, dairy management practices, and microbial interactions. This study aims to characterize and evaluate LAB isolates from raw cow milk to expand the understanding of their application in dairy fermentation and probiotic development.\u003c/p\u003e"},{"header":"2. MATERIALS AND METHODS","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Milk Sampling and Physicochemical \u0026amp; Compositional Analysis\u003c/h2\u003e \u003cp\u003eCow milk samples were collected from producers and sellers in the sub-cities of Mekelle using sterilized, sealed bottles. The samples were stored in an ice box and transported to the microbiology laboratory at Mekelle University, where they were kept at 4\u0026deg;C until analysis. The physicochemical properties (pH, density, freezing point, and moisture content) and compositional attributes (ash content, protein, fat, lactose, and titratable acidity) were analyzed using a Lacto-Scan milk analyzer (Olugbuyiro \u0026amp; Oseh, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e and Sharma et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Isolation and Morphological Characterization of LAB\u003c/h2\u003e \u003cp\u003eLAB isolation followed the method of Ismail1\u003cem\u003eet al\u003c/em\u003e., (2018), starting with serial dilutions of the samples. A 0.1 ml aliquot was spread onto de Man, Rogosa, and Sharpe (MRS) agar plates, which were incubated at 37\u0026deg;C under aerobic conditions for 48\u0026ndash;72 hours (Taye et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Morphological characterization of the isolates involved observing colony and cell morphology (shape, size, and color) under a microscope (Lingga et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Gram staining of pure cultures was performed by applying a bacterial culture to a heat-fixed slide, followed by sequential treatment with crystal violet (1 minute, rinsed), iodine solution (1 minute, rinsed), 95% alcohol (30 seconds, rinsed), and safranin (30 seconds, rinsed). After a final rinse, the slide was observed under a 100x objective lens, with Gram-positive bacteria appearing purple and Gram-negative bacteria appearing red.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Biochemical Characterization and Identification of LAB Isolates\u003c/h2\u003e \u003cp\u003eThe LAB's biochemical characterization involved tests to assess their metabolic capabilities, with the catalase test indicating the presence of the enzyme by producing gas bubbles when 3% hydrogen peroxide was added to a pure isolate (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). To assess their fermentative pathways, the gas production test utilized nutrient broth with an inverted Durham tube, where incubation at 37\u0026deg;C for 48 hours allowed for the differentiation of homo-fermentative LAB, which produce no gas, from hetero-fermentative LAB, which generate gas trapped in the Durham tube (Prastujati et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Further carbohydrate fermentation was evaluated using Triple Sugar Iron Agar (TSIA), with a yellow color change after 24 hours at 37\u0026deg;C signifying the fermentation of glucose, sucrose, and/or lactose, and the absence of blackening indicating no hydrogen sulfide production (Tigistu et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). The ability of the isolates to utilize citrate as a sole carbon source was tested using Simmons citrate medium, where a color change from green to blue or yellow after 24 hours at 37\u0026deg;C indicated a positive result (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eAdditional biochemical characteristics were assessed using SIM and MR tests. Motility was determined in SIM media by observing the spreading of bacterial colonies after 24 hours of incubation at 37\u0026deg;C. The production of indole was detected in the same incubated SIM media by the addition of Kovac's reagent, with the formation of a red layer at the top indicating a positive result (Ngene et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Finally, the Methyl Red (MR) test, performed on isolates grown in MR-VP media for 24 hours at 37\u0026deg;C, identified acid producers by a change in the medium's color to red upon the addition of the MR indicator (Ngene et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Physiological Characterization of LAB isolates\u003c/h2\u003e \u003cp\u003eThe physiological characteristics of the LAB isolates were evaluated through several tolerance tests. Temperature resistance was assessed by culturing the isolates in MRS broth and observing their growth after 48 hours of incubation at 15\u0026deg;C, 27\u0026deg;C, 37\u0026deg;C, and 45\u0026deg;C (Lingga et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Sodium chloride (NaCl) tolerance was determined by initially culturing the isolates in MRS broth for 24 hours, followed by subsequent 24-hour incubation in MRS broth supplemented with 1%, 4%, and 6% NaCl. These cultures were then plated on MRS agar and incubated for 48 hours, after which colony growth was observed and recorded (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). To test survival in acidic conditions, the isolates were incubated in MRS broth at pH levels of 6.5, 6, 4, and 2 for 24 hours, then streaked onto MRS agar and incubated at 37\u0026deg;C for 48 hours to monitor colony growth (Safi et al., 2023). Finally, phenol resistance was assessed by culturing the LAB isolates in MRS broth for 24 hours and then transferring them onto MRS agar supplemented with 0.4% and 0.6% phenol solution, with growth observed after 48 hours of incubation at 37\u0026deg;C.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5 Safety Evaluation of LAB isolates\u003c/h2\u003e \u003cp\u003eThe safety evaluation of LAB isolates involved hemolysis, antibacterial activity, and antibiotic susceptibility tests. The hemolysis test was conducted by growing LAB colonies on blood agar supplemented with sheep blood and incubating them at 37\u0026deg;C for 24 hours (Amenu \u0026amp; Bacha, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Hemolytic activity was assessed by holding the plate up to a light source, with control strains including \u003cem\u003eE. coli\u003c/em\u003e (ATCC 29922), \u003cem\u003eS. aureus\u003c/em\u003e (ATCC 25923), and \u003cem\u003eS. typhimurium\u003c/em\u003e (ATCC 13311) Meradji et al., (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Antibacterial activity was tested against \u003cem\u003eS. aureus\u003c/em\u003e (ATCC 25923), \u003cem\u003eE. coli\u003c/em\u003e (ATCC 25922), and \u003cem\u003eP. aeruginosa\u003c/em\u003e (ATCC 27853) using the agar well diffusion method (Amenu \u0026amp; Bacha, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Inhibition zones around the wells were categorized as resistant (R) (no zone), weak inhibition (\u0026lt;\u0026thinsp;1\u0026ndash;4 mm), moderate inhibition (4\u0026ndash;8 mm) (intermediate), or strong inhibition (\u0026gt;\u0026thinsp;8\u0026ndash;12 mm) (sensitive) (Akabanda et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Additionally, the antibiotic susceptibility of the isolates to penicillin G (10 \u0026micro;g), ampicillin (10 \u0026micro;g), cotrimoxazole (25 \u0026micro;g), vancomycin (30 \u0026micro;g), meropenem (10 \u0026micro;g), tetracycline (30 \u0026micro;g), ceftriaxone (30 \u0026micro;g), clindamycin (2 \u0026micro;g), ciprofloxacin (5 \u0026micro;g), erythromycin (15 \u0026micro;g), levofloxacin (5 \u0026micro;g), and amikacin (10 \u0026micro;g) was determined using the agar disc diffusion method (Bauer et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1966\u003c/span\u003e), with inhibition zones categorized as susceptible (S) (\u0026ge;\u0026thinsp;21 mm), intermediate (I) (16\u0026ndash;20 mm), or resistant (R) (\u0026le;\u0026thinsp;15 mm) (Bauer et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1966\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.6 Evaluation of Potent isolate as a Starter Culture\u003c/h2\u003e \u003cp\u003eThe evaluation of potent LAB isolates as a starter culture involved the formation of fermented milk (yogurt). Following (Ismail et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), fresh raw cow milk was procured from a local farm in Mekelle. The milk was pasteurized at 80\u0026ndash;90\u0026deg;C for 15 minutes to eliminate existing microbes, then cooled to approximately 40\u0026deg;C (Lee \u0026amp; Lucey, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). It was then inoculated with one colony or 1% of the LAB strain, ensuring a cell count of 1 \u0026times; 10⁶, and poured into sterilized bottles (Sharma et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). A total of 550 mL of milk was used for yogurt formation, with 50 mL allocated per sample. Fermentation occurred at room temperature for 20\u0026ndash;24 hours (Ismail1\u003cem\u003eet al\u003c/em\u003e., 2018).\u003c/p\u003e \u003cp\u003eThe physiochemical properties of yogurt, including pH, freezing point, titratable acidity, and density, were analyzed using a Lacto-Scan milk analyzer (Olugbuyiro \u0026amp; Oseh, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Sharma et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The analyzer was cleaned using \u003cem\u003eLacto Daily\u003c/em\u003e and \u003cem\u003eLacto Weekly\u003c/em\u003e solutions and rinsed three times with distilled water before analysis. A 25 mL sample was placed in a sample holder, activated via the start button, and measured, with results displayed digitally. The composition of yogurt, including ash content, protein, lactose, fat, and moisture, was also assessed using the Lacto-Scan milk analyzer (Olugbuyiro \u0026amp; Oseh, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Sharma et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.7 Sensory Evaluation of Yogurt\u003c/h2\u003e \u003cp\u003eSensory evaluation of yogurt was conducted following a previous method (Ademosun et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Twelve trained panelists\u0026mdash;six women and six men, aged 22\u0026ndash;45\u0026mdash;participated in the evaluation, including undergraduate and graduate students, as well as teaching and non-teaching staff members of Mekelle University. All panelists were familiar with conventional natural cow milk yogurt. The yogurt samples were assessed for organoleptic characteristics and overall acceptability using a 7-point hedonic scale: like extremely (7), like moderately (6), like slightly (5), neither like nor dislike (4), dislike slightly (3), dislike moderately (2), and dislike extremely (1). Sensory properties were evaluated based on fermented color, fermented odor, and taste quality. To minimize bias, each sample group was coded before being served as cooled yogurt to the panelists. The results were generated based on two trials and recorded using a table with mean values.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e2.8 Identification of LAB at Genus Level\u003c/h2\u003e \u003cp\u003eThe identification and characterization of LAB isolates relied on their morphological, biochemical, and physiological properties. Morphological analysis included observing colony shape, size, and color. Biochemical tests assessed metabolic activities such as catalase production and carbohydrate fermentation profiles. Physiological characterization evaluated their adaptability and functional potential by determining their resistance to various temperatures, tolerance to sodium chloride, survival in acidic conditions, and resistance to phenol (Yelnetty et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Hafsan and Mustami, 2015).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e2.9 Data Analysis\u003c/h2\u003e \u003cp\u003eData were recorded using Microsoft Excel, and differences between the means were determined through one-way analysis of variance (ANOVA). Quantitative data were further analyzed using the Fisher Least Significant Difference (LSD) method with Minitab software (Version 19). Statistical significance was established at a P-value of less than 0.05.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. RESULTS AND DISCUSSION","content":"\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Physicochemical Properties and Composition of Raw Cow Milk\u003c/h2\u003e \u003cp\u003eThe study found significant variations in protein and lactose content in milk samples, with highest protein levels in S2 and lowest in S3. These average values differ from previous studies but fall within the ranges of other studies, likely influenced by factors like forage quality (Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Meskel \u0026amp; Sebho, 2018). The study found that lactose content in S5 was highest, while in S1 and S4, it was lower, indicating the significant impact of environmental conditions and animal breeding, with crossbred animals showing higher lactose levels (Elnour \u003cem\u003eet al\u003c/em\u003e., 2022; Ibrahim \u003cem\u003eet al\u003c/em\u003e., 2021), Ketema \u003cem\u003eet al\u003c/em\u003e. (2018).\u003c/p\u003e \u003cp\u003eThe physical and chemical properties of the milk samples exhibited notable variations. Density was highest in S5 and lowest in S3 (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e3\u003c/span\u003e), aligning with previous findings linking it to fat and solids-not-fat content (Meskel \u0026amp; Sebho, 2018). In contrast, the freezing point (FP) varied, with S2 displaying the highest value, while S3, S4, and S6 had the lowest (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e3\u003c/span\u003e), a finding that underscores the influence of soluble milk constituents, particularly lactose and minerals, and contradicts Asefa \u0026amp; Teshome (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe pH values of the milk ranged from the highest in S6 to the lowest in S4 (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e3\u003c/span\u003e), consistent with previously reported ranges (Elnour et al., 2022; Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Salt content was most elevated in S7, with S1, S2, and S4 exhibiting the lowest levels (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e2\u003c/span\u003e), a finding influenced by genetic factors, feed composition, and environmental conditions. Likewise, fat content was highest in S2 and lowest in S1 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e2\u003c/span\u003e), and these variations align with findings from other studies (Zebib et al., 2023; Ibrahim et al., 2021; Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Meskel \u0026amp; Sebho, 2018), underscoring the significant impact of feed, genetics, breed, and lactation stage (Ketema et al., 2018).\u003c/p\u003e \u003cp\u003eTitratable acidity (T.A), ash, and moisture content varied across the milk samples. T.A, a key indicator of freshness, peaked in S7 and was lowest in S6 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e2\u003c/span\u003e), aligning with the range reported by Asefa \u0026amp; Teshome (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Ash content, reflecting mineral composition, was highest in S7 and lowest in S1, S2, and S4 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e2\u003c/span\u003e), showing discrepancies with previous findings (Elnour et al., 2022; Asefa \u0026amp; Teshome, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), likely due to variations in mineral matter, as noted by Zebib et al. (2023). Moisture content, influenced by animal species and feed, was highest in S1 and S4, while S7 had the lowest value (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e2\u003c/span\u003e), aligning with reported findings (Ibrahim et al., 2021; Elnour et al., 2022).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePhysiochemical properties and Compositional Analysis of Milk Samples\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"11\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" 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\u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSamples\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"6\" nameend=\"c7\" namest=\"c2\"\u003e \u003cp\u003eComposition (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c11\" namest=\"c8\"\u003e \u003cp\u003ePhysiochemical properties\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eP\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eA\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eT.A\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003epH\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eM\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eD\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003eF.P\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e3.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e6.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e87.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e27.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e3.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e3.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e5.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e86.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e27.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e6.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e85.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e25.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.83\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e3.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e3.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e5.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e87.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e26.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e6.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e84.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e27.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e3.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.29\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e6.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e86.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e26.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c8\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c9\"\u003e \u003cp\u003e83.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c10\"\u003e \u003cp\u003e26.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.46\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c11\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"11\"\u003e\u003cb\u003eKey\u003c/b\u003e: Values were given mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD from triplicate experiment. S-sample, P-protein, L-lactose, F-fat, A- ash, S-salt \u0026amp; T.A- Taitrable acidity, D-density, F.P-freezing point and M-moisture\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Morphological Characterization and Identification of LAB\u003c/h2\u003e \u003cp\u003eThe study identified 23 pure bacterial isolates, with 78% showing gram-positive patterns and 22% showing gram-negative ones. Microscopic examination identified them as cocci (35%), rods or bacilli (26%), and coccobacilli (39%). Cocci and coccobacilli were observed in similar percentages within the gram-positive group (Hafsan \u0026amp; Mustami, 2015).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eCultural and Microscopic Characteristics of LAB Isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eG-staining\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMorphology\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eColor\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRed\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRed\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRed\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRed\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCoccibacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRed\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCocci\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIsolate 23\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBacillus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurple\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Biochemical Characterization and Identification of LAB\u003c/h2\u003e \u003cp\u003eThe study found that out of the total bacterial isolates, only nine isolates were catalase-negative, indicating that 61% were catalase-positive and 39% were catalase-negative. Therefore, only Gram-positive isolates without gas bubble formation were selected for further investigation (Kabir et al., 2022; Nandha \u0026amp; Shukla, 2023; Goicochea-Vargas et al., 2024). Nine selected isolates, numbered M1 through M9, were identified based on their Gram-positive, catalase-negative characteristics, which align with the defining traits of lactic acid bacteria (LAB), which are rod- or coccus-shaped and catalase-negative (Das et al., 2024; Nasr \u0026amp; Abd, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTable\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e4\u003c/span\u003e reveals that two isolates (M5 and M9) were classified as hetero-fermentative LAB due to gas production, while seven isolates (M1, M2, M3, M4, M6, M7, and M8) were categorized as homo-fermentative. Homo-fermentative LAB do not generate gas, while hetero-fermentative LAB do. These findings align with Kavitha \u0026amp; Jeevaratnam's (2016) report on homo-fermentative Lactobacillus isolates from piper beetle leaves.\u003c/p\u003e \u003cp\u003eGram-positive and catalase-negative isolates' fermentation type was determined by glucose CO₂ production (Kabir et al., 2022). LABs are classified as homo- or hetero-fermentative based on carbohydrate fermentation end products (Nasr \u0026amp; Abd, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). All tested isolates fermented TSIA media containing glucose, lactose, and sucrose, producing acid. Positive LAB isolates are identified by a yellow color change in these sugars (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe citrate test confirmed that isolates M1\u0026ndash;M9 could not utilize citrate as a carbon source, as indicated by the absence of a color change (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e4\u003c/span\u003e). This suggests a deficiency in citrate-catabolizing enzymes (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), aligning with Hafsan and Mustami (2015), despite some LAB's ability to metabolize citrate (Castro \u0026amp; Poveda, 2024). Additionally, isolates M1\u0026ndash;M9 were non-motile and indole-negative (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e4\u003c/span\u003e), indicating the absence of flagella (Prastujati et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) and tryptophanase, preventing tryptophan breakdown (Hafsan \u0026amp; Mustami, 2015). Moreover, these isolates tested MR-negative (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e4\u003c/span\u003e), confirming the lack of mixed acid fermentation. This finding is consistent with Ngene et al. (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) but differs from some LAB strains found in fermented foods (Sirisha et al., 2021), suggesting that these isolates are unable to produce organic acids such as lactic, succinic, acetic, and formic acid from glucose.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eBiochemical Characteristics of LAB Isolate\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"11\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eCatalase test\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eCitrate test\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eMR test\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c8\" namest=\"c5\"\u003e \u003cp\u003eTSIA test\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c10\" namest=\"c9\"\u003e \u003cp\u003eSIM test\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eFermentative type\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eB\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eGas\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eH\u003csub\u003e2\u003c/sub\u003eS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eM\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eI\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1 (M1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4 (M2)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5 (M3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7 (M4)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8 (M5)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eYR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHetro\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11 (M6)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e14 (M7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e15 (M8)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHomo\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e17 (M9)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eYR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eY\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eHetro\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"11\"\u003e\u003cb\u003eKey\u003c/b\u003e: \u0026ldquo;S\u0026rdquo; slant, \u0026ldquo;B\u0026rdquo; butt, \u0026ldquo;M\u0026rdquo; motility, \u0026ldquo;I\u0026rdquo; indole, \u0026ldquo;Y\u0026rdquo; yellow, \u0026ldquo;R\u0026rdquo; red, \u0026ldquo;\u0026minus;\u0026rdquo; negative, \u0026ldquo;+\u0026rdquo; positive\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e3.4 Physiological Characteristics of LAB Isolates\u003c/h2\u003e \u003cp\u003eMost LAB isolates (77.8%, M1-M3, M5-M7, M9) tolerated all tested temperatures (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e5\u003c/span\u003e), consistent with prior LAB temperature tolerance research (Khalil \u0026amp; Anwar, 2016; Terpou et al., 2019). Temperature control is vital for maintaining probiotic potential in food applications (Sionek et al., 2024). All LAB isolates (M1-M9) grew at 1% and 4% NaCl, with 88.9% (M1-M7, M9) tolerating 6% NaCl (M8 failed at 6%). This broad salt tolerance suggests potential for gastrointestinal survival, crucial for probiotic efficacy (Asha et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Tsega et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Genetic analysis will confirm probiotic safety (Tsega et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2023\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIsolates M3 and M4 survived all tested acidic conditions (pH 2-6.5, Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e5\u003c/span\u003e), with 22% overall tolerance to this range and 88.9% tolerance to pH 4-6.5, suggesting potential gastrointestinal survival (Chen et al., 2020; Iraporda et al., 2024; Hanifa et al., 2020). Acid tolerance, crucial for probiotic efficacy given stomach pH (Russell et al., 1993; Dressman et al., 1990; Sahadeva et al., 2011; El-Beksh et al., 2021), makes M3 and M4 promising candidates in Mekele, Tigray, Ethiopia. All isolates tolerated 0.4% phenol (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e5\u003c/span\u003e), while 78% (M2-M3, M5-M9) resisted 0.6% (M1 and M4 failed). This phenol resistance, important due to inhibitory effects on LAB (Nandha \u0026amp; Shukla, 2023) and gut phenol presence, further supports their probiotic potential in the local context.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePhysiological Characteristics of LAB Isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"11\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" morerows=\"1\" nameend=\"c2\" namest=\"c1\" rowspan=\"2\"\u003e \u003cp\u003ePhysiological\u003c/p\u003e \u003cp\u003echaracteristics\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"9\" nameend=\"c11\" namest=\"c3\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eM4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eM5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eM6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eTemperature (℃)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e27\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e37\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e45\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eNaCl (%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003epH level\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ePhenol conc (%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"11\"\u003e\u003cb\u003eKey\u003c/b\u003e: \u0026ldquo;+\u0026rdquo; indicates bacterial growth, while \u0026ldquo;\u0026minus;\u0026rdquo; indicates no bacterial growth\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e3.5 Safety Evaluation of LAB Isolates\u003c/h2\u003e \u003cp\u003eHemolysis testing confirmed the safety of LAB isolates M1\u0026ndash;M9, as they exhibited γ-hemolysis (non-hemolytic), unlike the β-hemolytic pathogenic controls (Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e6\u003c/span\u003e) (Nandha \u0026amp; Shukla, 2023; Meradji et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). The absence of hemolysin, likely due to the lack of cytolysin genes (Meradji et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), indicates non-virulence (Tsega et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Given that hemolysis testing is a crucial criterion for probiotic selection, these isolates hold promising potential for applications in dairy, non-dairy, and medicinal products, pending further analysis.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 6\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eHemolysis Effect\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"13\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c13\" colnum=\"13\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eHemolytic Effect\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"9\" nameend=\"c10\" namest=\"c2\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"3\" nameend=\"c13\" namest=\"c11\"\u003e \u003cp\u003eControls\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eM4\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eM5\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eM6\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003e\u003cem\u003eE. coli\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c12\"\u003e \u003cp\u003e\u003cem\u003eS. aureus\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c13\"\u003e \u003cp\u003e\u003cem\u003eS. typhimurium\u003c/em\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eα\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eβ\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eγ\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e\u0026minus;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"13\"\u003e\u003cb\u003eKey\u003c/b\u003e: (\u0026minus;) indicates no hemolytic effect, (+) there is hemolytic effect\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cdiv id=\"Sec18\" class=\"Section3\"\u003e \u003ch2\u003e3.5.1 Antibacterial Activity Test\u003c/h2\u003e \u003cp\u003eIn this study, all tested isolates (M1-M9) showed no zone of inhibition (resistance) against \u003cem\u003eE. coli\u003c/em\u003e ATCC 25922 (a) (Table\u0026nbsp;\u003cspan refid=\"Tab6\" class=\"InternalRef\"\u003e7\u003c/span\u003e). This suggests these isolates lack the ability to inhibit \u003cem\u003eE. coli\u003c/em\u003e growth, possibly due to their co-existence without antagonism in milk. Against \u003cem\u003eS. aureus\u003c/em\u003e ATCC 25923 (b), four isolates (M2, M4, M7, and M9) showed no inhibition, four (M3, M5, M6, and M8) showed intermediate inhibition, and only M1 showed sensitivity. Against \u003cem\u003eP. aeruginosa\u003c/em\u003e ATCC 27853 (c), six isolates (M1, M2, M3, M4, M7, and M9) showed no inhibition, M5 showed intermediate inhibition, and M6 and M8 showed sensitivity. These results demonstrate varying degrees of inhibition against selected indicator organisms, consistent with Akabanda et al., (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Consequently, no isolate inhibited \u003cem\u003eE. coli\u003c/em\u003e ATCC 25922, only M1 inhibited \u003cem\u003eS. aureus\u003c/em\u003e ATCC 25923, and only M6 and M8 inhibited \u003cem\u003eP. aeruginosa\u003c/em\u003e ATCC 27853. Rattanachaikunsopon \u0026amp; Phumkhachorn (2010) suggest that LAB antimicrobial activity stems from organic acids and other compounds like ethanol, H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e, diacetyl, reuterin, and bacteriocins. Akabanda et al. (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) also demonstrated that LAB antimicrobial compounds can inhibit pathogenic bacteria growth.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab6\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 7\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntibacterial Test\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"10\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eAntibacterial effect\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"9\" nameend=\"c10\" namest=\"c2\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eM4\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eM5\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eM6\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS. \u003cem\u003eaureus\u003c/em\u003e (ATCC 25923)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(11)S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(7)I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(6)I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(4)I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(3)I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eE. coli\u003c/em\u003e (ATCC 25922)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP. aeruginosa\u003c/em\u003e (ATCC 27853)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(6)I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(9)S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(13)S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eKey\u003c/strong\u003e \u003cp\u003eResults mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD from triplicate experiment. Findings were recorded as no detectable inhibition or resistance (R), weak inhibition zone (\u0026lt;\u0026thinsp;1\u0026ndash;4 mm), moderate inhibition zone (4\u0026ndash;8 mm) or intermediate (I), and strong inhibition zone or sensitive (S) (\u0026gt;\u0026thinsp;8\u0026ndash;12mm) (Akabanda et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section3\"\u003e \u003ch2\u003e3.5.2 Antibiotic Susceptibility Test\u003c/h2\u003e \u003cp\u003eIsolate M1 showed varied antibiotic sensitivity. Isolates M2-M4 and M8 were resistant to all tested antibiotics. Isolates M5, M6, and M7 exhibited similar resistance patterns (clindamycin, ceftriaxone, amikacin, tetracycline), with M6 additionally resistant to erythromycin. Isolate M9 displayed broad resistance, with susceptibility only to penicillin and amikacin. These results align with previous findings: LAB sensitivity to ampicillin (Jadhav \u0026amp; Narayanaswamy, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), and varying sensitivity to ampicillin, erythromycin, tetracycline, and clindamycin (Ramu \u003cem\u003eet al\u003c/em\u003e., 2022). Clindamycin was the most prevalent resistance, followed by ceftriaxone, amikacin, and tetracycline. This is consistent with amikacin resistance reported by Szutowska \u0026amp; Gwiazdowska (2021), but differs from reports of kanamycin and vancomycin resistance (Marchwińska \u0026amp; Gwiazdowska, 2022). Ciprofloxacin resistance was also noted (Jadhav \u0026amp; Narayanaswamy, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Antibiotic resistance may stem from genetic factors (Okoye et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). While antibiotic sensitive is desirable in probiotics, genetic characterization is crucial for safety assurance.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab7\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 8\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntibiotic Susceptibility Test of Isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"13\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c13\" colnum=\"13\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eIsolates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"12\" nameend=\"c13\" namest=\"c2\"\u003e \u003cp\u003eInhibition Zone of Each Antibiotics (mm)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCln\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMe\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eE\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eAmp\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCro\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eP\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eA\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eVan\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eCip\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003eTe\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c12\"\u003e \u003cp\u003eCot\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c13\"\u003e \u003cp\u003eLiv\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(21) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(13) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e(29) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(18) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(27) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e(15) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(18) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e(23) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e(16) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e(22)S\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(10) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(22) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(22) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e(32) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(29) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(19) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e(23) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e(23) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e(24)S\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(21) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e(29) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(18) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(30) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e(10)R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(19) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e(25) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e(27)S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e(26) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e(22)S\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(8) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(23) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e(22) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e(30) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(30) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e(30) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e(20) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e(25) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e(6) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e(16) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003e(24)S\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e(13) R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e(31) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e(21) S\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c12\"\u003e \u003cp\u003e(17) I\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c13\"\u003e \u003cp\u003eR\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"13\"\u003e\u003cb\u003eKey\u003c/b\u003e: Cln-Clindamycin, Me-Meropinium, E-Erythromycin, Amp-Ampicillin, Cro-Ceftriaxone, P-Penicillin G, A-Amikacin, Van-Vancomycin, Cip-Ciprofloxacin, Te-Tetracycline, Cot-Cotrimoxazole, Liv-Levofloxacillin, R-Resistance, I-Intermediate susceptible, S-Susceptible\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003e3.6 Evaluation of Potent LAB as a Starter Culture\u003c/h2\u003e \u003cdiv id=\"Sec21\" class=\"Section3\"\u003e \u003ch2\u003e3.6.1 Formation of Yogurt\u003c/h2\u003e \u003cp\u003eIn this study, 67% of LAB isolates (M1, M2, M3, M7, M8, and M9) successfully fermented pasteurized milk into yogurt, while 33% (M4, M5, and M6) failed, likely due to lactase deficiency. The positive control (Y7) fermented, whereas the negative control (Y8) remained raw. The six yogurt-producing isolates show promise as starter cultures, pending further research. LAB plays a vital role in milk fermentation, typically involving \u003cem\u003eStreptococcus thermophilus\u003c/em\u003e and \u003cem\u003eLactobacillus bulgaricus\u003c/em\u003e, and has broad applications in dairy, food, and feed industries (Widyastuti \u0026amp; Febrisiantosa, 2014).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec22\" class=\"Section3\"\u003e \u003ch2\u003e3.6.2 Physiochemical Properties of Yogurt\u003c/h2\u003e \u003cp\u003eAs shown in Table\u0026nbsp;\u003cspan refid=\"Tab9\" class=\"InternalRef\"\u003e10\u003c/span\u003e, M8 exhibited the highest pH, while M3 had the lowest. M7's pH of 4.6 aligns with Lee \u0026amp; Lucey (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Iranian yogurt pH ranges from 4.06 to 4.64 (Nemati \u003cem\u003eet al\u003c/em\u003e., 2023). Yogurt consumption benefits lactose-intolerant individuals due to lactose conversion to lactic acid during fermentation (Ademosun et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). During milk acidification, pH decreases from 6.7 to \u0026le;\u0026thinsp;4.6 (Lee \u0026amp; Lucey, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). The low pH in all yogurt products is significant, as it inhibits most pathogenic microorganisms (Madora \u003cem\u003eet al\u003c/em\u003e., 2019). However, excessively low pH can also hinder beneficial microorganisms, necessitating optimal pH maintenance.\u003c/p\u003e \u003cp\u003eM2, M7, and M8 showed the highest freezing points (FP), while M9 and Y8 had the lowest. FP, indicating the solidification point, is influenced by sugars, additives, and starter cultures. Pasteurization-induced changes in calcium phosphate complexes and CO\u003csub\u003e2\u003c/sub\u003e pressure can also affect FP. M8 had the highest density, and M3 the lowest, demonstrating the starter culture's impact on yogurt density. Y8 had the highest titratable acidity (TA), and M8 the lowest. Iranian yogurt TA ranges from 0.91 to 1.57% (Nemati \u003cem\u003eet al\u003c/em\u003e., 2023). TA variations are attributed to starter culture differences. Increased TA is crucial for inhibiting undesirable microbial growth and ensuring proper fermentation (Madora \u003cem\u003eet al\u003c/em\u003e., 2019).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab8\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 9\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePhysiochemical Properties of Yogurt Fermented by the Isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003ePhysiochemical Properties\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003epH\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFP\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eTA\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eD\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e40.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e4.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e30.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e10.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.10\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e40.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e3.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e44.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e-0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e27.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.04\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eY7 (\u003cb\u003e+\u003c/b\u003e\u0026thinsp;control)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e4.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e29.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eY8 (-control)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e6.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e-0.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e7.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e27.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"5\"\u003e\u003cb\u003eKey\u003c/b\u003e: Values were given mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD from triplicate experiment. Y-yogurt, D-density, FP-freezing point, TA-titratable acidity\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec23\" class=\"Section3\"\u003e \u003ch2\u003e3.6.3 C\u003cb\u003eompositions of Yogurt\u003c/b\u003e\u003c/h2\u003e \u003cp\u003eAs shown in Table\u0026nbsp;\u003cspan refid=\"Tab9\" class=\"InternalRef\"\u003e10\u003c/span\u003e, M8 had the highest protein content, while M3 had the lowest. Previous reports on cow milk yogurt protein ranged from 2.56 to 4.07 (Ibrahim \u003cem\u003eet al\u003c/em\u003e., 2021; Wulansari \u0026amp; Kusmayadi, 2016; Elnour \u003cem\u003eet al\u003c/em\u003e., 2022). This variability is attributed to the bacterial strains' ability to break down milk protein. M8 also exhibited the highest fat content and M9 the lowest. Fat content in previous studies ranged from 3.61 to 4.51 (Wulansari \u0026amp; Kusmayadi, 2016; Elnour \u003cem\u003eet al\u003c/em\u003e., 2022). Variations in fat are due to starter culture differences and milk composition. M8 had the highest lactose content and M3 the lowest. Elnour \u003cem\u003eet al\u003c/em\u003e. (2022) reported yogurt lactose at 3.20, with differences attributed to starter culture activity (Ayivi \u003cem\u003eet al\u003c/em\u003e., 2020).\u003c/p\u003e \u003cp\u003eM3 had the highest salt content and M1 the lowest. Starter cultures indirectly affect salt perception through flavor compound production. Y8 had the highest ash content and M9 the lowest, aligning with Olugbuyiro \u0026amp; Oseh (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Ash content in previous reports ranged from 0.39 to 0.70 (Elnour \u003cem\u003eet al\u003c/em\u003e., 2022; Ibrahim \u003cem\u003eet al\u003c/em\u003e., 2021). Ash represents the inorganic residue after incineration. M9 had the highest moisture content, and Y7 the lowest, consistent with reports of 87.27% to 87.77% moisture (Wulansari \u0026amp; Kusmayadi, 2016; Ibrahim \u003cem\u003eet al\u003c/em\u003e., 2021). Yogurt composition is influenced by various factors, including carbohydrate (lactose) conversion to lactic acid during fermentation, resulting in lower carbohydrate levels (Elnour \u003cem\u003eet al\u003c/em\u003e., 2022; Ademosun et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab9\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 10\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eCompositions of Yogurts Fermented by the LAB Isolates\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"6\" nameend=\"c7\" namest=\"c2\"\u003e \u003cp\u003eComposition in Percent (%)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eP\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eA\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eM\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eS\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e2.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e9.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e88.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e3.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e87.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e1.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e1.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e2.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e87.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e1.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e6.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e9.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e89.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e5.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e7.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e10.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e88.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e3.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.58\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e89.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e0.7\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e+\u003c/b\u003eve control\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e3.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e2.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e0.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e86.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e1.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e-ve control\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e2.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e3.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e2.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e87.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c7\"\u003e \u003cp\u003e1.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"7\"\u003e\u003cb\u003eKey\u003c/b\u003e: Values were given mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD from triplicate experiment. M-isolate, Y-yogurt P-protein, L-lactose, F-fat, A-ash, M-moisture, S-salt\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec24\" class=\"Section2\"\u003e \u003ch2\u003e3.7 Sensory Evaluation of Yogurt\u003c/h2\u003e \u003cp\u003eAs shown in Table\u0026nbsp;\u003cspan refid=\"Tab10\" class=\"InternalRef\"\u003e11\u003c/span\u003e, Y7's color was rated \"like moderately\" and was well-accepted, while Y8's color was \"dislike extremely\" and rejected. M7's odor was rated \"like extremely\" and was highly accepted. M1, M9, M3, M2, and M8's odors were rated \"like moderately,\" compared to Y7. Y8's odor was unpleasant and rated \"dislike extremely.\" M1's taste was rated \"like extremely,\" while M8 was rated \"like slightly.\" Y8's taste was unpleasant and rejected. M1, M2, M9, and Y7 had good shape/size, resembling normal yogurt. M3, M7, and M8 had slightly less ideal shapes/sizes. Y8 had an abnormal shape/size and was rated \"dislike moderately.\u003c/p\u003e \u003cp\u003eOverall acceptability (OA) was also evaluated. M1, M3, M8, and M9 were rated \"like moderately,\" and Y7 was rated \"like slightly.\" Y8 was rated \"dislike extremely\" in OA, reflecting its poor sensory scores due to the absence of a starter culture. LAB isolates significantly influence aroma, quality, and consistency through varying fermentation potentials (Zapasnik \u003cem\u003eet al\u003c/em\u003e., 2022). Sensory scores are closely linked to free fatty acid concentration and amino acid degradation in products like yogurt. They also play a crucial role in host health (Fijan, 2014).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab10\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 11\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSensory Scores of the Yogurt\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"5\" nameend=\"c6\" namest=\"c2\"\u003e \u003cp\u003eSensory scores\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eColor\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eOdor\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eTest\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eShape/size\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eOverall acceptability\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e5.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e6.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e6.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e6.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e5.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e5.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e5.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e6.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e5.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e6.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e6.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e4.8\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e5.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.42\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e4.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e4.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e6.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e5.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e6.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e6.0\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e+\u003c/b\u003eve control\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e6.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.4\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e6.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e5.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e5.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e-ve control)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e1.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e1.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.71\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e2.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e1.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.42\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eKey\u003c/strong\u003e \u003cp\u003eM-isolate, Y-yogurt, 7\u0026thinsp;=\u0026thinsp;like extremely, 6\u0026thinsp;=\u0026thinsp;like moderately, 5\u0026thinsp;=\u0026thinsp;like slightly, 4\u0026thinsp;=\u0026thinsp;neither like nor dislike, 3\u0026thinsp;=\u0026thinsp;dislike slightly, 2\u0026thinsp;=\u0026thinsp;dislike moderately and 1\u0026thinsp;=\u0026thinsp;dislike extremely). Values mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD from duplicate experiment.\u003c/p\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec25\" class=\"Section2\"\u003e \u003ch2\u003e3.8 Identification of LAB Probiotic at Genus level\u003c/h2\u003e \u003cp\u003eThe LAB isolates in this study were tentatively grouped under the genera \u003cem\u003eLactobacillus\u003c/em\u003e, \u003cem\u003eEnterococcus\u003c/em\u003e, and \u003cem\u003eLeuconostoc\u003c/em\u003e. This classification was primarily based on morphology (rods, cocci, tetrads), glucose fermentation mode (homo- or hetero-fermentative), growth at 15\u0026deg;C and 45\u0026deg;C, and salt tolerance. Yelnetty et al., (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) reported that \u003cem\u003eLactobacillus\u003c/em\u003e was the dominant genus in fermented cow and lamb milk. LAB is typically gram-positive, catalase-negative, non-motile, non-spore-forming rods or cocci, and can be homo- or hetero- fermentative. \u003cem\u003eLactobacillus\u003c/em\u003e species exhibit rod-shaped cells, ranging from short to long, with some species displaying coccobacilli. \u003cem\u003eLactobacillus\u003c/em\u003e and \u003cem\u003eEnterococcus\u003c/em\u003e share similar characteristics, differing mainly in cell shape: rods for \u003cem\u003eLactobacillus\u003c/em\u003e and cocci for \u003cem\u003eEnterococcus\u003c/em\u003e (Ismail et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). \u003cem\u003eLeuconostoc\u003c/em\u003e species are hetero-fermentative cocci.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec26\" class=\"Section2\"\u003e \u003ch2\u003e3.9 Conclusion\u003c/h2\u003e \u003cp\u003eThis study analyzed cow milk composition in Mekele, Ethiopia, identifying and characterizing potential lactic acid bacteria (LAB) for probiotic and yogurt-producing applications. Milk samples showed significant compositional and physicochemical variations, influenced by genetic, environmental, and feed factors. Nine Gram-positive, catalase-negative LAB isolates (M1-M9) were selected from 23 due to their tolerance to temperature, salt, pH, and phenol, and their non-hemolytic activity, indicating safety. Antibacterial and antibiotic susceptibility tests revealed diverse inhibitory effects and resistance patterns. Six isolates successfully fermented pasteurized milk into yogurt, with M1, M3, M8, and M9 receiving favorable sensory evaluations. Biochemical tests classified seven isolates as homo-fermentative and two as hetero-fermentative, with preliminary identification linking them to \u003cem\u003eLactobacillus, Enterococcus\u003c/em\u003e, and \u003cem\u003eLeuconostoc\u003c/em\u003e, common in Ethiopian fermented dairy. The study highlights promising LAB isolates for yogurt production, recommending genetic analysis to confirm safety, probiotic potential, and antibiotic resistance, alongside in vivo studies for efficacy. Standardized starter cultures are essential for industrial fermentation, with future research needed on probiotic mechanisms and their impact on gut microbiota and immune function.\u003c/p\u003e \u003c/div\u003e"},{"header":"Declarations","content":" \u003cp\u003e \u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e \u003cp\u003eThis study was approved by the Mekelle University Institutional Review Board (Approve No. MU-IRB 2305/2024). All methods were conducted according to relevant guidelines and regulations. The study was carried out ethically in accordance with the World Medical Association Declaration of Helsinki (2024 version). Written informed consent was also obtained from zonal and district administrations of dairy farms and milk collection to inform all participants in the research. Informed consent to participate was obtained from all participants in the study.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eConsent for publication\u003c/strong\u003e \u003cp\u003eNot applicable.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eCompeting interest\u003c/strong\u003e \u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eFunding\u003c/h2\u003e \u003cp\u003eThis work was supported by Mekele University (Ethiopia), Hawassa University (Ethiopia), Norwegian University of Life Sciences (Norway) collaboration program phase V (Grant No. MU-HU-NMBU RADO/EXTERNAL/SS003/2024).\u003c/p\u003e\u003ch2\u003eAuthors\u0026rsquo; contributions\u003c/h2\u003e \u003cp\u003eGS wrote and develop the manuscript. GG provided constructive comments. GS and GG edited and finalized the manuscript. DB reviewed the manuscript. All authors strongly contribute to enhance the quality of manuscript. Finally, all authors read and approved the final manuscript.\u003c/p\u003e\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eThe authors thank the Mekelle University for granting M.Sc. to Goiteom Senay Niguse and thank the Scholars Rescue Fund of the International Institute of Education (SRF-IIF) for granting a fellowship to Dr Desta Berhe Sbhatu and the UiTM for hosting him during the preparation of this manuscript.\u003c/p\u003e\u003ch2\u003eAvailability of data and materials\u003c/h2\u003e \u003cp\u003eAll data and materials are available for this work and can be accessed from the corresponding author\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAdemosun OT, Ajanaku KO, Adebayo AH, Oloyede MO, Okere DU, Akinsiku AA, Ajayi SO, Ajanaku CO, Dokunmu TM, Owolabi AO (2019) Physico-Chemical, Microbial and Organoleptic Properties of Yoghurt Fortified with Tomato Juice. J Food Nutr Res 7(11):810\u0026ndash;814. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.12691/jfnr-7-11-9\u003c/span\u003e\u003cspan address=\"10.12691/jfnr-7-11-9\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAkabanda F, Owusu-kwarteng J, Tano-debrah K, Parkouda C, Jespersen L (2014) The Use of Lactic Acid Bacteria Starter Culture in the Production of Nunu, a Spontaneously Fermented Milk Product in Ghana. 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Heliyon 9(7):e17453. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.heliyon.2023.e17453\u003c/span\u003e\u003cspan address=\"10.1016/j.heliyon.2023.e17453\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYelnetty A, Purnomo H, Purwadi MA (2014) Biochemical Characteristics of Lactic Acid Bacteria with Proteolytic Activity and Capability as Starter Culture Isolated From Spontaneous Fermented Local Goat Milk. J Nat Sci R 4(10):137\u0026ndash;146\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Mekelle University","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Lactic acid bacteria, Milk, Probiotics, Starter culture, Yogurt","lastPublishedDoi":"10.21203/rs.3.rs-8906693/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8906693/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eLactic acid bacteria (LAB) have diverse applications in dairy, food, feed, and health, yet their probiotic potential from cow milk remains under-researched. This study aimed to identify and characterize LAB isolates from raw cow milk and evaluate their probiotic suitability. A total of 23 pure bacterial isolates were obtained from seven milk samples, with nine gram-positive, catalase-negative isolates selected for further analysis. These isolates were citrate-negative, non-motile, and indole-negative, with seven exhibiting homo-fermentative traits, and two (M5 and M9) identified as hetero-fermentative. Physicochemical analysis of milk samples showed notable variation in composition, including protein (2.2\u0026ndash;3.2%), lactose (3.3\u0026ndash;4.4%), fat (1.6\u0026ndash;2.6%), ash (0.4\u0026ndash;0.6%), salt (0.4\u0026ndash;0.6%), titratable acidity (0.1\u0026ndash;0.4%), pH (5.7\u0026ndash;6.7%), moisture (83.9\u0026ndash;87.5%), density (25.9\u0026ndash;27.3), and freezing point (0.4\u0026ndash;0.6). Antibacterial tests revealed strong inhibition of \u003cem\u003eS. aureus\u003c/em\u003e by M1 (11 mm) and \u003cem\u003eP. aeruginosa\u003c/em\u003e by M6 and M8 (9 mm and 13 mm), while \u003cem\u003eE. coli\u003c/em\u003e showed complete resistance. Hemolysis tests confirmed LAB isolates as non-pathogenic, with no α- or β-hemolysis observed. Antibiotic susceptibility varied, with M2, M3, M4, and M8 completely resistant, while M1, M5, M6, M7, and M9 showed susceptibility to meropenem, ampicillin, penicillin G, and other antibiotics. Sensory evaluation indicated M1 and M7 had the highest overall acceptability, while M3 and M8 scored lower in color and taste. LAB isolates were tentatively classified under \u003cem\u003eLactobacillus, Enterococcus\u003c/em\u003e, and \u003cem\u003eLeuconostoc\u003c/em\u003e, indicating their probiotic potential and viability as starter cultures in dairy and food industries. Further molecular identification and characterization are needed to determine specific strains and commercial applications.\u003c/p\u003e","manuscriptTitle":"Isolation and Characterization of Lactic Acid Bacteria from Raw Cow Milk and Evaluation of Their Probiotic Potential","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-02-19 15:28:59","doi":"10.21203/rs.3.rs-8906693/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"6b0ab3e4-5aa0-4c55-bf6d-b25016061468","owner":[],"postedDate":"February 19th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-02-19T15:28:59+00:00","versionOfRecord":[],"versionCreatedAt":"2026-02-19 15:28:59","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8906693","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8906693","identity":"rs-8906693","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}