Oxidative Stress Biomarkers in Caprine Clinical Mastitis: Correlation with Somatic Cell Count and Evaluation of Antioxidant Therapy

Oxidative Stress Biomarkers in Caprine Clinical Mastitis: Correlation with Somatic Cell Count and Evaluation of Antioxidant Therapy | 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 Oxidative Stress Biomarkers in Caprine Clinical Mastitis: Correlation with Somatic Cell Count and Evaluation of Antioxidant Therapy Aditya Pratap, Amita Tiwari, Shashi Pradhan, Devendra Kumar Gupta, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6999531/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 Mastitis remains a prevalent and economically significant disease in dairy goats. Oxidative stress, mediated by reactive oxygen species (ROS), is implicated in its pathogenesis, yet limited studies have explored this association in goats. This study aimed to assess oxidative stress biomarkers in caprine clinical mastitis, their correlation with somatic cell count (SCC), and the therapeutic efficacy of antioxidant interventions. A total of 313 lactating goats from both organized and unorganized sectors in Jabalpur were screened over six months, and 27 were diagnosed with clinical mastitis. Milk and heparinized blood samples were analyzed for SCC, milk pH, malondialdehyde (MDA), and reduced glutathione (GSH). Affected goats showed significantly elevated MDA, SCC, and milk pH, along with reduced GSH levels compared to healthy controls. A significant positive correlation (P < 0.05) was observed between SCC and MDA levels. Therapeutic trials compared the efficacy of ascorbic acid and tocopherol-sodium selenite combination alongside antibiotics. The tocopherol-selenium group demonstrated superior clinical recovery and normalization of oxidative stress markers. These results suggest that oxidative stress plays a pivotal role in clinical mastitis in goats and that antioxidant therapy, particularly tocopherol with selenium, can effectively mitigate oxidative damage when used with conventional treatment. Oxidative stress Clinical Mastitis Goats MDA GSH Antioxidants Figures Figure 1 INTRODUCTION Goats are among the most economically significant livestock species worldwide, often referred to as the "poor man's cow" due to their vital role in the livelihoods of landless laborers and marginal farmers, particularly in rural India. Their adaptability, frugality, and multipurpose utility—including meat, milk, and fiber production—make them indispensable in smallholder agricultural systems. Goat milk, in particular, is of considerable nutritional value, bearing a close resemblance to human breast milk in composition. It has gained increasing consumer interest owing to its high levels of oligosaccharides, medium-chain triglycerides, β-casein, and lower αs1-casein content compared to cow milk, conferring various health benefits (Turkmen, 2017 ). Among the diseases affecting dairy goats, mastitis remains a persistent and economically detrimental condition. It is an inflammation of the mammary gland, most commonly caused by bacterial infections but also influenced by fungal or viral pathogens and poor hygiene. Mastitis is broadly classified into clinical mastitis (CM) and subclinical mastitis (SCM). Numerous predisposing factors, including age, breed, parity, stage of lactation, environmental conditions, udder trauma, and suboptimal management practices, contribute to disease onset (Kumar et al., 2018 ). Mastitis not only disrupts milk yield and quality but also induces profound alterations in milk composition, particularly affecting biochemical and oxidative stress parameters. Cells constantly generate reactive oxygen species (ROS) during aerobic metabolism, which are typically neutralized by endogenous antioxidant defences. However, an imbalance favouring oxidants leads to oxidative stress, a state implicated in various pathophysiological conditions including mastitis (Jhambh et al., 2013 ). Milk contains intrinsic antioxidant systems, both enzymatic (catalase, lactoperoxidase, glutathione peroxidase) and non-enzymatic (vitamins A, C, E, selenium, carotenoids) which help preserve milk quality and mammary gland health. During intramammary infections, these antioxidant mechanisms are compromised, and specific oxidative stress markers can serve as potential indicators of mastitis severity (Andrei et al., 2016 ). While antibiotic therapy remains central to mastitis treatment, there is growing interest in the adjunct use of antioxidant supplementation, such as vitamins C and E, selenium, and other trace elements, to mitigate oxidative damage and enhance recovery. Despite this, most existing research has focused on bovine species, with limited information available on goats. Therefore, the present study was undertaken with the following objectives: To assess oxidative stress in clinical mastitis in goats To investigate the relationship between oxidative stress and clinical mastitis To evaluate the efficacy of different antioxidant therapies in affected goats MATERIALS AND METHODS Location, place and duration of work The presented research work was conducted in the Department of Veterinary Medicine, College of Veterinary Science and Animal Husbandry, Nanaji Deshmukh Veterinary Science University, Jabalpur, Madhya Pradesh for a period of six months. Experimental design The study was conducted in two phases:- Phase I - A total of 313 lactating goats were screened for the presence of mastitis by using California mastitis test (CMT). Out of which 27 goats were found positive for clinical mastitis. Phase II – After confirmation of clinical mastitis, 18 goats having CMT score (+ 3) were subjected to estimation of somatic cell count (SCC), parameters of oxidative stress (Malondialdehyde and Reduced Glutathione) and therapy. Additionally, 6 apparently healthy goats were selected to serve as healthy control group. For therapeutic study, 18 goats positive for CM were randomly selected and categorized into three treatment groups (G1, G2 and G3), each group comprised of 6 goats. Six apparently healthy goats were selected as healthy control group (G4). Two different antioxidants viz. Ascorbic acid @ 25mg/kg b.wt. I/M daily for 5 days and Tocopherol & Sodium selenite @1ml/25kg b.wt. S/C once was administered in groups G2 and G3, respectively. Moreover, Goats of all the three groups were commonly treated with parenteral antibiotic (Amoxycilin + sulbactum @10 mg/kg b.wt. I/M) and intramammary infusion of procaine penicillin G, streptomycin sulphate, sulphamerazine and hydrocortisone daily for 5 days. Fluid therapy, symptomatic and supportive therapy was given to goats as per the clinical condition in all the treatment groups as and when required. Parameters of the study History Complete history and patient specific data i.e. age, breed, stage of lactation, parity, drop in milk yield, number of teats affected etc. was recorded in each goat. Clinical examination All the goats under therapeutic trial were clinically examined for presence of symptoms like swelling of udder, abnormal milk secretion and/or enlargement of supramammary lymph node. Clinical parameters i.e. body temperature (ºF), pulse rate (per minute) and respiratory rate (per minute) were measured prior to and post treatment on day 0 and on day 6. Collection of samples Milk samples In confirmed cases of clinical mastitis, milk samples were collected on day ‘0’ pre-treatment and on day ‘6’ post-treatment. For collection of milk, udder of each goat was thoroughly washed with potassium permanganate (0.01 percent) and wiped with clean cloth. About 7 ml of midstream milk were collected aseptically from each half separately in sterilized vials and was transported (in ice box) to the department for further examination. Blood samples Blood samples were collected in confirmed cases of clinical mastitis on day ‘0’ pre-treatment and on day ‘6’ post-treatment. About 3 ml of blood was collected aseptically from jugular vein of the goats in heparinized vaccutainer for the estimation of oxidative indices. Processing of milk samples Somatic cell count (SCC) The somatic cell count was performed in milk samples on day ‘0’ (pre-treatment) and on day ‘6’ (post-treatment) in all the goats of different treatment group. The SCC was performed with the help of automatic milk analyzer (Lactoscan) using lactoscan SCC kit. Procedure The milk sample was first homogenized using a vortex mixer for one minute. Subsequently, 20 µl of the homogenized sample was transferred into an Eppendorf tube using a micropipette and further homogenized for 30 seconds. From this, 0.8 µl of milk was drawn and loaded onto the cartridge. Any excess milk was gently removed using cotton to ensure accurate measurement. The cartridge was then inserted into the cartridge slot of the Lactoscan machine, and the “Continue” button was pressed to initiate the analysis. The results were then displayed on the screen. Processing of blood samples Estimation of lipid peroxidation and reduced glutathione was done on day ‘0’ (pre-treatment) and on day ‘6’ (post-treatment) in all the goats under therapeutic trial. Separation of erythrocytes The heparinized blood samples were centrifuged at 2000 rpm for 15 minutes. Plasma and buffy coat was discarded. The resulting RBCS palate was washed thrice with 0.15 M NaCI. Dilution of 33% RBC was made by using phosphate buffer saline (PBS) and kept it at 4°C until, further analysis. A. Malondialdehyde (MDA) Membrane peroxidative damage in erythrocytes was determined in terms of Malondialdehyde (MDA) production as method suggested by Rehman ( 1984 ). Briefly, one ml of 33 percent of erythrocyte was incubated at 37°C for 2hrs. To each sample, 1ml of 10 percent w/v Trichloroacetic acid (TCA) was added. After thorough mixing, the reaction mixture was centrifuged at 2000 rpm for 10 minutes. Then 1 ml of 0.67 percent w/v of Thiobarbituric acid (TBA) was added in 1 ml of supernatant and kept in boiling water bath for 10 minutes. Reaction was allowed to cooled and diluted with 1ml of distilled water. Blank was made by adding all the reagents except packed erythrocyte. The absorbance of these samples was read at wavelength 535 nm using Bio-spectrometer. Calculation Calculation was done by using following formula. The amount of LPO expressed as nanomole (nM) of MDA formed per ml of packed RBCs. OD- Optical density of test; VR - Total volume of reaction mixture; VS - Volume of sample taken; T I - Time of incubation (2 hours); DF - Dilution factor; EC- Molar extinction coefficient of MDA-TBA complex i.e.1.56x 1011 /M/cm B. Reduced glutathione (GSH) GSH was estimated by the 5, 5 dithiobis (2-nitrobenzoic acid) (DTNB) method as described by Prins and Loos ( 1969 ). 0.2 ml of RBCS pack (33 percent dilution in PBS) was added to 4 ml of 0.08 N H₂SO4, and mixed carefully. After 10 minutes of standing at room temperature, 0.5 ml of tungstate solution was added to clear haemolysate. The tube was stoppered and mixture was shaken vigorously for 5 minutes. The stopper was removed and suspension was allowed to stand for 5 minutes in order to avoid bubble formation on top of the supernatant. The suspension was then centrifuge for 15 minutes at 2000 rpm at room temperature. After centrifugation, 2 ml of supernatant was added to 2.5 ml of tris-buffer, 0.2 ml of DTNB reagent was added and mixed well. Within a minute, absorbance was measured at 412 nm against blank by using Biospectrometer in which 2 ml of distilled water was substituted for supernatant. Calculation Calculation was done by using the following formula and results were expressed as milimole (mM) of GSH per ml of packed RBCs. VR – Total volume of reaction mixture; VS – Volume of sample taken; EC - Extinction coefficient i.e., 13100 /M/cm The response of therapy in different groups was evaluated on the basis of improvement in clinical parameters, milk parameters, oxidative stress parameters and compared with goats of healthy control group. Statistical Analysis The recorded data was analyzed by applying one-way ANOVA and mean comparisons were made by Duncan’s multiple range test and paired t-test was applied for comparison within group between interval as per the standard procedure IBM SPSS computer software version 25.0. RESULT AND DISCUSSION Clinical parameters Clinical parameters were recorded in all the 18 goats under therapeutic trial on day 0 (pre-treatment) and day 6 (post treatment) and compared with goats of healthy control group. All the three clinical parameters were significantly higher in goats of different treatment groups when compared with healthy control group on day 0 (pre treatment) although, significant improvement were noticed in all treatment groups on day 6 (post treatment). The detailed variations in clinical parameters in different treatment groups at different intervals are outlined in table 01. Table 01: Mean±SE value of clinical parameters in goats in different treatment groups at different intervals Groups (n=6) Body temperature (Mean±SE) Respiration rate (Mean±SE) Pulse rate (Mean±SE) Day 0 Day 6 Day 0 Day 6 Day 0 Day 6 G 1 104.0 aA ±0.15 102.6 aB ±0.17 44.7 aA ±1.94 27.3 aB ±1.15 94.2 aA ±1.64 72.8 aB ±1.33 G 2 104.1 aA ±0.19 102.5 aB ±0.14 45.7 aA ±1.87 26.5 aB ±1.48 93.3 aA ±2.91 72.7 aB ±1.52 G 3 104.1 aA ±0.18 102.3 aB ±0.20 45.8 aA ±1.82 24.8 aB ±1.01 95.2 aA ±2.83 71.7 aB ±1.05 G 4 102.1 bA ±0.19 102.2 aA ±0.16 24.2 bA ±0.95 24.3 aA ±0.99 71.3 bA ±1.54 71.2 aA ±1.17 Mean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p<0.05) As most of the studies are conducted on SCM in goats and the literature on the study of clinical parameters in clinical mastitis in goats is meager. Although Constable et al . (2007) described three categories of CM as abnormal milk, abnormal animal and abnormal udder. Abnormal animal shows systemic disease and is pyrexic. Macun et al . (2010) reported transient fever, rise in pulse and respiration rate along with other symptoms in goats having CM due to Mycoplasma agalactia. When compared with goats of healthy control group in the present study all the clinical parameters were raised significantly on day 0 (pre treatment). Raised clinical parameters in ailing goats are an indication of ongoing systemic infection and improvement in these parameters after treatment showed the efficacy of drugs used in the present study. Milk parameters Somatic cell count (SCC) and milk pH in all the 18 lactating goats under therapeutic trial was recorded on day 0 (pre-treatment) and day 6 (post treatment) and compared with the healthy control goats. Significant increase in mean SCC as well as mean milk pH were observed in goats of all treatment groups at day 0 pre treatment as compared to healthy control group. A significant improvement in milk parameters of goats was noticed in all treatment groups on day 6 post treatment. The detailed variations in milk parameters in different treatment groups at different intervals are depicted in table 02. The results of present study correlate well with the findings of Nava et al . (2008), Persson and Olofsson (2011) and Raikwar and Shukla (2015) who have observed an increase in SCC in the milk of goats affected with mastitis. It is the most widely used indicator for udder health in cow, sheep and goat milk. As compared to sheep and cow, SCC in goat milk is relatively high even in the healthy udder (Persson and Olofsson, 2011). However, Poutrel et al . (1997) stated that elevated SCC is mainly a response to infection. Therefore, measurement of somatic cell count is a reliable indicator to detect mastitis and other intramammary infection in goats. Table 02: Mean value of milk parameters in goats in different treatment groups at different intervals Groups (n=6) SCC (Mean±SE) pH (Mean±SE) Day 0 Day 6 Day 0 Day 6 G 1 19.48 aA ±2.84 4.75 aB ±0.36 7.49 aA ±0.10 6.65 aB ±0.03 G 2 19.40 aA ±3.20 4.37 aB ±0.47 7.40 aA ±0.08 6.62 aB ±0.08 G 3 19.60 aA ±3.58 3.80 aB ±0.35 7.47 aA ±0.09 6.61 aB ±0.05 G 4 03.70 bA ±0.37 3.72 aA ±0.34 6.60 bA ±0.04 6.60 aA ±0.04 Mean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p<0.05) The results of present study are in accordance with Islam et al . (2012), Hasan (2013) and Tomar et al . (2018) who also reported higher pH of milk in goats affected with mastitis. The probable reason for increase in milk pH in mastitis may be the damage to the blood-milk barrier in mastitis leading to leakage of epithelial tight junctions. As a result blood and extracellular fluid components mix with secreted milk (Zhao and Lacasse, 2008) and showed the trends of increased milk pH with the increase in the severity of inflammatory process (Qayyum et al ., 2016). Thus, milk pH may serve as one of the good indicator to assess udder health and a difference in milk pH may indicate severity of intramammary infection. The improvement in milk pH after therapy in the present work may indicate recovery of animals after treatment. Oxidative stress parameters Oxidative stress parameters i.e. malondialdehyde (MDA) and reduced glutathione (GSH) value were calculated in blood samples in all the 18 lactating goats under therapeutic trial on day 0 (pre-treatment) and day 6 (post treatment) and compared with that of healthy control goats. MDA value on day 0 pre-treatment was significantly higher in goats of all treatment groups in comparison to healthy control group. A significant improvement in the MDA value of goats was noticed in all treatment groups on day 6 post treatment. However maximum improvement was seen in goats of group G3. The detail of MDA value in different treatment groups at different intervals are outlined in table 03. The findings of present study are in agreement with the findings of El-deeb, (2013), who reported significantly higher level of MDA with gangrenous mastitis in goats. Moreover, Kumar et al . (2007) and Mahapatra et al . (2018) also reported significant elevation of blood MDA level in clinical mastitis in bovines. Malondialdehyde is a main defined reaction product in biological system which undergoes lipid peroxidation and its level are the measures of the lipid peroxidation. The elevated level of MDA in the present study indicated a significant increase in lipid peroxidation as a result of the involvement of oxidative stress and the possible oxidative damage in clinical mastitis. The post treatment improvement in the level of MDA in all the goats indicated the effect of antioxidant used in the therapeutic regimen. Table 03: Mean value of oxidative stress parameters in goats in different treatment groups at different intervals Groups (n=6) MDA (Mean±SE) GSH (Mean±SE) Day 0 Day 6 Day 0 Day 6 G 1 18.85 aA ±1.98 8.99 aB ±1.16 0.06 bB ±0.01 0.14 bA ±0.01 G 2 19.77 aA ±2.38 7.06 abB ±1.06 0.07 bB ±0.01 0.17 abA ±0.01 G 3 20.61 aA ±1.56 5.97 bB ±0.63 0.07 bB ±0.01 0.18 aA ±0.01 G 4 5.29 bA ±0.92 5.44 bA ±0.90 0.18 aA ±0.01 0.18 aA ±0.01 Mean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p<0.05) GSH value on day 0 pre-treatment was significantly lower in goats of all treatment groups in comparison to healthy control group. A significant improvement in the GSH value of goats was noticed in all treatment groups on day 6 post treatment. However maximum improvement was noticed in goats of group G3. The detail of GSH values in different treatment groups at different intervals are outlined in table 03. The results of present investigation correlates well with findings of Jhambh et al . (2013) and El-deeb (2013) who reported significant decrease in GSH concentration in mastitic cow and does as compared to healthy ones. Glutathione, a thiol-containing tripeptide is present in living cells at high concentration in its reduced form (GSH). Upon reaction with reactive oxygen species, it got oxidized to glutathione radical which can be regenerated to its reduced form by glutathione reductase (Kohen and Nyska, 2002). A significant decrease in GSH concentration in goats during mastitis in the present study may be due to conversion of reduced form to oxidized form by excessive production of reactive oxygen species from inflamed gland (Jhambh et al ., 2013) and the improvement in its level might be due to the effect of therapy with antioxidants. Correlation between oxidative stress parameters and milk parameters A positive and significant correlation (P < 0.05) was observed between SCC and MDA levels. Thus, it was concluded that oxidative stress parameters, specifically MDA and GSH, were significantly altered in goats suffering from clinical mastitis, with a positive correlation observed with SCC. Parameter GSH MDA SCC SCC -0.83 0.88* 1 MDA -0.52 1 0.88* GSH 1 -0.52 -0.83 *Significant correlation between the parameters (P<0.05) In mastitic milk, the concentration of MDA is higher than in healthy milk, demonstrating that during infectious processes there is an accumulation of ROS and products resulting from oxidative degradation of biomolecules (Andrei et al., 2016). Furthermore, increased SCC may be correlated with an increase in MDA levels and a decrease in the proportion of unsaturated fatty acids, as studies conducted on bovine milk show (Guerrero, et al., 2015). Yehia et al ., (2024) also recorded the same finding and indicate that a significant correlation between different parameters in blood and milk can be useful as a proper clinical indicator for the early detection of mastitis in cows. Therapeutic response evaluation The response of therapy in different treatment group was evaluated on the basis of improvement in clinical abnormalities, milk parameters and oxidative stress markers on day 6 post treatment in goats of all the treatment groups. The antibiotic used in all the treatment groups in the present study i.e. combination of amoxycilin and sulbactum @10 mg/kg b.wt. intramuscularly for 5 days was found effective in alleviating the mastitis. The efficacy of amoxicillin and sulbactum against mastitis is well documented by Roberson et al . (2004) and Sharma et al . (2020) as it possesses excellent antibacterial activity and has high promise for clinical application particularly in mastitis in livestock in small ruminants. Further, Constable et al . (2007) stated that parenteral antimicrobial therapy should be considered in all cases of clinical mastitis and it must be used daily for 3 to 5 days. As multiple pathogens are responsible for causing mastitis. So, intramammary antimicrobial therapy is in common use for the treatment of certain causes of mastitis like streptococcus agalactia, which is very sensitive to intramammary therapy as compared to systemic therapy (Constable et al ., 2007). Mastitis is one such disease in which antioxidant defense system of body is compromised. Oxidative stress in the ailing animal occurs when the production of reactive oxygen metabolites exceed the capacity of antioxidant of the body. In case of clinical mastitis excessive reactive oxygen species production from the inflamed mammary gland occurs. The use of antioxidants viz. Ascorbic acid (in group G2) and Tocopherol and selenium (in group G3) in the present study ameliorated the oxidative stress in the mastitic goats. Ascorbic acid, used in the present study, is an important water soluble nonenzymatic antioxidants which can be synthesize in the body of goats. Goat’s milk contains on average 1.29 mg/100 g of ascorbic acid, more than cow’s milk (0.94 mg/100 g) (Park et al., 2007). The amount of ascorbic acid decreases during pathological processes, for example in mastitis (Andrei et al., 2016). Moreover, in milk containing a large number of somatic cells, the concentration of antioxidant vitamins is lower compared to milk with a low SCC (Andrei et al., 2016). Thus, in the case of subclinical mastitis, there is a pronounced reduction in the concentration of vitamin C; this is due to the fact that ascorbic acid is consumed during oxidative processes. In a healthy goat mammary gland, the concentration of vitamin C in the milk is higher by 36.4% compared to an infected udder tissue (Silanikove et al., 2014). A meager literature regarding use of ascorbic acid in goat mastitis is available. However, Klezkowski et al . (2005) reported that cow suffering from mastitis have lower concentration of vitamin C in their milk and plasma. Ranjan et al . (2005) have also reported the therapeutic value of vitamin C administered in cow with mastitis. Ascorbic acid has multidimensional bioactive role and its antioxidant role is to neutralize the toxic peroxides and reactive oxygen species released during cellular activation due to bacterial infection, which cause tissue injury (Mahapatra et al ., 2018). Tocopherol and selenium is a fat soluble antioxidant and has been shown to reduce the severity of mastitis by various scientists (Mukherjee, 2007 and Khangal et al ., 2015). Goat’s milk contains on average 0.04 mg/100 g of vitamin E, its concentration depending on diet, season or stage of lactation (Park et al., 2007). Vitamin E has an important role in lipid membrane protection from the attack of reactive oxygen species. It enhances the efficiency of neutrophils by protecting them from oxidative damage following intracellular killing of ingested bacteria (Yang and lee, 2015). In the present study both Ascorbic acid (Vitamin C) and Tocopherol (Vitamin E) & selenium ameliorated the effect of oxidative stress during the mastitis in goats. The effect of vitamin C was moderate but vitamin E and selenium was proved as better antioxidant as compared to vitamin C because its role can be better understood on mastitis in a way that it’s act as lipid soluble antioxidant, free radical scavenger and protects against lipid peroxidation (Yang et al ., 2011). CONCLUSION A significant positive correlation (P < 0.05) was found between SCC and MDA levels indicating that oxidative stress is linked to the severity of mastitis. Enzymes like MDA, GSH and SOD are elevated in clinical mastitis and could serve as biomarkers for early detection of mastitis. These enzymes offer potential for real-time, online monitoring of mastitis in large dairy herds. Better diagnostics could lead to early mastitis detection, improving herd health, milk quality and overall dairy farm productivity. On the basis of clinical recovery, milk parameters and restoration of oxidative stress parameters, Tocopherol & Sodium selenite was proved as a better antioxidant in comparison of Ascorbic acid for the correction of oxidative stress produced by clinical mastitis in goats. Declarations ACKNOWLEDGEMENT The authors are grateful to the Department of Veterinary Medicine, College of Veterinary Science and Animal Husbandry, Nanaji Deshmukh Veterinary Science University, Jabalpur, for providing the necessary facilities and support to carry out this research work. The authors also extend their sincere thanks to the livestock owners for their cooperation during sample collection. STATEMENT OF ANIMAL ETHICS All procedures involving animals were conducted in accordance with the guidelines of the Institutional Animal Ethics Committee (IAEC). The study protocol was reviewed and approved by the IAEC of Nanaji Deshmukh Veterinary Science University, Jabalpur, Madhya Pradesh, India (CPCSEA Registration No. 2071/GO/Re/S/19/CPCSEA). Adequate measures were taken to minimize animal discomfort and ensure humane handling during sample collection and clinical examination. 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Enhanced erythrocytic lipid peroxides and reduced plasma ascorbic acid and alteration in blood trace elements level in dairy cows with mastitis. Veterinary Research Communications , 29 (1): 27-34. Rehman, S.U. (1984). Lead induced regional lipid peroxidation in brain. Toxicology Letters , 21 (3): 333-337. Roberson, J.R., Warnick, L.D. and Moore, G. (2004). Mild to moderate clinical mastitis: efficacy of intramammary amoxicillin, frequent milk out, a combined intramammary amoxicillin and frequent milk out treatment versus no treatment. Journal of Dairy Science , 87 (3): 583-592. Sharma, S.K., Meena, H.K. and Joshi, M. (2020). Studies on antibiotic sensitivity pattern of bacterial isolates from subclinical mastitis of Sirohi goats. Indian Journal of Veterinary Science and Biotechnology , 16 (4): 95-97. Shrivastava, S. (2008). Studies on dual status of livestock both for food and family. Indian Journal of Veterinary Medicine, 35 (1): 23. Silanikove, N., Merin, U., Shapiro, F. and Leitner, G. (2014). Subclinical mastitis in goats is associated with upregulation of nitric oxide-derived oxidative stress that causes reduction of milk antioxidative properties and impairment of its quality. Journal of Dairy Science , 97 (6): 3449-3455. Tomar, A., Shukla, P.C., Singh, B. and Sheikh, A. (2018). Comparative therapeutic efficacy of various teat dip solutions in caprine mastitis . International Journal of Chemical Studies , 6 (4): 123-127. Turkmen, N. (2017). The nutritional value and health benefits of goat milk components. In Nutrients in Dairy and their Implications on Health and Disease (pp. 441-449). Academic Press. Yang, F.L. and Li, X.S. (2015). Role of antioxidants vitamins and trace elements in mastitis in dairy cows. Journal of Advanced Veterinary and Animal Research , 2 (1): 1-9. Yang, F.L., Li, X.S. and He, B.X. (2011). Effects of vitamins and trace-elements supplementation on milk production in dairy cows: a review. African Journal of Biotechnology, 10 : 2574-2578. Yehia, S.G., Saad, M.F., Mosallam, T.E., Abdel-Mobdy, A.E., Megahed, E.A., Aly, H.H. and Ramadan, E.S. (2024). Evaluation of oxidative stress, compositional and biochemical changes in milk and serum of cows with subclinical mastitis. Comparative Clinical Pathology , 33 (4): 643-652. Zhao, X. and Lacasse, P. (2008). Mammary tissue damage during bovine mastitis: Causes and control. Journal of Animal Science , 86 (13); 57-65. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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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-6999531","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":480387756,"identity":"48b5ed63-e3d3-43d7-ba18-537346b29a25","order_by":0,"name":"Aditya Pratap","email":"data:image/png;base64,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","orcid":"","institution":"NDVSU","correspondingAuthor":true,"prefix":"","firstName":"Aditya","middleName":"","lastName":"Pratap","suffix":""},{"id":480387757,"identity":"923d532e-aac4-4e3b-8a47-b5cfe8f06ab0","order_by":1,"name":"Amita Tiwari","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Amita","middleName":"","lastName":"Tiwari","suffix":""},{"id":480387758,"identity":"2aa959ef-b5ce-492b-808b-35446e45208a","order_by":2,"name":"Shashi Pradhan","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Shashi","middleName":"","lastName":"Pradhan","suffix":""},{"id":480387759,"identity":"4131a3b4-8b36-494f-80d9-49854b422a6f","order_by":3,"name":"Devendra Kumar Gupta","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Devendra","middleName":"Kumar","lastName":"Gupta","suffix":""},{"id":480387760,"identity":"15c083ab-f2a4-45fb-8545-7f12bbbe475f","order_by":4,"name":"Amita Dubey","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Amita","middleName":"","lastName":"Dubey","suffix":""},{"id":480387761,"identity":"9fe72275-a992-4f96-93c8-c9b7a22c444e","order_by":5,"name":"Salil Kumar Pathak","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Salil","middleName":"Kumar","lastName":"Pathak","suffix":""},{"id":480387762,"identity":"44df40d7-4e93-4756-96d9-4bc0205e3c88","order_by":6,"name":"Shashank Vishvakarma","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Shashank","middleName":"","lastName":"Vishvakarma","suffix":""},{"id":480387763,"identity":"f3bccf5a-80f5-438a-a0bf-36e055ee687e","order_by":7,"name":"Varsha Mishra","email":"","orcid":"","institution":"NDVSU","correspondingAuthor":false,"prefix":"","firstName":"Varsha","middleName":"","lastName":"Mishra","suffix":""}],"badges":[],"createdAt":"2025-06-28 19:23:10","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6999531/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6999531/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":88616204,"identity":"4ff2279c-e18b-4a09-9910-92d942623c57","added_by":"auto","created_at":"2025-08-08 10:51:21","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":292304,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eProcessing and estimation of somatic cell count in milk\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(a) Loading of 0.8µl milk sample of clinical mastitis affected goat into cartridge\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(b) Insertion of cartridge into cartridge slot of automatic lactoscan\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(c) Result display on screen attached with automatic lactoscan\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-6999531/v1/36dc98fd9d3c1f51d30bf420.png"},{"id":89719571,"identity":"1dc9b5cc-bb2d-46f0-810c-ea3495dc2a84","added_by":"auto","created_at":"2025-08-23 08:31:51","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1490472,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6999531/v1/d6987929-d049-461c-a7d2-16dc3bb38701.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Oxidative Stress Biomarkers in Caprine Clinical Mastitis: Correlation with Somatic Cell Count and Evaluation of Antioxidant Therapy","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eGoats are among the most economically significant livestock species worldwide, often referred to as the \"poor man's cow\" due to their vital role in the livelihoods of landless laborers and marginal farmers, particularly in rural India. Their adaptability, frugality, and multipurpose utility\u0026mdash;including meat, milk, and fiber production\u0026mdash;make them indispensable in smallholder agricultural systems. Goat milk, in particular, is of considerable nutritional value, bearing a close resemblance to human breast milk in composition. It has gained increasing consumer interest owing to its high levels of oligosaccharides, medium-chain triglycerides, β-casein, and lower αs1-casein content compared to cow milk, conferring various health benefits (Turkmen, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Among the diseases affecting dairy goats, mastitis remains a persistent and economically detrimental condition. It is an inflammation of the mammary gland, most commonly caused by bacterial infections but also influenced by fungal or viral pathogens and poor hygiene. Mastitis is broadly classified into clinical mastitis (CM) and subclinical mastitis (SCM). Numerous predisposing factors, including age, breed, parity, stage of lactation, environmental conditions, udder trauma, and suboptimal management practices, contribute to disease onset (Kumar et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eMastitis not only disrupts milk yield and quality but also induces profound alterations in milk composition, particularly affecting biochemical and oxidative stress parameters. Cells constantly generate reactive oxygen species (ROS) during aerobic metabolism, which are typically neutralized by endogenous antioxidant defences. However, an imbalance favouring oxidants leads to oxidative stress, a state implicated in various pathophysiological conditions including mastitis (Jhambh et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eMilk contains intrinsic antioxidant systems, both enzymatic (catalase, lactoperoxidase, glutathione peroxidase) and non-enzymatic (vitamins A, C, E, selenium, carotenoids) which help preserve milk quality and mammary gland health. During intramammary infections, these antioxidant mechanisms are compromised, and specific oxidative stress markers can serve as potential indicators of mastitis severity (Andrei et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). While antibiotic therapy remains central to mastitis treatment, there is growing interest in the adjunct use of antioxidant supplementation, such as vitamins C and E, selenium, and other trace elements, to mitigate oxidative damage and enhance recovery. Despite this, most existing research has focused on bovine species, with limited information available on goats. Therefore, the present study was undertaken with the following objectives:\u003c/p\u003e\u003cp\u003e\u003cul\u003e\u003cli\u003e\u003cp\u003eTo assess oxidative stress in clinical mastitis in goats\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eTo investigate the relationship between oxidative stress and clinical mastitis\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eTo evaluate the efficacy of different antioxidant therapies in affected goats\u003c/p\u003e\u003c/li\u003e\u003c/ul\u003e\u003c/p\u003e"},{"header":"MATERIALS AND METHODS","content":"\u003cp\u003e\u003cstrong\u003eLocation, place and duration of work\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe presented research work was conducted in the Department of Veterinary Medicine, College of Veterinary Science and Animal Husbandry, Nanaji Deshmukh Veterinary Science University, Jabalpur, Madhya Pradesh for a period of six months.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExperimental design\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was conducted in two phases:-\u003c/p\u003e\n\u003cp\u003ePhase I - A total of 313 lactating goats were screened for the presence of mastitis by using California mastitis test (CMT). Out of which 27 goats were found positive for clinical mastitis.\u003c/p\u003e\n\u003cp\u003ePhase II \u0026ndash; After confirmation of clinical mastitis, 18 goats having CMT score (+\u0026thinsp;3) were subjected to estimation of somatic cell count (SCC), parameters of oxidative stress (Malondialdehyde and Reduced Glutathione) and therapy. Additionally, 6 apparently healthy goats were selected to serve as healthy control group.\u003c/p\u003e\n\u003cp\u003eFor therapeutic study, 18 goats positive for CM were randomly selected and categorized into three treatment groups (G1, G2 and G3), each group comprised of 6 goats. Six apparently healthy goats were selected as healthy control group (G4). Two different antioxidants viz. Ascorbic acid @ 25mg/kg b.wt. I/M daily for 5 days and Tocopherol \u0026amp; Sodium selenite @1ml/25kg b.wt. S/C once was administered in groups G2 and G3, respectively. Moreover, Goats of all the three groups were commonly treated with parenteral antibiotic (Amoxycilin\u0026thinsp;+\u0026thinsp;sulbactum @10 mg/kg b.wt. I/M) and intramammary infusion of procaine penicillin G, streptomycin sulphate, sulphamerazine and hydrocortisone daily for 5 days. Fluid therapy, symptomatic and supportive therapy was given to goats as per the clinical condition in all the treatment groups as and when required.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eParameters of the study\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHistory\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eComplete history and patient specific data i.e. age, breed, stage of lactation, parity, drop in milk yield, number of teats affected etc. was recorded in each goat.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical examination\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll the goats under therapeutic trial were clinically examined for presence of symptoms like swelling of udder, abnormal milk secretion and/or enlargement of supramammary lymph node.\u003c/p\u003e\n\u003cp\u003eClinical parameters i.e. body temperature (\u0026ordm;F), pulse rate (per minute) and respiratory rate (per minute) were measured prior to and post treatment on day 0 and on day 6.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCollection of samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMilk samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn confirmed cases of clinical mastitis, milk samples were collected on day \u0026lsquo;0\u0026rsquo; pre-treatment and on day \u0026lsquo;6\u0026rsquo; post-treatment. For collection of milk, udder of each goat was thoroughly washed with potassium permanganate (0.01 percent) and wiped with clean cloth. About 7 ml of midstream milk were collected aseptically from each half separately in sterilized vials and was transported (in ice box) to the department for further examination.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eBlood samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBlood samples were collected in confirmed cases of clinical mastitis on day \u0026lsquo;0\u0026rsquo; pre-treatment and on day \u0026lsquo;6\u0026rsquo; post-treatment. About 3 ml of blood was collected aseptically from jugular vein of the goats in heparinized vaccutainer for the estimation of oxidative indices.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eProcessing of milk samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSomatic cell count (SCC)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe somatic cell count was performed in milk samples on day \u0026lsquo;0\u0026rsquo; (pre-treatment) and on day \u0026lsquo;6\u0026rsquo; (post-treatment) in all the goats of different treatment group. The SCC was performed with the help of automatic milk analyzer (Lactoscan) using lactoscan SCC kit.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eProcedure\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe milk sample was first homogenized using a vortex mixer for one minute. Subsequently, 20 \u0026micro;l of the homogenized sample was transferred into an Eppendorf tube using a micropipette and further homogenized for 30 seconds. From this, 0.8 \u0026micro;l of milk was drawn and loaded onto the cartridge. Any excess milk was gently removed using cotton to ensure accurate measurement. The cartridge was then inserted into the cartridge slot of the Lactoscan machine, and the \u0026ldquo;Continue\u0026rdquo; button was pressed to initiate the analysis. The results were then displayed on the screen.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eProcessing of blood samples\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEstimation of lipid peroxidation and reduced glutathione was done on day \u0026lsquo;0\u0026rsquo; (pre-treatment) and on day \u0026lsquo;6\u0026rsquo; (post-treatment) in all the goats under therapeutic trial.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSeparation of erythrocytes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe heparinized blood samples were centrifuged at 2000 rpm for 15 minutes. Plasma and buffy coat was discarded. The resulting RBCS palate was washed thrice with 0.15 M NaCI. Dilution of 33% RBC was made by using phosphate buffer saline (PBS) and kept it at 4\u0026deg;C until, further analysis.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA. Malondialdehyde (MDA)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMembrane peroxidative damage in erythrocytes was determined in terms of Malondialdehyde (MDA) production as method suggested by Rehman (\u003cspan class=\"CitationRef\"\u003e1984\u003c/span\u003e). Briefly, one ml of 33 percent of erythrocyte was incubated at 37\u0026deg;C for 2hrs. To each sample, 1ml of 10 percent w/v Trichloroacetic acid (TCA) was added. After thorough mixing, the reaction mixture was centrifuged at 2000 rpm for 10 minutes. Then 1 ml of 0.67 percent w/v of Thiobarbituric acid (TBA) was added in 1 ml of supernatant and kept in boiling water bath for 10 minutes. Reaction was allowed to cooled and diluted with 1ml of distilled water. Blank was made by adding all the reagents except packed erythrocyte. The absorbance of these samples was read at wavelength 535 nm using Bio-spectrometer.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCalculation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCalculation was done by using following formula. The amount of LPO expressed as nanomole (nM) of MDA formed per ml of packed RBCs.\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"data:image/png;base64,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\" width=\"363\" height=\"114\"\u003e\u003c/p\u003e\n\u003cp\u003eOD- Optical density of test; VR - Total volume of reaction mixture; VS - Volume of sample taken; T\u003csub\u003eI\u003c/sub\u003e - Time of incubation (2 hours); DF - Dilution factor; EC- Molar extinction coefficient of MDA-TBA complex i.e.1.56x 1011 /M/cm\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eB. Reduced glutathione (GSH)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eGSH was estimated by the 5, 5 dithiobis (2-nitrobenzoic acid) (DTNB) method as described by Prins and Loos (\u003cspan class=\"CitationRef\"\u003e1969\u003c/span\u003e). 0.2 ml of RBCS pack (33 percent dilution in PBS) was added to 4 ml of 0.08 N H₂SO4, and mixed carefully. After 10 minutes of standing at room temperature, 0.5 ml of tungstate solution was added to clear haemolysate. The tube was stoppered and mixture was shaken vigorously for 5 minutes. The stopper was removed and suspension was allowed to stand for 5 minutes in order to avoid bubble formation on top of the supernatant. The suspension was then centrifuge for 15 minutes at 2000 rpm at room temperature. After centrifugation, 2 ml of supernatant was added to 2.5 ml of tris-buffer, 0.2 ml of DTNB reagent was added and mixed well. Within a minute, absorbance was measured at 412 nm against blank by using Biospectrometer in which 2 ml of distilled water was substituted for supernatant.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCalculation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCalculation was done by using the following formula and results were expressed as milimole (mM) of GSH per ml of packed RBCs.\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"data:image/png;base64,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\" width=\"467\" height=\"115\"\u003e\u003c/p\u003e\n\u003cp\u003eVR \u0026ndash; Total volume of reaction mixture; VS \u0026ndash; Volume of sample taken; EC - Extinction coefficient i.e., 13100 /M/cm\u003c/p\u003e\n\u003cp\u003eThe response of therapy in different groups was evaluated on the basis of improvement in clinical parameters, milk parameters, oxidative stress parameters and compared with goats of healthy control group.\u003c/p\u003e\n\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\n \u003ch2\u003eStatistical Analysis\u003c/h2\u003e\n \u003cp\u003eThe recorded data was analyzed by applying one-way ANOVA and mean comparisons were made by Duncan\u0026rsquo;s multiple range test and paired t-test was applied for comparison within group between interval as per the standard procedure IBM SPSS computer software version 25.0.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"RESULT AND DISCUSSION","content":"\u003cp\u003e\u003cstrong\u003eClinical parameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eClinical parameters were recorded in all the 18 goats under therapeutic trial on day 0 (pre-treatment) and day 6 (post treatment) and compared with goats of healthy control group. \u0026nbsp; All the three clinical parameters were significantly higher in goats of different treatment groups when compared with healthy control group on day 0 (pre treatment) although, significant improvement were noticed in all treatment groups on day 6 (post treatment). The detailed variations in clinical parameters in different treatment groups at different intervals are outlined in table 01.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 01: Mean\u0026plusmn;SE value of clinical parameters in goats in different treatment groups at different intervals\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 69px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(n=6)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 197px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBody temperature (Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 181px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eRespiration rate\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 169px;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePulse rate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 69px;\"\u003e\n \u003cp\u003eG \u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e104.0\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e102.6\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e44.7\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e27.3\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e94.2\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.64\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e72.8\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.33\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 69px;\"\u003e\n \u003cp\u003eG \u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e104.1\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e102.5\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e45.7\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.87\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e26.5\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e93.3\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;2.91\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e72.7\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.52\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 69px;\"\u003e\n \u003cp\u003eG \u003csub\u003e3\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e104.1\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e102.3\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e45.8\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.82\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e24.8\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e95.2\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;2.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e71.7\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 69px;\"\u003e\n \u003cp\u003eG \u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e102.1\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e102.2\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e24.2\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.95\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 91px;\"\u003e\n \u003cp\u003e24.3\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e71.3\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;1.54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 84px;\"\u003e\n \u003cp\u003e71.2\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.17\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eMean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p\u0026lt;0.05) \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAs most of the studies are conducted on SCM in goats and the literature on the study of clinical parameters in clinical mastitis in goats is meager. Although Constable \u003cem\u003eet al\u003c/em\u003e. (2007) described three categories of CM as abnormal milk, abnormal animal and abnormal udder. Abnormal animal shows systemic disease and is pyrexic. Macun \u003cem\u003eet al\u003c/em\u003e. (2010) reported transient fever, rise in pulse and respiration rate along with other symptoms in goats having CM due to Mycoplasma agalactia. When compared with goats of healthy control group in the present study all the clinical parameters were raised significantly on day 0 (pre treatment). Raised clinical parameters in ailing goats are an indication of ongoing systemic infection and improvement in these parameters after treatment showed the efficacy of drugs used in the present study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMilk parameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSomatic cell count (SCC) and milk pH in all the 18 lactating goats under therapeutic trial was recorded on day 0 (pre-treatment) and day 6 (post treatment) and compared with the healthy control goats. Significant increase in mean SCC as well as mean milk pH were observed in goats of all treatment groups at day 0 pre treatment as compared to healthy control group. A significant improvement in milk parameters of goats was noticed in all treatment groups on day 6 post treatment. The detailed variations in milk parameters in different treatment groups at different intervals are depicted in table 02.\u003c/p\u003e\n\u003cp\u003eThe results of present study correlate well with the findings of Nava \u003cem\u003eet al\u003c/em\u003e. (2008), Persson and Olofsson (2011) and Raikwar and Shukla (2015) who have observed an increase in SCC in the milk of goats affected with mastitis. It is the most widely used indicator for udder health in cow, sheep and goat milk. As compared to sheep and cow, SCC in goat milk is relatively high even in the healthy udder (Persson and Olofsson, 2011). However, Poutrel \u003cem\u003eet al\u003c/em\u003e. (1997) stated that elevated SCC is mainly a response to infection. Therefore, measurement of somatic cell count is a reliable indicator to detect mastitis and other intramammary infection in goats.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 02: Mean value of milk parameters in goats in different treatment groups at different intervals\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(n=6)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 246px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSCC\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 247px;\"\u003e\n \u003cp\u003e\u003cstrong\u003epH\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e19.48\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;2.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e4.75\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e7.49\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e6.65\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.03\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e19.40\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;3.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e4.37\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e7.40\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.08\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e6.62\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.08\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e3\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e19.60\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;3.58\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e3.80\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e7.47\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e6.61\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e03.70\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e3.72\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e6.60\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e6.60\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.04\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eMean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p\u0026lt;0.05)\u003c/p\u003e\n\u003cp\u003eThe results of present study are in accordance with Islam \u003cem\u003eet al\u003c/em\u003e. (2012), Hasan (2013) and Tomar \u003cem\u003eet al\u003c/em\u003e. (2018) who also reported higher pH of milk in goats affected with mastitis. The probable reason for increase in milk pH in mastitis may be the damage to the blood-milk barrier in mastitis leading to leakage of epithelial tight junctions. As a result blood and extracellular fluid components mix with secreted milk (Zhao and Lacasse, 2008) and showed the trends of increased milk pH with the increase in the severity of inflammatory process (Qayyum \u003cem\u003eet al\u003c/em\u003e., 2016). Thus, milk pH may serve as one of the good indicator to assess udder health and a difference in milk pH may indicate severity of intramammary infection. The improvement in milk pH after therapy in the present work may indicate recovery of animals after treatment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eOxidative stress parameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOxidative stress parameters i.e. malondialdehyde (MDA) and reduced glutathione (GSH) value were calculated in blood samples in all the 18 lactating goats under therapeutic trial on day 0 (pre-treatment) and day 6 (post treatment) and compared with that of healthy control goats.\u003c/p\u003e\n\u003cp\u003eMDA value on day 0 pre-treatment was significantly higher in goats of all treatment groups in comparison to healthy control group. A significant improvement in the MDA value of goats was noticed in all treatment groups on day 6 post treatment. However maximum improvement was seen in goats of group G3. The detail of MDA value in different treatment groups at different intervals are outlined in table 03. The findings of present study are in agreement with the findings of El-deeb, (2013), who reported significantly higher level of MDA with gangrenous mastitis in goats. Moreover, Kumar \u003cem\u003eet al\u003c/em\u003e. (2007) and Mahapatra \u003cem\u003eet al\u003c/em\u003e. (2018) also reported significant elevation of blood MDA level in clinical mastitis in bovines. Malondialdehyde is a main defined reaction product in biological system which undergoes lipid peroxidation and its level are the measures of the lipid peroxidation. The elevated level of MDA in the present study indicated a significant increase in lipid peroxidation as a result of the involvement of oxidative stress and the possible oxidative damage in clinical mastitis. The post treatment improvement in the level of MDA in all the goats indicated the effect of antioxidant used in the therapeutic regimen.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 03: Mean value of oxidative stress parameters in goats in different treatment groups at different intervals\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroups\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(n=6)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 246px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMDA\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 247px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGSH\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e(Mean\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDay 6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e18.85\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.98\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e8.99\u003csup\u003eaB\u003c/sup\u003e\u0026plusmn;1.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.06\u003csup\u003ebB\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.14\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e19.77\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;2.38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e7.06\u003csup\u003eabB\u003c/sup\u003e\u0026plusmn;1.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.07\u003csup\u003ebB\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.17\u003csup\u003eabA\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e3\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e20.61\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;1.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e5.97\u003csup\u003ebB\u003c/sup\u003e\u0026plusmn;0.63\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.07\u003csup\u003ebB\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.18\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003eG \u003csub\u003e4\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e5.29\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e5.44\u003csup\u003ebA\u003c/sup\u003e\u0026plusmn;0.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.18\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 123px;\"\u003e\n \u003cp\u003e0.18\u003csup\u003eaA\u003c/sup\u003e\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eMean values with different superscripts between group (lowercase) and between days (uppercase) differ significantly (p\u0026lt;0.05)\u003c/p\u003e\n\u003cp\u003eGSH value on day 0 pre-treatment was significantly lower in goats of all treatment groups in comparison to healthy control group. A significant improvement in the GSH value of goats was noticed in all treatment groups on day 6 post treatment. However maximum improvement was noticed in goats of group G3. The detail of GSH values in different treatment groups at different intervals are outlined in table 03. The results of present investigation correlates well with findings of Jhambh \u003cem\u003eet al\u003c/em\u003e. (2013) and El-deeb (2013) who reported significant decrease in GSH concentration in mastitic cow and does as compared to healthy ones. Glutathione, a thiol-containing tripeptide is present in living cells at high concentration in its reduced form (GSH). Upon reaction with reactive oxygen species, it got oxidized to glutathione radical which can be regenerated to its reduced form by glutathione reductase (Kohen and Nyska, 2002). A significant decrease in GSH concentration in goats during mastitis in the present study may be due to conversion of reduced form to oxidized form by excessive production of reactive oxygen species from inflamed gland (Jhambh \u003cem\u003eet al\u003c/em\u003e., 2013) and the improvement in its level might be due to the effect of therapy with antioxidants.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCorrelation between oxidative stress parameters and milk parameters\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA positive and significant correlation (P \u0026lt; 0.05) was observed between SCC and MDA levels. Thus, it was concluded that oxidative stress parameters, specifically MDA and GSH, were significantly altered in goats suffering from clinical mastitis, with a positive correlation observed with SCC.\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"616\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eParameter\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGSH\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMDA\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 146px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSCC\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSCC\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e-0.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e0.88*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 146px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMDA\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e-0.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 146px;\"\u003e\n \u003cp\u003e0.88*\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGSH\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 156px;\"\u003e\n \u003cp\u003e-0.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 146px;\"\u003e\n \u003cp\u003e-0.83\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e*Significant correlation between the parameters (P\u0026lt;0.05)\u003c/p\u003e\n\u003cp\u003eIn mastitic milk, the concentration of MDA is higher than in healthy milk, demonstrating that during infectious processes there is an accumulation of ROS and products resulting from oxidative degradation of biomolecules (Andrei \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2016). Furthermore, increased SCC may be correlated with an increase in MDA levels and a decrease in the proportion of unsaturated fatty acids, as studies conducted on bovine milk show (Guerrero, \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2015). Yehia \u003cem\u003eet al\u003c/em\u003e., (2024) also recorded the same finding and indicate that a significant correlation between different parameters in blood and milk can be useful as a proper clinical indicator for the early detection of mastitis in cows.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTherapeutic response evaluation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe response of therapy in different treatment group was evaluated on the basis of improvement in clinical abnormalities, milk parameters and oxidative stress markers on day 6 post treatment in goats of all the treatment groups.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe antibiotic used in all the treatment groups in the present study i.e. combination of amoxycilin and sulbactum @10 mg/kg b.wt. intramuscularly for 5 days was found effective in alleviating the mastitis. The efficacy of amoxicillin and sulbactum against mastitis is well documented by Roberson \u003cem\u003eet al\u003c/em\u003e. (2004) and Sharma \u003cem\u003eet al\u003c/em\u003e. (2020) as it possesses excellent antibacterial activity and has high promise for clinical application particularly in mastitis in livestock in small ruminants. Further, Constable \u003cem\u003eet al\u003c/em\u003e. (2007) stated that parenteral antimicrobial therapy should be considered in all cases of clinical mastitis and it must be used daily for 3 to 5 days. As multiple pathogens are responsible for causing mastitis. So, intramammary antimicrobial therapy is in common use for the treatment of certain causes of mastitis like streptococcus agalactia, which is very sensitive to intramammary therapy as compared to systemic therapy (Constable \u003cem\u003eet al\u003c/em\u003e., 2007).\u003c/p\u003e\n\u003cp\u003eMastitis is one such disease in which antioxidant defense system of body is compromised. Oxidative stress in the ailing animal occurs when the production of reactive oxygen metabolites exceed the capacity of antioxidant of the body. In case of clinical mastitis excessive reactive oxygen species production from the inflamed mammary gland occurs. The use of antioxidants viz. Ascorbic acid (in group G2) and Tocopherol and selenium (in group G3) in the present study ameliorated the oxidative stress in the mastitic goats.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAscorbic acid, used in the present study, is an important water soluble nonenzymatic antioxidants which can be synthesize in the body of goats. Goat\u0026rsquo;s milk contains on average 1.29 mg/100 g of ascorbic acid, more than cow\u0026rsquo;s milk (0.94 mg/100 g) (Park \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2007). The amount of ascorbic acid decreases during pathological processes, for example in mastitis (Andrei \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2016). Moreover, in milk containing a large number of somatic cells, the concentration of antioxidant vitamins is lower compared to milk with a low SCC (Andrei \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2016). Thus, in the case of subclinical mastitis, there is a pronounced reduction in the concentration of vitamin C; this is due to the fact that ascorbic acid is consumed during oxidative processes. In a healthy goat mammary gland, the concentration of vitamin C in the milk is higher by 36.4% compared to an infected udder tissue (Silanikove \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2014). A meager literature regarding use of ascorbic acid in goat mastitis is available. However, Klezkowski \u003cem\u003eet al\u003c/em\u003e. (2005) reported that cow suffering from mastitis have lower concentration of vitamin C in their milk and plasma. Ranjan \u003cem\u003eet al\u003c/em\u003e. (2005) have also reported the therapeutic value of vitamin C administered in cow with mastitis. Ascorbic acid has multidimensional bioactive role and its antioxidant role is to neutralize the toxic peroxides and reactive oxygen species released during cellular activation due to bacterial infection, which cause tissue injury (Mahapatra \u003cem\u003eet al\u003c/em\u003e., 2018).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTocopherol and selenium is a fat soluble antioxidant and has been shown to reduce the severity of mastitis by various scientists (Mukherjee, 2007 and Khangal \u003cem\u003eet al\u003c/em\u003e., 2015). Goat\u0026rsquo;s milk contains on average 0.04 mg/100 g of vitamin E, its concentration depending on diet, season or stage of lactation (Park \u003cem\u003eet al.,\u0026nbsp;\u003c/em\u003e2007). Vitamin E has an important role in lipid membrane protection from the attack of reactive oxygen species. It enhances the efficiency of neutrophils by protecting them from oxidative damage following intracellular killing of ingested bacteria (Yang and lee, 2015).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn the present study both Ascorbic acid (Vitamin C) and Tocopherol (Vitamin E) \u0026amp; selenium ameliorated the effect of oxidative stress during the mastitis in goats. The effect of vitamin C was moderate but vitamin E and selenium was proved as better antioxidant as compared to vitamin C because its role can be better understood on mastitis in a way that it\u0026rsquo;s act as lipid soluble antioxidant, free radical scavenger and protects against lipid peroxidation (Yang \u003cem\u003eet al\u003c/em\u003e., 2011).\u003c/p\u003e"},{"header":"CONCLUSION","content":"\u003cp\u003eA significant positive correlation (P \u0026lt; 0.05) was found between SCC and MDA levels indicating that oxidative stress is linked to the severity of mastitis. Enzymes like MDA, GSH and SOD are elevated in clinical mastitis and could serve as biomarkers for early detection of mastitis. These enzymes offer potential for real-time, online monitoring of mastitis in large dairy herds. Better diagnostics could lead to early mastitis detection, improving herd health, milk quality and overall dairy farm productivity. On the basis of clinical recovery, milk parameters and restoration of oxidative stress parameters, Tocopherol \u0026amp; Sodium selenite was proved as a better antioxidant in comparison of Ascorbic acid for the correction of oxidative stress produced by clinical mastitis in goats.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eACKNOWLEDGEMENT\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors are grateful to the Department of Veterinary Medicine, College of Veterinary Science and Animal Husbandry, Nanaji Deshmukh Veterinary Science University, Jabalpur, for providing the necessary facilities and support to carry out this research work. The authors also extend their sincere thanks to the livestock owners for their cooperation during sample collection.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSTATEMENT OF ANIMAL ETHICS\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll procedures involving animals were conducted in accordance with the guidelines of the Institutional Animal Ethics Committee (IAEC). The study protocol was reviewed and approved by the IAEC of Nanaji Deshmukh Veterinary Science University, Jabalpur, Madhya Pradesh, India (CPCSEA Registration No. 2071/GO/Re/S/19/CPCSEA). Adequate measures were taken to minimize animal discomfort and ensure humane handling during sample collection and clinical examination.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCONFLICT OF INTEREST STATEMENT\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFUNDING DECLARATION\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors received no financial support for the research, authorship, and/or publication of this article.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAndrei, S., Matei, S., Fit, N., Cernea, C., Ciupe, S., Bogdan, S. and Groza, I.S. (2011). Glutathione peroxidase activity and its relationship with somatic cell count, number of colony forming unit and protein content in subclinical mastitis cow\u0026rsquo;s milk. \u003cem\u003eRomanian Biotechnological Letters\u003c/em\u003e, \u003cstrong\u003e16\u003c/strong\u003e(3): 6209-6217.\u003c/li\u003e\n\u003cli\u003eAndrei, S., Pintea, A., Bunea, A., Groza, I., Bogdan, L., Ciupe, S. and Crainic, D. (2009). Non-Enzymatic Antioxidants Concentration and Lipids Peroxidation Level in Milk From Cows with Subclinical Mastitis. \u003cem\u003eBulletin of the University of Agricultural Sciences \u0026amp; Veterinary Medicine Cluj-Napoca. Veterinary Medicine\u003c/em\u003e, \u003cstrong\u003e66\u003c/strong\u003e(1):196-201.\u003c/li\u003e\n\u003cli\u003eAndrei, S., Matei, S., Rugina, D., Bogdan, L. and Ștefănuț, C. (2016). 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Mammary tissue damage during bovine mastitis: Causes and control. \u003cem\u003eJournal of Animal Science\u003c/em\u003e, \u003cstrong\u003e86\u003c/strong\u003e(13); 57-65.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","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":"Oxidative stress, Clinical Mastitis, Goats, MDA, GSH, Antioxidants","lastPublishedDoi":"10.21203/rs.3.rs-6999531/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6999531/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eMastitis remains a prevalent and economically significant disease in dairy goats. Oxidative stress, mediated by reactive oxygen species (ROS), is implicated in its pathogenesis, yet limited studies have explored this association in goats. This study aimed to assess oxidative stress biomarkers in caprine clinical mastitis, their correlation with somatic cell count (SCC), and the therapeutic efficacy of antioxidant interventions. A total of 313 lactating goats from both organized and unorganized sectors in Jabalpur were screened over six months, and 27 were diagnosed with clinical mastitis. Milk and heparinized blood samples were analyzed for SCC, milk pH, malondialdehyde (MDA), and reduced glutathione (GSH). Affected goats showed significantly elevated MDA, SCC, and milk pH, along with reduced GSH levels compared to healthy controls. A significant positive correlation (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05) was observed between SCC and MDA levels. Therapeutic trials compared the efficacy of ascorbic acid and tocopherol-sodium selenite combination alongside antibiotics. The tocopherol-selenium group demonstrated superior clinical recovery and normalization of oxidative stress markers. These results suggest that oxidative stress plays a pivotal role in clinical mastitis in goats and that antioxidant therapy, particularly tocopherol with selenium, can effectively mitigate oxidative damage when used with conventional treatment.\u003c/p\u003e","manuscriptTitle":"Oxidative Stress Biomarkers in Caprine Clinical Mastitis: Correlation with Somatic Cell Count and Evaluation of Antioxidant Therapy","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-08-08 10:51:16","doi":"10.21203/rs.3.rs-6999531/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":"c3aaf4ca-5323-4eb9-b961-f2729f22ca3b","owner":[],"postedDate":"August 8th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-08-23T08:23:42+00:00","versionOfRecord":[],"versionCreatedAt":"2025-08-08 10:51:16","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6999531","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6999531","identity":"rs-6999531","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}