Zoonotic threat of Proteus mirabilis in Xinjiang swine: antimicrobial resistance and virulence risks

Zoonotic threat of Proteus mirabilis in Xinjiang swine: antimicrobial resistance and virulence risks | 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 Zoonotic threat of Proteus mirabilis in Xinjiang swine: antimicrobial resistance and virulence risks Nana LI, Shenghui WAN, Yibo WANG, Yan LIANG, Yuwan LI, Pei Zheng, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6592294/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 Proteus mirabilis ( P. mirabilis ) is an opportunistic pathogen causing zoonotic diseases like diarrhea, keratitis, and urinary tract infections, sometimes leading to animal miscarriages. This study examined P. mirabilis prevalence in Xinjiang’s pig farms and pork, analyzing its drug resistance and pathogenicity to address food safety concerns. From 216 samples, 42 P. mirabilis strains were isolated (19.44%, 42/216), mostly from pig farms (95.24%, 40/42) rather than pork (4.76%, 2/42). All strains were multidrug-resistant, showing over 90.00% resistance to sulfonamides, tetracyclines, chloramphenicol, macrolides, polypeptides, and glycopeptides. Among them, 54.76% (23/42) produced extended-spectrum β-lactamases (ESBLs), enhancing resistance. The strains carried 19 resistance genes (7 categories), with CTX and OXA type ESBL related resistance genes exceeding 30.00% detection rate. Additionally, 95.24% (40/42) of the isolated strains carried class I integron genes. Nine virulence genes ( ureC , zapA , atfA , ucaA , pmfA , mrpA , r sbA , fliL , and hpmA ) were identified, and most strains (80.95%, 34/42) had strong biofilm-forming ability. These findings indicate that multidrug-resistant P. mirabilis in pigs, carrying diverse resistance and virulence genes, may spread via the food chain, posing health risks to consumers. Pigs Proteus mirabilis Antimicrobial resistance Virulence genes Pathogenicity Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Introduction Proteus mirabilis ( P. mirabilis ) is a Gram-negative bacterium with cell migration phenomenon, widely present in soil, water, and sewage [ 1 ]. It infects the digestive, respiratory, and urinary systems in humans and animals, with high infection rates in both populations [ 2 , 3 ]. Notably, it is a leading cause of urinary tract infections second only to Escherichia coli [ 4 ]. A major global concern is its resistance to β-lactam antibiotics due to extended-spectrum β-lactamases (ESBLs), which confer resistance to penicillin and cephalosporins. ESBLs mediated by plasmids can spread among different Enterobacteriaceae [ 5 ], and multidrug-resistant (MDR) P. mirabilis strains producing ESBLs and AmpC enzymes are increasing worldwide [ 6 ]. These strains can lead to clonal spread, causing outbreaks in hospitals and across regions [ 7 ]. Critically, infections caused by ESBL-producing P. mirabilis are associated with significantly higher rates of treatment failure and patient mortality [ 8 ]. Since animal-derived ESBL-producing P. mirabili threatens public health, this study isolated P. mirabilis strains from healthy pigs and pork to assess their antibiotics resistance, ESBL-producing, and genetic determinants (resistance/virulence genes). The findings provide insights into MDR mechanisms and zoonotic risks associated with porcine P. mirabilis . Methods Sample Collection Samples were collected from randomly selected healthy pigs in Xinjiang swine farms. Anal and nasal swabs were obtained by gentle rotation (2–3 times) using sterile swabs. Diseased pig lung tissues were also collected. All samples were labeled, stored at -4°C, and transported to the lab. Retail pork samples were collected from Shihezi morning markets. A total of 216 samples were obtained (108 farm samples: swabs and tissues; 108 pork samples). See Table 1 for details. Table 1 Sample source Breeding Type Sample Size Sample Type Swine farm A 23 Healthy swine nasal swab sample 19 Healthy swine anal swab sample Swine farm B 50 Healthy swine nasal swab sample Swine farm C 21 Healthy swine anal swab sample Swine farm D 24 Healthy swine anal swab sample Swine farm E 31 Infected swine lung tissue A fresh meat market in Shihezi City 48 Fresh meat in the market Experimental Animals Thirty 6-week-old Kunming mice were obtained from Xinjiang Medical University's Laboratory Animal Center for pathogenicity testing. All animal experiments described in this study were approved by the Biology Ethics Committee of Shihezi University (Ref. No. A2025-650) and strictly complied with the guidelines for the care and use of laboratory animals. The mice were euthanized by cervical dislocation performed by trained personnel. Prior to euthanasia, the mice were rendered unconscious via intraperitoneal injection of 1% sodium pentobarbital at 50 mg/kg dosage to ensure anesthesia before cervical dislocation. Bacterial Isolation and Gram staining examination Samples were inoculated into LB broth (5 mL, 37°C, 180 rpm, overnight) and plated on SS agar. Characteristic colonies (transparent periphery with black centers) were purified through 3–5 subcultures and confirmed by Gram staining. Biochemical Identification Isolates were identified using a biochemical test kit (Hangzhou Microbial Reagent Co., Hangzhou, China) following the manufacturer’s instructions. Hemolytic activity was assessed on 5% sheep blood agar (37°C, 24h). Molecular Confirmation Purified bacterial isolates were cultured in LB broth (5 mL, 180 rpm, 12–24 h), followed by genomic DNA extraction via boiling. The DNA was used for 16S rDNA amplification (primers in Table 2 ) under the following conditions: initial denaturation at 95°C for 5 min; 35 cycles of 95°C for 30 s, 58°C for 30 s, and 72°C for 90 s; with a final extension at 72°C for 10 min. In parallel, PCR was performed to amplify the P. mirabilis -specific pm0395 gene for confirming the presence of P. mirabilis , using primers listed in Table 2 . PCR products (5–6 µL) were electrophoresed on 1.0% agarose gels and visualized, then sent for sequencing (General Bio Ltd., Anhui). Sequences were analyzed by BLAST (NCBI database). Table 2 Primer information for specific genes Primer name Primer sequence (5 '-3') Fragment size (bp) Annealing temperature (℃) 16S rDNA F:AGAGTTTGATCCTGGCTCAG R:TACGGCTACCTTGTTACGACTT 1476 58 pm0395 F:ATCCAGTCACCACTAATCT R:TCTGACATCAACAGTAATTG 539 57 Antimicrobial susceptibility testing of P. mirabilis isolates The antibiotic susceptibility profiles of all isolated strains were determined using the Kirby-Bauer disk diffusion method on Mueller-Hinton agar (MHA) in accordance with CLSI guidelines, including phenotypic screening and confirmation tests for ESBL production [ 9 ]. The tested panel comprised 25 antibiotics from major classes: β-lactams (Cefotaxime/CTX 30 µg, Ceftazidime/CAZ 30 µg, Cephalosporin/CA 30 µg, Cefepime/CP 30 µg, Penicillin G/PG 10 µg, Amoxicillin/AM 10 µg, Ampicillin/AMP 10 µg), fluoroquinolones (Enrofloxacin/ENR 5 µg, Ciprofloxacin/CIP 5 µg, Norfloxacin/NF 5 µg), folate pathway inhibitors (Compound sulfamethoxazole/SXT 1.25/23.75 µg), tetracyclines (Tetracycline/TC 30 µg, Doxycycline/DOX 10 µg), aminoglycosides (Gentamicin/GN 10 µg, Kanamycin/KAN 30 µg, Amikacin/AMK 30 µg, Tobramycin/TOB 10 µg), phenicols (Fluphenicol/FFC 30 µg, Chloramphenicol/CL 30 µg), polymyxins (Polymyxin B/PB 30 µg), macrolides (Erythromycin/ERY 15 µg), glycopeptides (Vancomycin/VAN 30 µg), carbapenems (Imipenem/IMP 10 µg), and β-lactam/β-lactamase inhibitor combinations (Cefotaxime/clavulanic acid 10/10 µg, Ceftazidime/clavulanic acid 10/10 µg). Detection of antibiotic resistance genes in P. mirabilis isolates According to the references [ 10 – 12 ], the isolates were screened for major resistance genes including β-lactamases ( blaSHV , blaCMY , blaTEM , blaCIT , blaDHA , blaMOX , blaCTX-M-1/2/9 , blaOXA-1/48 ), sulfonamide ( sul1-3 ), tetracycline ( tetA/B ), quinolone ( qnrA/B/D/S ), chloramphenicol ( floR ), aminoglycoside [ acc(6')-1b-cr ], polymyxin ( vanA/B ), and macrolide resistance genes ( mefA/D ) using PCR with primers listed in Table 3 . The amplification protocol consisted of initial denaturation at 95°C (5 min), followed by 35 cycles of 95°C (30 s), gene-specific annealing (Table 3 , 30s), and 72°C extension (90 s), with final extension at 72°C (10 min). Table 3 Primers information of antibiotic resistance genes and integron genes Gene type Gene Name Primer sequence (5 '-3') Fragment size (bp) Annealing temperature (℃) β-lactams blaSHV F:AGCCGCTTGAGCAAATTAAAC R:ATCCCGCAGATAAATCACCAC 635 55 blaTEM F: CATTTCCGTGTCGCCCTTATTC R: CGTTCATCCATAGTTGCCTGAC 800 57 blaCIT F: CGAAGAGGCAATGACCAGAC R:ACGGACAGGGTTAGGATAGT 538 56 blaDHA F: TGATGGCACAGCAGGATATTC R: GCTTTGACTCTTTCGGTATTCG 997 55 blaMOX F: GCAACAACGACAATCCATCCT R:GGGATAGGCGTAACTCTCCCAA 895 57.5 blaCMY F:TGGCCGTTGCCGTTATCTAC R:CCCGTTTTATGCACCCATGA 868 55 blaCTX-M-1 F: TTAGGAARTGTGCCGCTGTA R: CGATATCGTTGGTGGTACCAT 688 54 blaCTX-M-2 F:CGTTAACGGCACGATGAC R:CGATATCGTTGGTGGTRCCAT 404 55 blaCTX-M-9 F:TCAAGCCTGCCGATCTGGT R:TGATTCTCGCCGCTGAAG 561 56 blaOXA-1 F:GACTTTATAAATTTAGTGTGTTTA R:TCCCAAGGGTTCCAGCA 816 55 blaOXA-48 F:GCTTGATCGCCCTCGATT R:GATTTGCTCCGTGGCCGAAA 281 56 Quinolones qnrA F:CCAGGATTTGAGTGACAGC R:GGCTCGTGTGTGCGGATG 592 69 qnrB F:GATCGTGAAAGCCAGAAAGG R:ATGAGCAACGATGCCTGGTA 476 65 qnrD F:CGAGATCAATTTACGGGGAATA R:TCTAAACCGTCGAGTTCGGCG 572 65 qnrS F:GCAAGTTCATTGAACAGGGT R:TGAAGGTCATCGATAGCAGG 428 65 Sulfonamide class sul1 F:CTTCGATGAGAGCCGGCGGC R:GCAAGGCGGAAACCCGCGCC 238 58 sul2 F:CGGCATCGTCAACATAACC R:GTGTGCGGATGAAGTCAG 722 60 sul3 F:AGATGTGATTGATTTGGGAGC R:TAGTTGTTTCTGGATTAGAGCCT 443 50 Tetracyclines tetA F:GCTACATCCTGCTTGCCTTC R:CATAGATCGCCGTGAAGAGG 210 63 tetB F:TGAAGGTCATCGATAGCAGG R:ATTTGCCGACTACCTTGGTG 391 55 Chloramphenicol class floR F:GGCTTTCGTCATTGCGTCTC R:ATCGGTAGGATGAAGGTGAGGA 650 60 Sminoglycosides Acc (6')—1b F:ATGACCTTGCGATGCTCTATGA R:CGAATGCCTGGCGTGTTT 486 54 Polymyxin vanA F:GTAGGCTGCGATATTCAAAGC R:CGATTCAATTGCGTAGTCCAA 231 56 vanB F:GTAGGCTGCGATATTCAAAGC R:GCCGACAATCAAATCATCCTC 330 56 Macrolides mefA F:AGTATCATTAATCACTAGTGC R:TTCTTCTGGTACTAAAAGT 348 53 mrsD F:GCCTTCCGGAGCTCCTACTT R:GCGTCCAATGTATCTCTAT 500 53 Integron intI1 F:ACGAGCGCAAGGTTTCGGT R:GAAAGGTCTGGTCATACATG 525 56 intI2 F:CACGGATATGCGACAAAAAGGT R:GTAGCAAACGACTGACGACAAAAAGGT 789 56 intI3 F:GCCTCCGGCAGCGACTTTCAG R:GATGCTGCCCAGGGCGCTCG 433 65 Detection of integrations in P. mirabilis isolates Integrations play an important role in the transmission of bacterial resistance. PCR amplification was performed to detect class I, II, and III integrons using published primer sequences [ 13 ] (see Table 3 for details). Detection of virulence genes in P. mirabilis isolates PCR amplification using published primers [ 14 ] (Table 4 ) was performed to detect 10 key virulence genes ( ureC , mrpA , rsbA , atfC , ucAA , pmfA , atfA , zapA , fⅠiL , and hpmA ) in the isolates, with reaction conditions following established protocols. Table 4 Primer sequence of virulence gene Gene Type Gene Name Primer sequence (5 '-3') Fragment size (bp) Urease ureC F:GTTATTCGTGATGGGATGGG R:ATAAAGGTGGTTACGCCAG 375 Mannose-resistant-like fimbriae mrpA F:ACACCTGCCCATATGGAAGATACTGGTACA R:AAGTGATGAAGCTTAGTGATGGTGATGGTG 550 Migration-ability-regulating gene rsbA F:TCGATTTCAGTGTTTGGCCAT R:TCGATTTCAGTGTTTGGCCAT 1647 Thermophilic fimbriae atfA F:CATAATTTCTAGACCTGCCCTAGCA R:CTGCTTGGATCCGTAATTTTTAACG 332 atfC F:AGAAAGGGATCCTACAAATTAA R:TATAGCATGCATTTAAATTGCC 472 Uroepithelial cell adhesin ucaA F:GTAAAGTTGTTGCGCAAAC R:TTGAGCCACTGTGGATACA 587 Fimbrial gene pmfA F:GGATCATCTATAATGAAACTG R:CTGATAATCAACTTGGAAGTT 512 Metalloproteinase zapA F:ACCGCAGGAAAACATATAGCCC R:GCGACTATCTTCCGCATAATCA 543 Flagellar gene fⅠiL F:CTCTGCTCGTGGTGGTGTCG R:GCGTCGTCACCTGATGTGTC 770 Hemolysin hpmA F:GTTGAGGGGCGTTATCAAGAGTC R:GATAACTGTTTTGCCCTTTTGTGC 709 Biofilm formation ability assay of P. mirabilis isolates The biofilm-forming capacity of isolates was determined using a 96-well microtiter plate assay [ 15 – 16 ]. Briefly, 180 µL LB medium and 20 µL bacterial culture were added per well, incubated at 37°C for 24 h, washed with PBS, and fixed with methanol (200 µL, 15 min). After crystal violet staining (1%, 5 min) and acetic acid dissolution (33%, 30 min), absorbance was measured at 590 nm. Biofilm formation was categorized as: non (OD ≤ negative control), weak (OD ≤ 2× control), moderate (OD ≤ 4× control), or strong (OD > 4× control), with triplicate wells per strain. Mouse pathogenicity assay The multidrug-resistant, ESBL-producing P. mirabilis strain Pm-07 (carrying 6 virulence and 8 resistance genes) was selected for challenge. Thirty 6-week-old Kunming mice were randomized into six groups (n = 5/group). After 24-hour fasting, mice were intraperitoneally injected with 0.5 mL of 10-fold serially diluted bacterial suspensions (2.6×10⁴–2.6×10⁹ CFU), while controls received sterile saline. Clinical signs and mortality were monitored every 4 hours. LD₅₀ was calculated using the modified Karber method. Deceased mice underwent necropsy for lesion evaluation, bacterial re-isolation (confirmed by PCR), and histopathology (4% formaldehyde-fixed, paraffin-embedded, H&E-stained tissues). Results Isolation and identification of P. mirabilis A total of 42 P. mirabilis strains (19.44% isolation rate) were obtained from 216 samples (168 swine swabs and 48 pork/lung tissues) collected from five swine farms, with varying isolation rates across farms (11.90-35.48%) and lower detection in pork (4.16%) (Fig. 1). The isolates exhibited characteristic migration on BHI agar (> 18h incubation) and typical black-centered colonies on SS agar, with Gram-negative rod morphology. Biochemical profiling confirmed xylose fermentation, urea/ornithine/lysine decomposition, H₂S production, and β-hemolysis on 5% sheep blood agar, while testing negative for cottonseed sugar fermentation and other markers (Fig. 2). Molecular identification via 16S rDNA sequencing revealed 99.65–100% homology with known P. mirabilis strains among the 42 isolates. The associated sequence datasets generated in this study have been deposited in the GenBank repository under accession numbers PV686792 to PV686828 and PV700941 to PV700945. Species-specific PCR consistently amplified the expected 539 bp fragment, conclusively identifying all isolates as P. mirabilis (Fig. 3). Antimicrobial resistance profile of P. mirabilis isolates All 42 P. mirablis isolates exhibited multidrug resistance, demonstrating > 90% resistance to sulfonamides, tetracyclines, chloramphenicol, macrolides, polypeptides, and glycopeptides. Resistance rates varied by class: β-lactams (ampicillin 69.05%, penicillin 88.10%, amoxicillin 92.86%), quinolones (59.52–85.71%), and aminoglycosides (gentamicin 50.00%, tobramycin 78.57%, kanamycin 80.95%), while showing relative sensitivity to amikacin (21.43%) and cefepime (9.52%) (Fig. 4A, B). Among these, 54.76% (23/42) were ESBL-producers, displaying broader resistance (11 antibiotics across 8 classes) compared to non-ESBL strains (6 antibiotics in 5 classes), confirming ESBL production significantly enhances resistance (Fig. 4C, D). All isolates were confirmed as multidrug-resistant, highlighting widespread resistance in porcine P. mirabilis . Antibiotic resistance gene profile of P. mirabilis isolates PCR screening of 42 P. mirabilis isolates identified 19 resistance genes across 7 classes, with β-lactamase genes being most prevalent (83.33% ESBL-carriers). Key findings include: universal presence of sul1 (100%, sulfonamide resistance) and high prevalence of floR (85.71%, chloramphenicol), aac (6')-Ib-cr (73.81%, aminoglycoside), and CTX-M-type ESBLs (85.71%, including CTX-M-2 (45.24%), CTX-M-9 (35.71%), and CTX-M-1 (33.33%). Notably, 54.76% isolates carried ≥ 3 ESBL genotypes, while qnrD (33.33%, quinolone) and tetA (38.10%, tetracycline) showed significant detection rates. No polymyxin resistance genes or TEM genotypes were detected, highlighting specific resistance patterns in these porcine isolates (Fig. 5). Correlation between phenotypic and genotypic resistance in P. mirabilis isolates The consistency between the resistance phenotype and resistance genotype of the P. mirabilis isolates is shown in Table 5. The 42 P. mirabilis isolates demonstrated varying concordance rates between resistance genes and phenotypes: high for sulfonamides (100.00%, 42/42), chloramphenicols (85.71%, 36/42), β-lactams (83.33%, 35/42), aminoglycosides (73.81%, 31/42), quinolones (54.76%, 23/42), and tetracyclines (40.48%, 17/42), but markedly lower for macrolides (26.19%,11/42) and polymyxins (0.00%, 0/42). Table 5 Concordance between genotypic and phenotypic resistance profiles in P. mirabilis isolates Category of antibiotics Number of drug-resistant bacteria Number of bacterial strains containing resistance genes Drug resistance compliance rate (%) β-lactams 42 35 83.33% Quinolones 42 23 54.76% Sulfonamide class 42 42 100.00% Tetracyclines 42 17 40.48% Chloramphenicol class 42 36 85.71% Aminoglycosides 42 31 73.81% Polymyxin 42 0 0.00% Macrolides 42 11 26.19% Integron and virulence gene distribution in P. mirabilis Isolates Among 42 P. mirabilis isolates, class I integrons were predominant (95.2%), while class II (2.4%) and III (9.5%) were rare. Virulence gene profiling revealed near-universal presence of fliL (97.6%), ureC (92.9%), atfA (92.9%), and zapA (90.5%), with lower frequencies for hpmA (88.1%), pmfA (76.2%), ucaA (47.6%), mrpA (28.6%), and rsbA (23.8%). Notably, atfC was absent in all isolates, demonstrating a conserved virulence gene profile with specific exceptions. Virulence gene profile of P. mirabilis isolates All 42 P. mirabilis isolates carried varying combinations of nine virulence genes, with near-ubiquitous prevalence of four core virulence factors: flagellar gene fliL (97.6%), urease gene ureC (92.9%), thermoregulated fimbriae gene atfA (92.9%), and metalloproteinase gene zapA (90.5%). Secondary virulence determinants included hemolysin hpmA (88.1%) and fimbrial genes pmfA (76.2%) and ucaA (47.6%), while mannose-resistant fimbriae mrpA (28.6%) and motility regulator rsbA (23.8%) were less common. Notably, atfC was absent in all isolates, revealing a conserved virulence gene pattern with distinct prevalence tiers among clinical strains(Fig. 6). Biofilm formation capacity of P. mirabilis isolates The biofilm-forming capacity of 42 P. mirabilis isolates was quantitatively assessed via microtiter plate assay (OD₅₉₀), revealing three distinct tiers: 80.95% (34/42) exhibited strong biofilm production (OD > 4×ODc),14.29% (6/42) showed moderate formation (2×ODc < OD ≤ 4×ODc), while only 4.76% (2/42) pork-derived isolates demonstrated weak capability (1×ODc < OD ≤ 2×ODc). (Fig. 7). Statistical correlation between phenotypic resistance patterns and genotypic resistance determinants for β-lactam antibiotics Correlation analysis was performed to assess the relationship between β-lactam antibiotic resistance phenotypes and resistance genes in 42 P. mirabilis strains, including both ESBL-producing and non-ESBL-producing isolates. As demonstrated in Fig. 8, the overall correlation between resistance phenotypes and resistance genes to β-lactamase antibiotics was not statistically significant. Among ESBL-producing strains, phenotypic resistance showed a marginal positive correlation with the blaSHV gene (Pearson's r = 0.36) and a slight negative correlation with the blaCMY gene (r = -0.31). In contrast, non-ESBL-producing strains exhibited weak negative correlations with both blaSHV (r = -0.36) and blaCMY (r = -0.31). No significant correlations were observed between resistance phenotypes and other β-lactamase resistance genes in either group. Correlation analysis of virulence genes and biofilm formation in P. mirabilis Biofilm formation and virulence gene profiles were analyzed in 42 clinical P. mirabilis isolates to assess their potential association. As shown in Fig. 9, most virulence genes demonstrated weak correlations with biofilm-forming capacity. Notably, only the temperature-adapted fimbrial gene ( atfA ) showed a modest positive correlation (r = 0.40) with biofilm formation, suggesting a potential role in bacterial adhesion and biofilm development. Mouse pathogenicity test Comprehensive characterization of 42 P. mirabilis isolates identified strain Pm-07 for murine pathogenicity testing. Intraperitoneal inoculation (2.6×10⁸ CFU/mL) induced clinical signs (lethargy, anorexia, diarrhea) within 6 hours, with mortality commencing at 24 hours. Necropsy revealed significant organ enlargement (lungs, liver, spleen, kidneys) with hemorrhaging and duodenal wall thinning (Fig. 10). PCR confirmed bacterial re-isolation from affected tissues (Fig. 11), with calculated LD₅₀ = 7.05×10⁷ CFU/mL. Histopathology demonstrated alveolar hyperplasia, hepatic disorganization, splenic inflammatory infiltration, glomerular congestion, and intestinal villi damage with neutrophil recruitment (Fig. 12). Discussion P. mirabilis , along with Klebsiella pneumoniae and Escherichia coli , ranks among the most clinically significant Enterobacteriaceae responsible for both nosocomial and community-acquired infections globally [ 17 ]. Our study identified P. mirabilis in Xinjiang farming environments with an overall prevalence of 19.4%, showing notable farm-to-farm variation. These findings complement existing epidemiological data demonstrating variable detection rates across China - from 28.9% in Xinjiang sheep farms [ 18 ] to 7.07% in Shandong poultry operations [ 19 ]. The observed geographical disparities in prevalence likely reflect regional differences in animal husbandry practices and environmental conditions, while the consistent detection across diverse livestock species underscores P. mirabilis 's adaptability and zoonotic potential. The escalating use of broad-spectrum antibiotics has precipitated a concerning rise in antimicrobial resistance among Proteus mirabilis isolates, particularly with the increasing prevalence of ESBL-producing strains that substantially complicate treatment regimens [ 20 – 21 ]. Our study of porcine-derived P. mirabilis isolates demonstrated alarming resistance profiles, exhibiting > 90% resistance to 10 antimicrobial agents spanning 7 classes, with retained susceptibility only to amikacin (aminoglycoside) and cefepime (4th-generation cephalosporin). All isolates exhibited multidrug resistance (MDR) to ≥ 5 antibiotic classes, mirroring Sun et al.'s [ 22 ] findings of 100% resistance to 6 antibiotic classes in bovine-derived strains, which remained sensitive solely to ceftazidime, gentamicin and amikacin. This convergence of severe MDR patterns across livestock species and geographical regions signals an urgent need for comprehensive antibiotic stewardship programs in animal agriculture, enhanced surveillance of resistance gene transmission, and development of novel therapeutic strategies to address this critical public health challenge. β-lactamase genes represent a major mechanism of bacterial resistance. Studies reveal varying prevalence of these genes: Sun [ 22 ] reported blaCTX-M (45.1%), blaTEM (13.7%), and blaSHV (21.6%) in clinical isolates, while Cao [ 12 ] found higher rates in fur animals ( blaTEM 83.0%, blaOXA-1 60.4%). Our study detected blaCTX-M-2 in 45.2% of isolates-higher than South African poultry (38%) [ 23 ] but showing distinct regional patterns, with CTX-M variants demonstrating geographical predominance ( CTX-M-2 in Latin America [ 24 ] vs CTX-M-1/9 in other regions [ 11 ]). Notably, ESBL-producing strains showed weak correlations with specific genes ( blaSHV : r = 0.36; blaCMY : r=-0.31), contrasting with Liu's findings of strong cefotaxime- blaCTX-M correlations (r = 0.95-1) in canine isolates. This variability likely reflects complex interactions among β-lactamase genes, where compensatory mechanisms may obscure individual gene effects on resistance phenotypes [ 25 ]. Integrons play a critical role in the development of multidrug resistance (MDR) through their ability to acquire, integrate, and express multiple antibiotic resistance gene cassettes, with over 80 such cassettes identified in class I integrons alone. The high prevalence of integrons across different P. mirabilis strains is particularly concerning, as demonstrated by Shi's finding of 100% class I integron carriage in porcine isolates [ 26 ], Chen's report of 60.7% integron positivity in human clinical strains (including 20% class I, 14.7% class II, and 26% carrying both) [ 27 ], and our observation of 95.2% class I integron prevalence with 9.5% co-carriage of multiple integron types. These findings collectively highlight integrons as key vectors for resistance gene dissemination and MDR development [ 28 ], emphasizing the urgent need for enhanced resistance surveillance and stricter antibiotic stewardship in both agricultural and clinical settings to curb the spread of these mobile genetic elements. The P. mirabilis isolates carried multiple virulence genes (ureC, pmfA, zapA, fliL, atfA, hpmA, ucaA), with fliL (97.62%, 41/42) and ureC (92.86%, 39/42) being most prevalent. The fliL gene encodes an ~ 18kDa transmembrane protein near the flagellar basal body [ 29 ], crucial for bacterial motility [ 30 ]. Similar to Qu et al.'s findings [ 31 ], which identified 20 flagellar virulence factors (including flgI , fliM , flgC ) in all 30 strains, and consistent with Chengdu studies showing high ureC prevalence in pets [ 10 ]. Biofilm formation, a key virulence factor [ 32 ], was observed in all isolates: 80.95% (34/42) strong, 14.29% (6/42) moderate, and 4.76% (2/42) weak producers. This aligns with Jia's report of universal biofilm formation in chicken-derived strains (15.38% strong) [ 33 ] and Sun's findings in diarrheic animals (92.05% producers) [ 3 ]. Biofilm-positive strains showed higher pathogenicity, significantly correlating with ureC , zapA , rsmA , hmpA , mrpA , atfA , and pmfA ( P < 0.05). Specifically, biofilm formation weakly correlated with atfA (r = 0.40). Further research should investigate strain-specific biofilm-virulence interactions to optimize clinical management. Conclusion This study reveals that swine-derived P. mirabilis strains are multidrug-resistant, carrying various resistance and virulence genes, with 80.95% exhibiting strong biofilm formation. These findings highlight significant zoonotic risks through potential food chain transmission, calling for improved antimicrobial stewardship and surveillance in livestock production to mitigate public health threats. Abbreviations P. mirabilis , Proteus mirabilis ; PEN, Penicillin; AMX, Amoxicillin; CFB, Cefazolin; CTX, Cefotaxime; K, Kanamycin; GM, Gentamicin; ENR, Enrofloxacin; CIP, Ciprofloxacin; DX, Doxycycline; TMP, Trimethoprim-sulfamethoxazole; FON, Florfenicol; PB, Polymyxin B; AMP, Ampicillin; IPM, Imipenem; TCY, Tetracycline; CRO, Ceftriaxone; CA, Vancomycin Declarations Acknowledgments The authors wish to express their gratitude to the College of Animal Science and Technology, Xinjiang Tecon Animal Husbandry Technology Co., Ltd., for facilitating this study. Funding This research was funded by the Natural Science Support Program Project of the Xinjiang Production and Construction Corps (2024DA007) and the Shihezi University Technology Transfer and Promotion Program (CGZH202309). The APC was funded by the Natural Science Support Program Project of the Xinjiang Production and Construction Corps (2024DA007). Author information Nana LI, Shenghui WAN and Yibo WANG contributed equally to this work. Authors and Affiliations 1 College of Animal Science and Technology，Shihezi University, Shihezi 832003, China Yonggang QU, Gaoming HE, Yanfang Li, Yan LIANG, Nana LI, Shenghui WAN 2 College of Food and Bioengineering, Henan University of Science and Technology Yuwan LI, Yibo WANG 3 Xinjiang Tecon Animal Husbandry Technology Co., Ltd., Changji 831399, China Pei ZHENG Contributions Writing-original draft preparation, Nana Li. Shenghui Wan; the acquisition, analysis, Yibo WANG; resources, Pei Zheng; review and editing, Yuwan LI;review and editing, Yanfang Li; resources, Gaoming HE;resources, Yan Liang; Supervision, Yonggang Qu; Project administration, Yonggang Qu; Funding acquisition, Yonggang Qu. All the authors have read and agreed to the published version of the manuscript. Data availability The original contributions presented in the study are included in the article, further inquiries can be directed to the corresponding author after reasonable request. Ethics approval and consent to participate This study received ethical approval from the Biology Ethics Committee of Shihezi University (Approval No. A2025-650). Written informed consent was obtained from all animal owners prior to their participation in the research. Consent for publication Not Applicable. Competing interests The authors declare no competing financial interests. References Adeolu M, Alnajar S, Naushad S, Gupta S. Genome-based phylogeny and taxonomy of the 'Enterobacteriales': proposal for Enterobacterales ord. nov. divided into the families Enterobacteriaceae, Erwiniaceae fam. nov., Pectobacteriaceae fam. nov., Yersiniaceae fam. nov., Hafniaceae fam. nov., Morganellaceae fam. nov., and Budviciaceae fam. nov. Int J Syst Evol Microbiol. 2016;66(12):5575–99. Wang JT, Chen PC, Chang SC, Shiau YR, Wang HY, Lai JF, et al. Antimicrobial susceptibilities of Proteus mirabilis: a longitudinal nationwide study from the Taiwan surveillance of antimicrobial resistance (TSAR) program. BMC Infect Dis. 2014;14:486. Sun Y, Wen S, Zhao L, Xia Q, Pan Y, Liu H, et al. Association among biofilm formation, virulence gene expression, and antibiotic resistance in Proteus mirabilis isolates from diarrhetic animals in Northeast China. BMC Vet Res. 2020;16(1):176. Kwiecińska-Piróg J, Skowron K, Bogiel T, Białucha A, Przekwas J, Gospodarek-Komkowska E. 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Isolation, identification, virulence gene detection, and drug resistance analysis of Escherichia coli and Proteus mirabilis in calf diarrhea. Xianyang: Northwest A&F University; 2023. Jin H, Zhou R, Kang M, Luo R, Cai X, Chen H. Biofilm formation by field isolates and reference strains of Haemophilus parasuis. Vet Microbiol. 2006;118(1–2):117–23. Li Nana Y, Xingyu HG, Mingzhu L, Yang G, Yaqi Z, Pei, et al. Isolation, identification, drug resistance, and pathogenicity analysis of Escherichia coli from pigs. Chin Anim Husb veterinary Med. 2025;52(01):376–88. Abreu AG, Marques SG, Monteiro-Neto V, Carvalho RM, Gonçalves AG. Nosocomial infection and characterization of extended-spectrum β-lactamases-producing Enterobacteriaceae in Northeast Brazil. Rev Soc Bras Med Trop. 2011;44(4):441–6. Xiaojuan M, Xiaoxiao W, Jiaxin H, Zhaoran Y, Sheripu G, Jianlong L, et al. Wait Investigation of sheep diarrhea in some areas of Xinjiang and isolation and identification of pathogenic bacteria. Adv Vet Med. 2024;45(10):109–13. Li Z, Peng C, Zhang G, Shen Y, Zhang Y, Liu C, et al. Prevalence and characteristics of multidrug-resistant Proteus mirabilis from broiler farms in Shandong Province, China. Poult Sci. 2022;101(4):101710. Girlich D, Bonnin RA, Dortet L, Naas T. Genetics of Acquired Antibiotic Resistance Genes in Proteus spp. Front Microbiol. 2020;11:256. Sherchan JB, Hayakawa K, Miyoshi-Akiyama T, Ohmagari N, Kirikae T, Nagamatsu M, et al. Clinical epidemiology and molecular analysis of extended-spectrum-β-lactamase-producing Escherichia coli in Nepal: characteristics of sequence types 131 and 648. Antimicrob Agents Chemother. 2015;59(6):3424–32. Sun Shihao.Isolation, identification, and analysis of drug resistance and virulence genes of Xinjiang cattle derived Proteus mirabilis. Alar: Tarim University. 2021. Ramatla T, Ramaili T, Lekota K, Mileng K, Ndou R, Mphuthi M, et al. Antibiotic resistance and virulence profiles of Proteus mirabilis isolated from broiler chickens at abattoir in South Africa. Vet Med Sci. 2024;10(2):e1371. Sennati S, Santella G, Di Conza J, Pallecchi L, Pino M, Ghiglione B, et al. Changing epidemiology of extended-spectrum β-lactamases in Argentina: emergence of CTX-M-15. Antimicrob Agents Chemother. 2012;56(11):6003–5. Shi Letian. Study on the resistance characteristics of avian Escherichia coli and Klebsiella pneumoniae producing extended spectrum β-lactase. Zhengzhou: Henan Agricultural University; 2024. Shi Baoqiu Z, Di G, Shuo Z, Xiaoliang J, Jingliang Y, Guangfu, et al. Isolation and identification of Proteus mirabilis from pigs and detection of class I integrons. Chin J Anim Infect Dis. 2020;28(2):7–13. Chen Keda L, Qiaoping QW, Yanye T. Goryeo. Analysis of Homozygous Distribution and Drug Resistance in Proteus mirabilis. Chin J Antibiot. 2020;45(11):1148–52. Odumosu BT, Adeniyi BA, Chandra R. Analysis of integrons and associated gene cassettes in clinical isolates of multidrug resistant Pseudomonas aeruginosa from Southwest Nigeria. Ann Clin Microbiol Antimicrob. 2013;12:29. Lee YY, Belas R. Loss of FliL alters Proteus mirabilis surface sensing and temperature-dependent swarming. J Bacteriol. 2015;197(1):159–73. Attmannspacher U, Scharf BE, Harshey RM. FliL is essential for swarming: motor rotation in absence of FliL fractures the flagellar rod in swarmer cells of Salmonella enterica. Mol Microbiol. 2008;68(2):328–41. Qu X, Zhou J, Huang H, Wang W, Xiao Y, Tang B, et al. Genomic Investigation of Proteus mirabilis Isolates Recovered From Pig Farms in Zhejiang Province, China. Front Microbiol. 2022;13:952982. Fusco A, Coretti L, Savio V, Buommino E, Lembo F, Donnarumma G. Biofilm Formation and Immunomodulatory Activity of Proteus mirabilis Clinically Isolated Strains. Int J Mol Sci. 2017;18(2):414. Jia Yuanzheng. Biological characteristics of chicken derived Proteus mirabilis and its LuxS/AI-2 quorum sensing system. Yangzhou University; 2021. Additional Declarations No competing interests reported. Supplementary Files SupplementaryTableS1.xls Supplementaryfilefigure3.docx 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. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-6592294","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":473329731,"identity":"a0a8cfc9-3282-4da1-8863-9b54271fc4f5","order_by":0,"name":"Nana LI","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Nana","middleName":"","lastName":"LI","suffix":""},{"id":473329732,"identity":"67af14e1-9e00-4d84-b494-8e4b61882356","order_by":1,"name":"Shenghui WAN","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Shenghui","middleName":"","lastName":"WAN","suffix":""},{"id":473329733,"identity":"f227ffb8-4080-41df-bcf2-aec0026167ba","order_by":2,"name":"Yibo WANG","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Yibo","middleName":"","lastName":"WANG","suffix":""},{"id":473329734,"identity":"30b45744-0d8d-42ea-a15e-b52ea4da29cb","order_by":3,"name":"Yan LIANG","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Yan","middleName":"","lastName":"LIANG","suffix":""},{"id":473329735,"identity":"82766a06-8e5a-4dca-ac2f-4207ce872e17","order_by":4,"name":"Yuwan LI","email":"","orcid":"","institution":"Henan University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Yuwan","middleName":"","lastName":"LI","suffix":""},{"id":473329737,"identity":"d2578c67-be6e-4ea7-af2f-e8ebc25f3b48","order_by":5,"name":"Pei Zheng","email":"","orcid":"","institution":"Xinjiang Tecon Animal Husbandry Technology Co., Ltd","correspondingAuthor":false,"prefix":"","firstName":"Pei","middleName":"","lastName":"Zheng","suffix":""},{"id":473329738,"identity":"9b369da9-adca-46cf-9ef3-9ee57806d135","order_by":6,"name":"Yanfang Li","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Yanfang","middleName":"","lastName":"Li","suffix":""},{"id":473329739,"identity":"49cc3a43-f4ce-48ef-908c-68efaf2079f0","order_by":7,"name":"Yonggang QU","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA8UlEQVRIiWNgGAWjYHAD5gPMEEYC0VrYEkjWwmNAnBa+GzmGjwt+1cqb86/5/Lngz2EGfvYcA4afO3BrkbyRY2w8s++44c4Zb7dJz+A5zCDZ88aAsfcMbi0GN3LMpHl7jjFuuHF2GzOPxGGQiAEzYxthLfYbbpx5/JnH4DCDPVFaeH7UJG4438MgzZMAtEWCgBbJM8+KjXkbDiRvuMEG1HsgnUfizLOCg714tPAdT974mOdPne2G84eBDvtjLcffnrzxwU88WhgOcBgwMLYdZmCQSADzecCCeDQAZdkfMDD8qWNg4MevbhSMglEwCkYwAACcdVcmtmtVWwAAAABJRU5ErkJggg==","orcid":"","institution":"Shihezi University","correspondingAuthor":true,"prefix":"","firstName":"Yonggang","middleName":"","lastName":"QU","suffix":""},{"id":473329740,"identity":"453b3184-b717-41b4-ba2b-1739cf86ba7f","order_by":8,"name":"Gaoming HE","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Gaoming","middleName":"","lastName":"HE","suffix":""}],"badges":[],"createdAt":"2025-05-05 07:53:25","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6592294/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6592294/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":85140333,"identity":"bf706456-52a9-4314-ac51-e2b11a2ea7ce","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":156564,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSpatial epidemiology of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eProteus mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e in Xinjiang swine production systems\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote: \u003c/strong\u003eSampling locations are marked by proportional circles indicating \u003cem\u003eP. mirabilis\u003c/em\u003e isolation rates (range: 9.5-36.0%), with farm sites (A-E) shown as filled green triangles and retail markets as red circles. The size of the circle represents the high or low detection rate of \u003cem\u003eP. mirabilis\u003c/em\u003e. Key findings: (i) Higher prevalence in pig farms (B, E) versus pig farms (A, C, D), (ii) Minimal contamination in retail pork (4.2%).\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/d1e4d7ab2a79038b8ddafb20.png"},{"id":85140336,"identity":"0b0b9562-fbad-4079-9603-d2518636b0b5","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":321440,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMorphological and phenotypic characterization of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote: \u003c/strong\u003e\u0026nbsp;(A) Swarming motility on LB agar showing characteristic concentric growth rings (18 h incubation); (B) Typical colony morphology was observed on SS agar, and 5 × local magnification showed colonies with black edges in the center and transparent edges; (C) Gram-stained microscopy (1000×) revealing Gram-negative rods with rounded ends; (D) β-hemolysis on 5% sheep blood agar, indicated by complete clearing around colonies. Scale bars: 2 mm (A, B, D), 5 μm (C).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/17d0453eece923f8daa5aa9c.png"},{"id":85140643,"identity":"de9b8648-8de0-4f16-a406-3012e04e0e54","added_by":"auto","created_at":"2025-06-22 10:00:37","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":149805,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMolecular confirmation of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eProteus mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates by PM gene PCR amplification\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote: \u003c/strong\u003eAgarose gel (1.0%) electrophoresis of PM gene PCR products (expected size: 539 bp). Lanes: M, DL2000 DNA marker; 1-19, representative \u003cem\u003eP. mirabilis\u003c/em\u003e isolates; N, negative control (no template). All test strains showed the target amplicon, confirming their identity as \u003cem\u003eP. mirabilis\u003c/em\u003e. It should be noted that the positive control was unfortunately missing in this gel run. However, the negative control remained blank, demonstrating assay specificity.\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/2907da1643118f2aeee75958.png"},{"id":85140657,"identity":"a099ecce-4e46-47b3-94f0-7d3fbb7f70b5","added_by":"auto","created_at":"2025-06-22 10:00:39","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":116582,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eComprehensive antimicrobial resistance profile of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote: \u003c/strong\u003eA: Intuitively display the overall antimicrobial resistance of all isolated strains to different types of antibiotics, with color gradient indicating resistance intensity (red: resistant, blue: sensitive). B: Quantitative resistance rates (%) categorized by antibiotic class, demonstrating: (i) \u0026gt;90% resistance to sulfonamides, tetracyclines, and chloramphenicol; (ii) \u0026lt;25% resistance to cefepime and amikacin. ESBL-producing strains (n=23) showed significantly broader resistance spectra compared to non-ESBL strains (n=19), particularly against β-lactams (p\u0026lt;0.01). C: This histogram shows the resistance rates of ESBL-producing and non-ESBL-producing strains to different antibiotics (blue: ESBL strains; red: non-ESBL-producing strains). D: The histogram showed the distribution of the number of multi-drug resistant (MDR) isolates, and the results showed that all isolates were multi-drug resistant (MDR) strains.\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/96a88fba903140532cad6f74.png"},{"id":85140341,"identity":"c5e05ade-96fc-459c-8207-0ffcfbac7b42","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":13053,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eDistribution of antimicrobial resistance genes and integrons in \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCorrelation between phenotypic and genotypic resistance in \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe consistency between the resistance phenotype and resistance genotype of the \u003cem\u003eP. mirabilis\u003c/em\u003e isolates is shown in Table 5. The 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates demonstrated varying concordance rates between resistance genes and phenotypes: high for sulfonamides (100.00%, 42/42), chloramphenicols (85.71%, 36/42), β-lactams (83.33%, 35/42), aminoglycosides (73.81%, 31/42), quinolones (54.76%, 23/42), and tetracyclines (40.48%, 17/42), but markedly lower for macrolides (26.19%,11/42) and polymyxins (0.00%, 0/42).\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/ba48fcd08b737e8f271bcc8f.png"},{"id":85140349,"identity":"0307824f-355c-4be4-be5c-eef222bd2f79","added_by":"auto","created_at":"2025-06-22 09:52:38","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":11830,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eVirulence gene profile of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/03c5aa12dcc5a846c0974c74.png"},{"id":85140344,"identity":"9c01c059-bb56-4720-9b30-2e8691d8c979","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":9253,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eBiofilm formation by \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/f3f7657d648addebbf83af58.png"},{"id":85140646,"identity":"a071e8ed-bd68-4634-acb6-e5fb5a156cc9","added_by":"auto","created_at":"2025-06-22 10:00:38","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":84640,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCorrelation analysis of antimicrobial resistance phenotypes and resistance gene profiles in ESBL-producing versus non-ESBL-producing strains\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote:\u003c/strong\u003eCircle size and color intensity represent the Pearson correlation coefficient (r) magnitude. Correlation strength is categorized as: |r| ≥ 0.8 (strong), 0.5 ≤ |r| \u0026lt; 0.8 (moderate), 0.3 ≤ |r| \u0026lt; 0.5 (weak), and |r| \u0026lt; 0.3 (negligible). Positive/negative values indicate direct/inverse relationships respectively.\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/bfa8c84fc37bdb864a65a146.png"},{"id":85140337,"identity":"9afb11de-3318-420f-80ac-bc1656e6988c","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":80709,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCorrelation analysis between virulence gene profiles and biofilm formation capacity in \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e isolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNote: Circle size and color gradient represent the magnitude of Pearson's correlation coefficient (r). Correlation strength is categorized as: strong (|r| ≥ 0.8), moderate (0.5 ≤ |r| \u0026lt; 0.8), weak (0.3 ≤ |r| \u0026lt; 0.5), or negligible (|r| \u0026lt; 0.3). Positive values indicate direct correlations while negative values represent inverse relationships.\u003c/p\u003e","description":"","filename":"9.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/25cbcd3cd3c1dc926db1a75d.png"},{"id":85140353,"identity":"9f8dc67e-fe1a-4dc7-8b67-55ee467710a1","added_by":"auto","created_at":"2025-06-22 09:52:38","extension":"png","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":556767,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePathological examination of mice intraperitoneal inoculated \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e strain ( Pm-07)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNote: \u003c/strong\u003eA: Compared with the control group, the mice had subcutaneous capillary hemorrhage and pathological changes in internal organs; B: Mice have enlarged livers with bleeding spots; C: Severe hemorrhage in the lungs of mice, dark red; D: Mouse kidney enlargement; E: The spleen of the mouse is enlarged, severely hemorrhaged, and dark red; F: Large thinning of the intestinal wall of the duodenum of mice( upper figure: control group; lower figure: challenged group).\u003c/p\u003e","description":"","filename":"10.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/ab1c501360af24fd6a82f988.png"},{"id":85140359,"identity":"0913f7a8-926c-4abb-bcfb-379ad36a2d65","added_by":"auto","created_at":"2025-06-22 09:52:38","extension":"png","order_by":11,"title":"Figure 11","display":"","copyAsset":false,"role":"figure","size":5713,"visible":true,"origin":"","legend":"\u003cp\u003eLegend not included with this version.\u003c/p\u003e","description":"","filename":"11.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/660146d962883015e60de4fc.png"},{"id":85140356,"identity":"765ff1c2-7a5b-4c79-a95b-3f1620363b25","added_by":"auto","created_at":"2025-06-22 09:52:38","extension":"png","order_by":12,"title":"Figure 12","display":"","copyAsset":false,"role":"figure","size":259137,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHistopathological observation of mice\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNote: A, B, C, D, E: lung, liver, spleen, intestinal and small intestine of mice in the control group; a, b, c, d: lung, liver, spleen, intestinal and small intestine of 2.6×10\u003csup\u003e7\u003c/sup\u003e CFU/mL mice in the experimental group.\u003c/p\u003e","description":"","filename":"12.png","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/da4a0a249d027638eff19d6a.png"},{"id":101295642,"identity":"613d78ec-697d-4c90-9364-cb6bdf08df5f","added_by":"auto","created_at":"2026-01-28 08:58:49","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3690480,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/a131e074-c711-44aa-aa44-8843d8066a37.pdf"},{"id":85140335,"identity":"fc8f5010-491c-406c-b24d-9dc66d990ff2","added_by":"auto","created_at":"2025-06-22 09:52:37","extension":"xls","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":23552,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTableS1.xls","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/321b65bead1ebebcbdcd22cf.xls"},{"id":85140647,"identity":"8eb7a374-d16c-405b-a99c-28c4403aa4e4","added_by":"auto","created_at":"2025-06-22 10:00:38","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":356217,"visible":true,"origin":"","legend":"","description":"","filename":"Supplementaryfilefigure3.docx","url":"https://assets-eu.researchsquare.com/files/rs-6592294/v1/b0366eaf66693a93edfd92d3.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Zoonotic threat of Proteus mirabilis in Xinjiang swine: antimicrobial resistance and virulence risks","fulltext":[{"header":"Introduction","content":"\u003cp\u003e \u003cem\u003eProteus mirabilis\u003c/em\u003e (\u003cem\u003eP. mirabilis\u003c/em\u003e) is a Gram-negative bacterium with cell migration phenomenon, widely present in soil, water, and sewage [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. It infects the digestive, respiratory, and urinary systems in humans and animals, with high infection rates in both populations [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Notably, it is a leading cause of urinary tract infections second only to \u003cem\u003eEscherichia coli\u003c/em\u003e [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eA major global concern is its resistance to β-lactam antibiotics due to extended-spectrum β-lactamases (ESBLs), which confer resistance to penicillin and cephalosporins. ESBLs mediated by plasmids can spread among different \u003cem\u003eEnterobacteriaceae\u003c/em\u003e [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], and multidrug-resistant (MDR) \u003cem\u003eP. mirabilis\u003c/em\u003e strains producing ESBLs and \u003cem\u003eAmpC\u003c/em\u003e enzymes are increasing worldwide [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. These strains can lead to clonal spread, causing outbreaks in hospitals and across regions [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Critically, infections caused by ESBL-producing \u003cem\u003eP. mirabilis\u003c/em\u003e are associated with significantly higher rates of treatment failure and patient mortality [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Since animal-derived ESBL-producing \u003cem\u003eP. mirabili\u003c/em\u003e threatens public health, this study isolated \u003cem\u003eP. mirabilis\u003c/em\u003e strains from healthy pigs and pork to assess their antibiotics resistance, ESBL-producing, and genetic determinants (resistance/virulence genes). The findings provide insights into MDR mechanisms and zoonotic risks associated with porcine \u003cem\u003eP. mirabilis\u003c/em\u003e.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eSample Collection\u003c/h2\u003e \u003cp\u003eSamples were collected from randomly selected healthy pigs in Xinjiang swine farms. Anal and nasal swabs were obtained by gentle rotation (2\u0026ndash;3 times) using sterile swabs. Diseased pig lung tissues were also collected. All samples were labeled, stored at -4\u0026deg;C, and transported to the lab. Retail pork samples were collected from Shihezi morning markets. A total of 216 samples were obtained (108 farm samples: swabs and tissues; 108 pork samples). See Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e for details.\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 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSample source\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBreeding Type\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSample Size\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSample Type\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSwine farm A\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e23\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHealthy swine nasal swab sample\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHealthy swine anal swab sample\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSwine farm B\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHealthy swine nasal swab sample\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSwine farm C\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHealthy swine anal swab sample\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSwine farm D\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e24\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eHealthy swine anal swab sample\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSwine farm E\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e31\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eInfected swine lung tissue\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eA fresh meat market in Shihezi City\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e48\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFresh meat in the market\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\n\u003ch3\u003eExperimental Animals\u003c/h3\u003e\n\u003cp\u003eThirty 6-week-old Kunming mice were obtained from Xinjiang Medical University's Laboratory Animal Center for pathogenicity testing. All animal experiments described in this study were approved by the Biology Ethics Committee of Shihezi University (Ref. No. A2025-650) and strictly complied with the guidelines for the care and use of laboratory animals. The mice were euthanized by cervical dislocation performed by trained personnel. Prior to euthanasia, the mice were rendered unconscious via intraperitoneal injection of 1% sodium pentobarbital at 50 mg/kg dosage to ensure anesthesia before cervical dislocation.\u003c/p\u003e\n\u003ch3\u003eBacterial Isolation and Gram staining examination\u003c/h3\u003e\n\u003cp\u003eSamples were inoculated into LB broth (5 mL, 37\u0026deg;C, 180 rpm, overnight) and plated on SS agar. Characteristic colonies (transparent periphery with black centers) were purified through 3\u0026ndash;5 subcultures and confirmed by Gram staining.\u003c/p\u003e\n\u003ch3\u003eBiochemical Identification\u003c/h3\u003e\n\u003cp\u003eIsolates were identified using a biochemical test kit (Hangzhou Microbial Reagent Co., Hangzhou, China) following the manufacturer\u0026rsquo;s instructions. Hemolytic activity was assessed on 5% sheep blood agar (37\u0026deg;C, 24h).\u003c/p\u003e\n\u003ch3\u003eMolecular Confirmation\u003c/h3\u003e\n\u003cp\u003ePurified bacterial isolates were cultured in LB broth (5 mL, 180 rpm, 12\u0026ndash;24 h), followed by genomic DNA extraction via boiling. The DNA was used for 16S rDNA amplification (primers in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e) under the following conditions: initial denaturation at 95\u0026deg;C for 5 min; 35 cycles of 95\u0026deg;C for 30 s, 58\u0026deg;C for 30 s, and 72\u0026deg;C for 90 s; with a final extension at 72\u0026deg;C for 10 min. In parallel, PCR was performed to amplify the \u003cem\u003eP. mirabilis\u003c/em\u003e-specific pm0395 gene for confirming the presence of \u003cem\u003eP. mirabilis\u003c/em\u003e, using primers listed in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. PCR products (5\u0026ndash;6 \u0026micro;L) were electrophoresed on 1.0% agarose gels and visualized, then sent for sequencing (General Bio Ltd., Anhui). Sequences were analyzed by BLAST (NCBI database).\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 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePrimer information for specific genes\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=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePrimer name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePrimer sequence (5 '-3')\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFragment size (bp)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAnnealing temperature (℃)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e16S rDNA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eF:AGAGTTTGATCCTGGCTCAG\u003c/p\u003e \u003cp\u003eR:TACGGCTACCTTGTTACGACTT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e1476\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003epm0395\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eF:ATCCAGTCACCACTAATCT\u003c/p\u003e \u003cp\u003eR:TCTGACATCAACAGTAATTG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e539\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \n\u003cp\u003e\u003cb\u003eAntimicrobial susceptibility testing of\u003c/b\u003e \u003cb\u003eP. mirabilis\u003c/b\u003e \u003cb\u003eisolates\u003c/b\u003e\u003c/p\u003e\n\u003cp\u003eThe antibiotic susceptibility profiles of all isolated strains were determined using the Kirby-Bauer disk diffusion method on Mueller-Hinton agar (MHA) in accordance with CLSI guidelines, including phenotypic screening and confirmation tests for ESBL production [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. The tested panel comprised 25 antibiotics from major classes: β-lactams (Cefotaxime/CTX 30 \u0026micro;g, Ceftazidime/CAZ 30 \u0026micro;g, Cephalosporin/CA 30 \u0026micro;g, Cefepime/CP 30 \u0026micro;g, Penicillin G/PG 10 \u0026micro;g, Amoxicillin/AM 10 \u0026micro;g, Ampicillin/AMP 10 \u0026micro;g), fluoroquinolones (Enrofloxacin/ENR 5 \u0026micro;g, Ciprofloxacin/CIP 5 \u0026micro;g, Norfloxacin/NF 5 \u0026micro;g), folate pathway inhibitors (Compound sulfamethoxazole/SXT 1.25/23.75 \u0026micro;g), tetracyclines (Tetracycline/TC 30 \u0026micro;g, Doxycycline/DOX 10 \u0026micro;g), aminoglycosides (Gentamicin/GN 10 \u0026micro;g, Kanamycin/KAN 30 \u0026micro;g, Amikacin/AMK 30 \u0026micro;g, Tobramycin/TOB 10 \u0026micro;g), phenicols (Fluphenicol/FFC 30 \u0026micro;g, Chloramphenicol/CL 30 \u0026micro;g), polymyxins (Polymyxin B/PB 30 \u0026micro;g), macrolides (Erythromycin/ERY 15 \u0026micro;g), glycopeptides (Vancomycin/VAN 30 \u0026micro;g), carbapenems (Imipenem/IMP 10 \u0026micro;g), and β-lactam/β-lactamase inhibitor combinations (Cefotaxime/clavulanic acid 10/10 \u0026micro;g, Ceftazidime/clavulanic acid 10/10 \u0026micro;g).\u003c/p\u003e \u003cp\u003e \u003cb\u003eDetection of antibiotic resistance genes in\u003c/b\u003e \u003cb\u003eP. mirabilis\u003c/b\u003e \u003cb\u003eisolates\u003c/b\u003e\u003c/p\u003e \u003cp\u003eAccording to the references [\u003cspan additionalcitationids=\"CR11\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e], the isolates were screened for major resistance genes including β-lactamases (\u003cem\u003eblaSHV\u003c/em\u003e, \u003cem\u003eblaCMY\u003c/em\u003e, \u003cem\u003eblaTEM\u003c/em\u003e, \u003cem\u003eblaCIT\u003c/em\u003e, \u003cem\u003eblaDHA\u003c/em\u003e, \u003cem\u003eblaMOX\u003c/em\u003e, \u003cem\u003eblaCTX-M-1/2/9\u003c/em\u003e, \u003cem\u003eblaOXA-1/48\u003c/em\u003e), sulfonamide (\u003cem\u003esul1-3\u003c/em\u003e), tetracycline (\u003cem\u003etetA/B\u003c/em\u003e), quinolone (\u003cem\u003eqnrA/B/D/S\u003c/em\u003e), chloramphenicol (\u003cem\u003efloR\u003c/em\u003e), aminoglycoside [\u003cem\u003eacc(6')-1b-cr\u003c/em\u003e], polymyxin (\u003cem\u003evanA/B\u003c/em\u003e), and macrolide resistance genes (\u003cem\u003emefA/D\u003c/em\u003e) using PCR with primers listed in Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e. The amplification protocol consisted of initial denaturation at 95\u0026deg;C (5 min), followed by 35 cycles of 95\u0026deg;C (30 s), gene-specific annealing (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e, 30s), and 72\u0026deg;C extension (90 s), with final extension at 72\u0026deg;C (10 min).\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 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePrimers information of antibiotic resistance genes and integron genes\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=\"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=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGene type\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGene Name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePrimer sequence (5 '-3')\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eFragment size (bp)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eAnnealing temperature (℃)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"10\" rowspan=\"11\"\u003e \u003cp\u003eβ-lactams\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaSHV\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:AGCCGCTTGAGCAAATTAAAC\u003c/p\u003e \u003cp\u003eR:ATCCCGCAGATAAATCACCAC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e635\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaTEM\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF: CATTTCCGTGTCGCCCTTATTC\u003c/p\u003e \u003cp\u003eR: CGTTCATCCATAGTTGCCTGAC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e800\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaCIT\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF: CGAAGAGGCAATGACCAGAC\u003c/p\u003e \u003cp\u003eR:ACGGACAGGGTTAGGATAGT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e538\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaDHA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF: TGATGGCACAGCAGGATATTC\u003c/p\u003e \u003cp\u003eR: GCTTTGACTCTTTCGGTATTCG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e997\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaMOX\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF: GCAACAACGACAATCCATCCT\u003c/p\u003e \u003cp\u003eR:GGGATAGGCGTAACTCTCCCAA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e895\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e57.5\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaCMY\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:TGGCCGTTGCCGTTATCTAC\u003c/p\u003e \u003cp\u003eR:CCCGTTTTATGCACCCATGA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e868\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaCTX-M-1\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF: TTAGGAARTGTGCCGCTGTA\u003c/p\u003e \u003cp\u003eR: CGATATCGTTGGTGGTACCAT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e688\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e54\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaCTX-M-2\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CGTTAACGGCACGATGAC\u003c/p\u003e \u003cp\u003eR:CGATATCGTTGGTGGTRCCAT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e404\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaCTX-M-9\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:TCAAGCCTGCCGATCTGGT\u003c/p\u003e \u003cp\u003eR:TGATTCTCGCCGCTGAAG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e561\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaOXA-1\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GACTTTATAAATTTAGTGTGTTTA\u003c/p\u003e \u003cp\u003eR:TCCCAAGGGTTCCAGCA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e816\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eblaOXA-48\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GCTTGATCGCCCTCGATT\u003c/p\u003e \u003cp\u003eR:GATTTGCTCCGTGGCCGAAA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e281\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eQuinolones\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eqnrA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CCAGGATTTGAGTGACAGC\u003c/p\u003e \u003cp\u003eR:GGCTCGTGTGTGCGGATG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e592\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e69\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eqnrB\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GATCGTGAAAGCCAGAAAGG\u003c/p\u003e \u003cp\u003eR:ATGAGCAACGATGCCTGGTA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e476\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e65\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eqnrD\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CGAGATCAATTTACGGGGAATA\u003c/p\u003e \u003cp\u003eR:TCTAAACCGTCGAGTTCGGCG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e572\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e65\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eqnrS\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GCAAGTTCATTGAACAGGGT\u003c/p\u003e \u003cp\u003eR:TGAAGGTCATCGATAGCAGG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e428\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e65\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eSulfonamide class\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003esul1\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CTTCGATGAGAGCCGGCGGC\u003c/p\u003e \u003cp\u003eR:GCAAGGCGGAAACCCGCGCC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e238\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e58\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003esul2\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CGGCATCGTCAACATAACC\u003c/p\u003e \u003cp\u003eR:GTGTGCGGATGAAGTCAG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e722\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003esul3\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:AGATGTGATTGATTTGGGAGC\u003c/p\u003e \u003cp\u003eR:TAGTTGTTTCTGGATTAGAGCCT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e443\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e50\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTetracyclines\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003etetA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GCTACATCCTGCTTGCCTTC\u003c/p\u003e \u003cp\u003eR:CATAGATCGCCGTGAAGAGG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e210\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e63\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003etetB\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:TGAAGGTCATCGATAGCAGG\u003c/p\u003e \u003cp\u003eR:ATTTGCCGACTACCTTGGTG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e391\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e55\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eChloramphenicol class\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003efloR\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GGCTTTCGTCATTGCGTCTC\u003c/p\u003e \u003cp\u003eR:ATCGGTAGGATGAAGGTGAGGA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e650\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSminoglycosides\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eAcc (6')\u0026mdash;1b\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:ATGACCTTGCGATGCTCTATGA\u003c/p\u003e \u003cp\u003eR:CGAATGCCTGGCGTGTTT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e486\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e54\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ePolymyxin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003evanA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GTAGGCTGCGATATTCAAAGC\u003c/p\u003e \u003cp\u003eR:CGATTCAATTGCGTAGTCCAA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e231\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003evanB\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GTAGGCTGCGATATTCAAAGC\u003c/p\u003e \u003cp\u003eR:GCCGACAATCAAATCATCCTC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e330\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eMacrolides\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003emefA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:AGTATCATTAATCACTAGTGC\u003c/p\u003e \u003cp\u003eR:TTCTTCTGGTACTAAAAGT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e348\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e53\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003emrsD\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GCCTTCCGGAGCTCCTACTT\u003c/p\u003e \u003cp\u003eR:GCGTCCAATGTATCTCTAT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e500\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e53\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eIntegron\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eintI1\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:ACGAGCGCAAGGTTTCGGT\u003c/p\u003e \u003cp\u003eR:GAAAGGTCTGGTCATACATG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e525\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eintI2\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CACGGATATGCGACAAAAAGGT\u003c/p\u003e \u003cp\u003eR:GTAGCAAACGACTGACGACAAAAAGGT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e789\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e56\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eintI3\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GCCTCCGGCAGCGACTTTCAG\u003c/p\u003e \u003cp\u003eR:GATGCTGCCCAGGGCGCTCG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e433\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e65\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 \u003cb\u003eDetection of integrations in\u003c/b\u003e \u003cb\u003eP. mirabilis\u003c/b\u003e \u003cb\u003eisolates\u003c/b\u003e\u003c/p\u003e \u003cp\u003eIntegrations play an important role in the transmission of bacterial resistance. PCR amplification was performed to detect class I, II, and III integrons using published primer sequences [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e] (see Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e for details).\u003c/p\u003e \u003cp\u003e \u003cb\u003eDetection of virulence genes in\u003c/b\u003e \u003cb\u003eP. mirabilis\u003c/b\u003e \u003cb\u003eisolates\u003c/b\u003e\u003c/p\u003e \u003cp\u003ePCR amplification using published primers [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e] (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e) was performed to detect 10 key virulence genes (\u003cem\u003eureC\u003c/em\u003e, \u003cem\u003emrpA\u003c/em\u003e, \u003cem\u003ersbA\u003c/em\u003e, \u003cem\u003eatfC\u003c/em\u003e, \u003cem\u003eucAA\u003c/em\u003e, \u003cem\u003epmfA\u003c/em\u003e, \u003cem\u003eatfA\u003c/em\u003e, \u003cem\u003ezapA\u003c/em\u003e, \u003cem\u003efⅠiL\u003c/em\u003e, and \u003cem\u003ehpmA\u003c/em\u003e) in the isolates, with reaction conditions following established protocols.\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 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePrimer sequence of virulence gene\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=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGene Type\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGene Name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePrimer sequence (5 '-3')\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eFragment size (bp)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eUrease\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eureC\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GTTATTCGTGATGGGATGGG\u003c/p\u003e \u003cp\u003eR:ATAAAGGTGGTTACGCCAG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e375\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMannose-resistant-like fimbriae\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003emrpA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:ACACCTGCCCATATGGAAGATACTGGTACA\u003c/p\u003e \u003cp\u003eR:AAGTGATGAAGCTTAGTGATGGTGATGGTG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e550\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMigration-ability-regulating gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003ersbA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:TCGATTTCAGTGTTTGGCCAT\u003c/p\u003e \u003cp\u003eR:TCGATTTCAGTGTTTGGCCAT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e1647\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eThermophilic fimbriae\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eatfA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CATAATTTCTAGACCTGCCCTAGCA\u003c/p\u003e \u003cp\u003eR:CTGCTTGGATCCGTAATTTTTAACG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e332\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eatfC\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:AGAAAGGGATCCTACAAATTAA\u003c/p\u003e \u003cp\u003eR:TATAGCATGCATTTAAATTGCC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e472\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eUroepithelial cell adhesin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eucaA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GTAAAGTTGTTGCGCAAAC\u003c/p\u003e \u003cp\u003eR:TTGAGCCACTGTGGATACA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e587\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFimbrial gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003epmfA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GGATCATCTATAATGAAACTG\u003c/p\u003e \u003cp\u003eR:CTGATAATCAACTTGGAAGTT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e512\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMetalloproteinase\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003ezapA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:ACCGCAGGAAAACATATAGCCC\u003c/p\u003e \u003cp\u003eR:GCGACTATCTTCCGCATAATCA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e543\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFlagellar gene\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003efⅠiL\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:CTCTGCTCGTGGTGGTGTCG\u003c/p\u003e \u003cp\u003eR:GCGTCGTCACCTGATGTGTC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e770\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eHemolysin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003ehpmA\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eF:GTTGAGGGGCGTTATCAAGAGTC\u003c/p\u003e \u003cp\u003eR:GATAACTGTTTTGCCCTTTTGTGC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e709\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 \u003cb\u003eBiofilm formation ability assay of\u003c/b\u003e \u003cb\u003eP. mirabilis\u003c/b\u003e \u003cb\u003eisolates\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe biofilm-forming capacity of isolates was determined using a 96-well microtiter plate assay [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Briefly, 180 \u0026micro;L LB medium and 20 \u0026micro;L bacterial culture were added per well, incubated at 37\u0026deg;C for 24 h, washed with PBS, and fixed with methanol (200 \u0026micro;L, 15 min). After crystal violet staining (1%, 5 min) and acetic acid dissolution (33%, 30 min), absorbance was measured at 590 nm. Biofilm formation was categorized as: non (OD\u0026thinsp;\u0026le;\u0026thinsp;negative control), weak (OD\u0026thinsp;\u0026le;\u0026thinsp;2\u0026times; control), moderate (OD\u0026thinsp;\u0026le;\u0026thinsp;4\u0026times; control), or strong (OD\u0026thinsp;\u0026gt;\u0026thinsp;4\u0026times; control), with triplicate wells per strain.\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eMouse pathogenicity assay\u003c/h2\u003e \u003cp\u003eThe multidrug-resistant, ESBL-producing \u003cem\u003eP. mirabilis\u003c/em\u003e strain Pm-07 (carrying 6 virulence and 8 resistance genes) was selected for challenge. Thirty 6-week-old Kunming mice were randomized into six groups (n\u0026thinsp;=\u0026thinsp;5/group). After 24-hour fasting, mice were intraperitoneally injected with 0.5 mL of 10-fold serially diluted bacterial suspensions (2.6\u0026times;10⁴\u0026ndash;2.6\u0026times;10⁹ CFU), while controls received sterile saline. Clinical signs and mortality were monitored every 4 hours. LD₅₀ was calculated using the modified Karber method. Deceased mice underwent necropsy for lesion evaluation, bacterial re-isolation (confirmed by PCR), and histopathology (4% formaldehyde-fixed, paraffin-embedded, H\u0026amp;E-stained tissues).\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eIsolation and identification of\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA total of 42 \u003cem\u003eP. mirabilis\u003c/em\u003e strains (19.44% isolation rate) were obtained from 216 samples (168 swine swabs and 48 pork/lung tissues) collected from five swine farms, with varying isolation rates across farms (11.90-35.48%) and lower detection in pork (4.16%) (Fig. 1). The isolates exhibited characteristic migration on BHI agar (\u0026gt;\u0026thinsp;18h incubation) and typical black-centered colonies on SS agar, with Gram-negative rod morphology. Biochemical profiling confirmed xylose fermentation, urea/ornithine/lysine decomposition, H₂S production, and \u0026beta;-hemolysis on 5% sheep blood agar, while testing negative for cottonseed sugar fermentation and other markers (Fig. 2).\u003c/p\u003e\n\u003cp\u003eMolecular identification via 16S rDNA sequencing revealed 99.65\u0026ndash;100% homology with known \u003cem\u003eP. mirabilis\u003c/em\u003e strains among the 42 isolates. The associated sequence datasets generated in this study have been deposited in the GenBank repository under accession numbers PV686792 to PV686828 and PV700941 to PV700945. Species-specific PCR consistently amplified the expected 539 bp fragment, conclusively identifying all isolates as \u003cem\u003eP. mirabilis\u003c/em\u003e (Fig. 3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAntimicrobial resistance profile of\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll 42 \u003cem\u003eP. mirablis\u003c/em\u003e isolates exhibited multidrug resistance, demonstrating\u0026thinsp;\u0026gt;\u0026thinsp;90% resistance to sulfonamides, tetracyclines, chloramphenicol, macrolides, polypeptides, and glycopeptides. Resistance rates varied by class: \u0026beta;-lactams (ampicillin 69.05%, penicillin 88.10%, amoxicillin 92.86%), quinolones (59.52\u0026ndash;85.71%), and aminoglycosides (gentamicin 50.00%, tobramycin 78.57%, kanamycin 80.95%), while showing relative sensitivity to amikacin (21.43%) and cefepime (9.52%) (Fig. 4A, B). Among these, 54.76% (23/42) were ESBL-producers, displaying broader resistance (11 antibiotics across 8 classes) compared to non-ESBL strains (6 antibiotics in 5 classes), confirming ESBL production significantly enhances resistance (Fig. 4C, D). All isolates were confirmed as multidrug-resistant, highlighting widespread resistance in porcine \u003cem\u003eP. mirabilis\u003c/em\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAntibiotic resistance gene profile of\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePCR screening of 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates identified 19 resistance genes across 7 classes, with \u0026beta;-lactamase genes being most prevalent (83.33% ESBL-carriers). Key findings include: universal presence of \u003cem\u003esul1\u003c/em\u003e (100%, sulfonamide resistance) and high prevalence of \u003cem\u003efloR\u003c/em\u003e (85.71%, chloramphenicol), \u003cem\u003eaac (6\u0026apos;)-Ib-cr\u003c/em\u003e (73.81%, aminoglycoside), and \u003cem\u003eCTX-M-type\u003c/em\u003e ESBLs (85.71%, including \u003cem\u003eCTX-M-2\u003c/em\u003e (45.24%), \u003cem\u003eCTX-M-9\u003c/em\u003e (35.71%), and \u003cem\u003eCTX-M-1\u003c/em\u003e (33.33%). Notably, 54.76% isolates carried\u0026thinsp;\u0026ge;\u0026thinsp;3 ESBL genotypes, while \u003cem\u003eqnrD\u003c/em\u003e (33.33%, quinolone) and \u003cem\u003etetA\u003c/em\u003e (38.10%, tetracycline) showed significant detection rates. No polymyxin resistance genes or \u003cem\u003eTEM\u003c/em\u003e genotypes were detected, highlighting specific resistance patterns in these porcine isolates (Fig. 5).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCorrelation between phenotypic and genotypic resistance in\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe consistency between the resistance phenotype and resistance genotype of the \u003cem\u003eP. mirabilis\u003c/em\u003e isolates is shown in Table 5. The 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates demonstrated varying concordance rates between resistance genes and phenotypes: high for sulfonamides (100.00%, 42/42), chloramphenicols (85.71%, 36/42), \u0026beta;-lactams (83.33%, 35/42), aminoglycosides (73.81%, 31/42), quinolones (54.76%, 23/42), and tetracyclines (40.48%, 17/42), but markedly lower for macrolides (26.19%,11/42) and polymyxins (0.00%, 0/42).\u0026nbsp;\u003c/p\u003e\n\u003ctable border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv\u003eTable 5\u003c/div\u003e\n \u003cdiv\u003e\n \u003cp\u003eConcordance between genotypic and phenotypic resistance profiles in \u003cem\u003eP. mirabilis\u003c/em\u003e isolates\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCategory of antibiotics\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eNumber of drug-resistant bacteria\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eNumber of bacterial strains containing resistance genes\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eDrug resistance compliance rate (%)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-lactams\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e83.33%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eQuinolones\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e54.76%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSulfonamide class\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e100.00%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTetracyclines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e40.48%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eChloramphenicol class\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e85.71%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAminoglycosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e73.81%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ePolymyxin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.00%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMacrolides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e26.19%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIntegron and virulence gene distribution in\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eIsolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAmong 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates, class I integrons were predominant (95.2%), while class II (2.4%) and III (9.5%) were rare. Virulence gene profiling revealed near-universal presence of \u003cem\u003efliL\u003c/em\u003e (97.6%), \u003cem\u003eureC\u003c/em\u003e (92.9%), \u003cem\u003eatfA\u003c/em\u003e (92.9%), and \u003cem\u003ezapA\u003c/em\u003e (90.5%), with lower frequencies for \u003cem\u003ehpmA\u003c/em\u003e (88.1%), \u003cem\u003epmfA\u003c/em\u003e (76.2%), \u003cem\u003eucaA\u003c/em\u003e (47.6%), \u003cem\u003emrpA\u003c/em\u003e (28.6%), and \u003cem\u003ersbA\u003c/em\u003e (23.8%). Notably, \u003cem\u003eatfC\u003c/em\u003e was absent in all isolates, demonstrating a conserved virulence gene profile with specific exceptions.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eVirulence gene profile of\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates carried varying combinations of nine virulence genes, with near-ubiquitous prevalence of four core virulence factors: flagellar gene \u003cem\u003efliL\u003c/em\u003e (97.6%), urease gene \u003cem\u003eureC\u003c/em\u003e (92.9%), thermoregulated fimbriae gene \u003cem\u003eatfA\u003c/em\u003e (92.9%), and metalloproteinase gene \u003cem\u003ezapA\u003c/em\u003e (90.5%). Secondary virulence determinants included hemolysin \u003cem\u003ehpmA\u003c/em\u003e (88.1%) and fimbrial genes \u003cem\u003epmfA\u003c/em\u003e (76.2%) and \u003cem\u003eucaA\u003c/em\u003e (47.6%), while mannose-resistant fimbriae \u003cem\u003emrpA\u003c/em\u003e (28.6%) and motility regulator \u003cem\u003ersbA\u003c/em\u003e (23.8%) were less common. Notably, \u003cem\u003eatfC\u003c/em\u003e was absent in all isolates, revealing a conserved virulence gene pattern with distinct prevalence tiers among clinical strains(Fig. 6).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eBiofilm formation capacity of\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e \u003cstrong\u003eisolates\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe biofilm-forming capacity of 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates was quantitatively assessed via microtiter plate assay (OD₅₉₀), revealing three distinct tiers: 80.95% (34/42) exhibited strong biofilm production (OD\u0026thinsp;\u0026gt;\u0026thinsp;4\u0026times;ODc),14.29% (6/42) showed moderate formation (2\u0026times;ODc\u0026thinsp;\u0026lt;\u0026thinsp;OD\u0026thinsp;\u0026le;\u0026thinsp;4\u0026times;ODc), while only 4.76% (2/42) pork-derived isolates demonstrated weak capability (1\u0026times;ODc\u0026thinsp;\u0026lt;\u0026thinsp;OD\u0026thinsp;\u0026le;\u0026thinsp;2\u0026times;ODc). (Fig. 7).\u003c/p\u003e\n\u003ch3\u003eStatistical correlation between phenotypic resistance patterns and genotypic resistance determinants for \u0026beta;-lactam antibiotics\u003c/h3\u003e\n\u003cp\u003eCorrelation analysis was performed to assess the relationship between \u0026beta;-lactam antibiotic resistance phenotypes and resistance genes in 42 \u003cem\u003eP. mirabilis\u003c/em\u003e strains, including both ESBL-producing and non-ESBL-producing isolates. As demonstrated in Fig. 8, the overall correlation between resistance phenotypes and resistance genes to \u0026beta;-lactamase antibiotics was not statistically significant. Among ESBL-producing strains, phenotypic resistance showed a marginal positive correlation with the \u003cem\u003eblaSHV\u003c/em\u003e gene (Pearson\u0026apos;s r\u0026thinsp;=\u0026thinsp;0.36) and a slight negative correlation with the \u003cem\u003eblaCMY\u003c/em\u003e gene (r = -0.31). In contrast, non-ESBL-producing strains exhibited weak negative correlations with both \u003cem\u003eblaSHV\u003c/em\u003e (r = -0.36) and \u003cem\u003eblaCMY\u003c/em\u003e (r = -0.31). No significant correlations were observed between resistance phenotypes and other \u0026beta;-lactamase resistance genes in either group.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCorrelation analysis of virulence genes and biofilm formation in\u003c/strong\u003e \u003cstrong\u003eP. mirabilis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBiofilm formation and virulence gene profiles were analyzed in 42 clinical \u003cem\u003eP. mirabilis\u003c/em\u003e isolates to assess their potential association. As shown in Fig. 9, most virulence genes demonstrated weak correlations with biofilm-forming capacity. Notably, only the temperature-adapted fimbrial gene (\u003cem\u003eatfA\u003c/em\u003e) showed a modest positive correlation (r\u0026thinsp;=\u0026thinsp;0.40) with biofilm formation, suggesting a potential role in bacterial adhesion and biofilm development.\u003c/p\u003e\n\u003cdiv\u003e\n \u003ch2\u003eMouse pathogenicity test\u003c/h2\u003e\n \u003cp\u003eComprehensive characterization of 42 \u003cem\u003eP. mirabilis\u003c/em\u003e isolates identified strain Pm-07 for murine pathogenicity testing. Intraperitoneal inoculation (2.6\u0026times;10⁸ CFU/mL) induced clinical signs (lethargy, anorexia, diarrhea) within 6 hours, with mortality commencing at 24 hours. Necropsy revealed significant organ enlargement (lungs, liver, spleen, kidneys) with hemorrhaging and duodenal wall thinning (Fig. 10). PCR confirmed bacterial re-isolation from affected tissues (Fig. 11), with calculated LD₅₀ = 7.05\u0026times;10⁷ CFU/mL. Histopathology demonstrated alveolar hyperplasia, hepatic disorganization, splenic inflammatory infiltration, glomerular congestion, and intestinal villi damage with neutrophil recruitment (Fig. 12).\u003c/p\u003e\n\u003c/div\u003e\n"},{"header":"Discussion","content":"\u003cp\u003e \u003cem\u003eP. mirabilis\u003c/em\u003e, along with \u003cem\u003eKlebsiella pneumoniae\u003c/em\u003e and \u003cem\u003eEscherichia coli\u003c/em\u003e, ranks among the most clinically significant \u003cem\u003eEnterobacteriaceae\u003c/em\u003e responsible for both nosocomial and community-acquired infections globally [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Our study identified \u003cem\u003eP. mirabilis\u003c/em\u003e in Xinjiang farming environments with an overall prevalence of 19.4%, showing notable farm-to-farm variation. These findings complement existing epidemiological data demonstrating variable detection rates across China - from 28.9% in Xinjiang sheep farms [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e] to 7.07% in Shandong poultry operations [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. The observed geographical disparities in prevalence likely reflect regional differences in animal husbandry practices and environmental conditions, while the consistent detection across diverse livestock species underscores \u003cem\u003eP. mirabilis\u003c/em\u003e's adaptability and zoonotic potential.\u003c/p\u003e \u003cp\u003eThe escalating use of broad-spectrum antibiotics has precipitated a concerning rise in antimicrobial resistance among \u003cem\u003eProteus mirabilis\u003c/em\u003e isolates, particularly with the increasing prevalence of ESBL-producing strains that substantially complicate treatment regimens [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. Our study of porcine-derived \u003cem\u003eP. mirabilis\u003c/em\u003e isolates demonstrated alarming resistance profiles, exhibiting\u0026thinsp;\u0026gt;\u0026thinsp;90% resistance to 10 antimicrobial agents spanning 7 classes, with retained susceptibility only to amikacin (aminoglycoside) and cefepime (4th-generation cephalosporin). All isolates exhibited multidrug resistance (MDR) to \u0026ge;\u0026thinsp;5 antibiotic classes, mirroring Sun et al.'s [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e] findings of 100% resistance to 6 antibiotic classes in bovine-derived strains, which remained sensitive solely to ceftazidime, gentamicin and amikacin. This convergence of severe MDR patterns across livestock species and geographical regions signals an urgent need for comprehensive antibiotic stewardship programs in animal agriculture, enhanced surveillance of resistance gene transmission, and development of novel therapeutic strategies to address this critical public health challenge.\u003c/p\u003e \u003cp\u003eβ-lactamase genes represent a major mechanism of bacterial resistance. Studies reveal varying prevalence of these genes: Sun [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e] reported \u003cem\u003eblaCTX-M\u003c/em\u003e (45.1%), \u003cem\u003eblaTEM\u003c/em\u003e (13.7%), and \u003cem\u003eblaSHV\u003c/em\u003e (21.6%) in clinical isolates, while Cao [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e] found higher rates in fur animals (\u003cem\u003eblaTEM\u003c/em\u003e 83.0%, blaOXA-1 60.4%). Our study detected \u003cem\u003eblaCTX-M-2\u003c/em\u003e in 45.2% of isolates-higher than South African poultry (38%) [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] but showing distinct regional patterns, with \u003cem\u003eCTX-M\u003c/em\u003e variants demonstrating geographical predominance (\u003cem\u003eCTX-M-2\u003c/em\u003e in Latin America [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e] vs \u003cem\u003eCTX-M-1/9\u003c/em\u003e in other regions [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]).\u003c/p\u003e \u003cp\u003eNotably, ESBL-producing strains showed weak correlations with specific genes (\u003cem\u003eblaSHV\u003c/em\u003e: r\u0026thinsp;=\u0026thinsp;0.36; \u003cem\u003eblaCMY\u003c/em\u003e: r=-0.31), contrasting with Liu's findings of strong cefotaxime-\u003cem\u003eblaCTX-M\u003c/em\u003e correlations (r\u0026thinsp;=\u0026thinsp;0.95-1) in canine isolates. This variability likely reflects complex interactions among β-lactamase genes, where compensatory mechanisms may obscure individual gene effects on resistance phenotypes [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIntegrons play a critical role in the development of multidrug resistance (MDR) through their ability to acquire, integrate, and express multiple antibiotic resistance gene cassettes, with over 80 such cassettes identified in class I integrons alone. The high prevalence of integrons across different \u003cem\u003eP. mirabilis\u003c/em\u003e strains is particularly concerning, as demonstrated by Shi's finding of 100% class I integron carriage in porcine isolates [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e], Chen's report of 60.7% integron positivity in human clinical strains (including 20% class I, 14.7% class II, and 26% carrying both) [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e], and our observation of 95.2% class I integron prevalence with 9.5% co-carriage of multiple integron types. These findings collectively highlight integrons as key vectors for resistance gene dissemination and MDR development [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], emphasizing the urgent need for enhanced resistance surveillance and stricter antibiotic stewardship in both agricultural and clinical settings to curb the spread of these mobile genetic elements.\u003c/p\u003e \u003cp\u003eThe \u003cem\u003eP. mirabilis\u003c/em\u003e isolates carried multiple virulence genes (ureC, pmfA, zapA, fliL, atfA, hpmA, ucaA), with fliL (97.62%, 41/42) and ureC (92.86%, 39/42) being most prevalent. The fliL gene encodes an ~\u0026thinsp;18kDa transmembrane protein near the flagellar basal body [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e], crucial for bacterial motility [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. Similar to Qu et al.'s findings [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e], which identified 20 flagellar virulence factors (including \u003cem\u003eflgI\u003c/em\u003e, \u003cem\u003efliM\u003c/em\u003e, \u003cem\u003eflgC\u003c/em\u003e) in all 30 strains, and consistent with Chengdu studies showing high \u003cem\u003eureC\u003c/em\u003e prevalence in pets [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eBiofilm formation, a key virulence factor [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e], was observed in all isolates: 80.95% (34/42) strong, 14.29% (6/42) moderate, and 4.76% (2/42) weak producers. This aligns with Jia's report of universal biofilm formation in chicken-derived strains (15.38% strong) [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e] and Sun's findings in diarrheic animals (92.05% producers) [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Biofilm-positive strains showed higher pathogenicity, significantly correlating with \u003cem\u003eureC\u003c/em\u003e, \u003cem\u003ezapA\u003c/em\u003e, \u003cem\u003ersmA\u003c/em\u003e, \u003cem\u003ehmpA\u003c/em\u003e, \u003cem\u003emrpA\u003c/em\u003e, \u003cem\u003eatfA\u003c/em\u003e, and \u003cem\u003epmfA\u003c/em\u003e (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Specifically, biofilm formation weakly correlated with \u003cem\u003eatfA\u003c/em\u003e (r\u0026thinsp;=\u0026thinsp;0.40).\u003c/p\u003e \u003cp\u003eFurther research should investigate strain-specific biofilm-virulence interactions to optimize clinical management.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study reveals that swine-derived \u003cem\u003eP. mirabilis\u003c/em\u003e strains are multidrug-resistant, carrying various resistance and virulence genes, with 80.95% exhibiting strong biofilm formation. These findings highlight significant zoonotic risks through potential food chain transmission, calling for improved antimicrobial stewardship and surveillance in livestock production to mitigate public health threats.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003e\u003cem\u003eP. mirabilis\u003c/em\u003e, \u003cem\u003eProteus mirabilis\u003c/em\u003e; PEN, Penicillin; AMX, Amoxicillin; CFB, Cefazolin; CTX, Cefotaxime; K, Kanamycin; GM, Gentamicin; ENR, Enrofloxacin; CIP, Ciprofloxacin; DX, Doxycycline; TMP, Trimethoprim-sulfamethoxazole; FON, Florfenicol; PB, Polymyxin B; AMP, Ampicillin; IPM, Imipenem; TCY, Tetracycline; CRO, Ceftriaxone; CA, Vancomycin\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors wish to express their gratitude to the College of Animal Science and Technology, Xinjiang Tecon Animal Husbandry Technology Co., Ltd., for facilitating this study.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was funded by the Natural Science Support Program Project of the Xinjiang Production and Construction Corps (2024DA007) and the Shihezi University Technology Transfer and Promotion Program (CGZH202309). The APC was funded by the Natural Science Support Program Project of the Xinjiang Production and Construction Corps (2024DA007).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor information\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNana LI, Shenghui WAN and Yibo WANG contributed equally to this work.\u003c/p\u003e\n\u003cp\u003eAuthors and Affiliations\u003c/p\u003e\n\u003cp\u003e\u003csup\u003e1\u003c/sup\u003eCollege of Animal Science and Technology，Shihezi University, Shihezi 832003, China\u003c/p\u003e\n\u003cp\u003eYonggang QU, Gaoming HE, Yanfang Li, Yan LIANG, Nana LI, Shenghui WAN\u003c/p\u003e\n\u003cp\u003e\u003csup\u003e2\u003c/sup\u003eCollege of Food and Bioengineering, Henan University of Science and Technology\u003c/p\u003e\n\u003cp\u003eYuwan LI, Yibo WANG\u003c/p\u003e\n\u003cp\u003e\u003csup\u003e3\u003c/sup\u003eXinjiang Tecon Animal Husbandry Technology Co., Ltd., Changji 831399, China\u003c/p\u003e\n\u003cp\u003ePei ZHENG\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWriting-original draft preparation, Nana Li. Shenghui Wan; the acquisition, analysis, Yibo WANG; resources, Pei Zheng; review and editing, Yuwan LI;review and editing, Yanfang Li; resources, Gaoming HE;resources, Yan Liang; Supervision, Yonggang Qu; Project administration, Yonggang Qu; Funding acquisition, Yonggang Qu. All the authors have read and agreed to the published version of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe original contributions presented in the study are included in the article, further inquiries can be directed to the corresponding author after reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study received ethical approval from the Biology\u0026nbsp;Ethics Committee of Shihezi University (Approval No. A2025-650). Written informed consent was obtained from all animal owners prior to their participation in the research.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot Applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing financial interests.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAdeolu M, Alnajar S, Naushad S, Gupta S. Genome-based phylogeny and taxonomy of the 'Enterobacteriales': proposal for Enterobacterales ord. nov. divided into the families Enterobacteriaceae, Erwiniaceae fam. nov., Pectobacteriaceae fam. nov., Yersiniaceae fam. nov., Hafniaceae fam. nov., Morganellaceae fam. nov., and Budviciaceae fam. nov. 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Study on the resistance characteristics of avian Escherichia coli and Klebsiella pneumoniae producing extended spectrum β-lactase. Zhengzhou: Henan Agricultural University; 2024.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShi Baoqiu Z, Di G, Shuo Z, Xiaoliang J, Jingliang Y, Guangfu, et al. Isolation and identification of Proteus mirabilis from pigs and detection of class I integrons. Chin J Anim Infect Dis. 2020;28(2):7\u0026ndash;13.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen Keda L, Qiaoping QW, Yanye T. Goryeo. Analysis of Homozygous Distribution and Drug Resistance in Proteus mirabilis. Chin J Antibiot. 2020;45(11):1148\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOdumosu BT, Adeniyi BA, Chandra R. Analysis of integrons and associated gene cassettes in clinical isolates of multidrug resistant Pseudomonas aeruginosa from Southwest Nigeria. Ann Clin Microbiol Antimicrob. 2013;12:29.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee YY, Belas R. Loss of FliL alters Proteus mirabilis surface sensing and temperature-dependent swarming. J Bacteriol. 2015;197(1):159\u0026ndash;73.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAttmannspacher U, Scharf BE, Harshey RM. FliL is essential for swarming: motor rotation in absence of FliL fractures the flagellar rod in swarmer cells of Salmonella enterica. Mol Microbiol. 2008;68(2):328\u0026ndash;41.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eQu X, Zhou J, Huang H, Wang W, Xiao Y, Tang B, et al. Genomic Investigation of Proteus mirabilis Isolates Recovered From Pig Farms in Zhejiang Province, China. Front Microbiol. 2022;13:952982.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFusco A, Coretti L, Savio V, Buommino E, Lembo F, Donnarumma G. Biofilm Formation and Immunomodulatory Activity of Proteus mirabilis Clinically Isolated Strains. Int J Mol Sci. 2017;18(2):414.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJia Yuanzheng. Biological characteristics of chicken derived Proteus mirabilis and its LuxS/AI-2 quorum sensing system. Yangzhou University; 2021.\u003c/span\u003e\u003c/li\u003e\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":"Pigs, Proteus mirabilis, Antimicrobial resistance, Virulence genes, Pathogenicity","lastPublishedDoi":"10.21203/rs.3.rs-6592294/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6592294/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e \u003cem\u003eProteus mirabilis\u003c/em\u003e (\u003cem\u003eP. mirabilis\u003c/em\u003e) is an opportunistic pathogen causing zoonotic diseases like diarrhea, keratitis, and urinary tract infections, sometimes leading to animal miscarriages. This study examined \u003cem\u003eP. mirabilis\u003c/em\u003e prevalence in Xinjiang\u0026rsquo;s pig farms and pork, analyzing its drug resistance and pathogenicity to address food safety concerns. From 216 samples, 42 \u003cem\u003eP. mirabilis\u003c/em\u003e strains were isolated (19.44%, 42/216), mostly from pig farms (95.24%, 40/42) rather than pork (4.76%, 2/42). All strains were multidrug-resistant, showing over 90.00% resistance to sulfonamides, tetracyclines, chloramphenicol, macrolides, polypeptides, and glycopeptides. Among them, 54.76% (23/42) produced extended-spectrum β-lactamases (ESBLs), enhancing resistance. The strains carried 19 resistance genes (7 categories), with \u003cem\u003eCTX\u003c/em\u003e and \u003cem\u003eOXA\u003c/em\u003e type ESBL related resistance genes exceeding 30.00% detection rate. Additionally, 95.24% (40/42) of the isolated strains carried class I integron genes. Nine virulence genes (\u003cem\u003eureC\u003c/em\u003e, \u003cem\u003ezapA\u003c/em\u003e, \u003cem\u003eatfA\u003c/em\u003e, \u003cem\u003eucaA\u003c/em\u003e, \u003cem\u003epmfA\u003c/em\u003e, \u003cem\u003emrpA\u003c/em\u003e, r\u003cem\u003esbA\u003c/em\u003e, \u003cem\u003efliL\u003c/em\u003e, and \u003cem\u003ehpmA\u003c/em\u003e ) were identified, and most strains (80.95%, 34/42) had strong biofilm-forming ability. These findings indicate that multidrug-resistant \u003cem\u003eP. mirabilis\u003c/em\u003e in pigs, carrying diverse resistance and virulence genes, may spread via the food chain, posing health risks to consumers.\u003c/p\u003e","manuscriptTitle":"Zoonotic threat of Proteus mirabilis in Xinjiang swine: antimicrobial resistance and virulence risks","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-06-22 09:52:32","doi":"10.21203/rs.3.rs-6592294/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":"6c0a8e1d-f974-49d4-8e33-de24ce0dd223","owner":[],"postedDate":"June 22nd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-01-28T08:56:52+00:00","versionOfRecord":[],"versionCreatedAt":"2025-06-22 09:52:32","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6592294","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6592294","identity":"rs-6592294","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}