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Johne's disease (JD), chronic granulomatous disease caused by Mycobacterium avium subsp. paratuberculosis (MAP), is regarded as a potential public health issue and a significant threat to the dairy and agricultural food businesses. This study was conducted for the molecular investigation of MAP in cattle reared for small-scale dairy production in Sharkia Governorate, Egypt. Seventy-five fecal samples were collected from diarrheic and healthy dairy cows that came into contact with diarrheic ones for molecular screening. Clinically, 32 of 75 examined cattle showed variable degrees of body weight loss and diarrhea. The PCR targeting the insertion sequence IS900 gene revealed that 22.6% of examined cattle were positive for MAP. A rate was significantly higher in diseased older cattle than in young cattle with no symptoms of the disease. Furthermore, DNA sequencing and phylogenetic analysis revealed that our strains (ON816021 and ON816022) were 100% identical showed complete identity with MAP-C (M. paratuberculosis of cattle origin) as Japanese (CP066812) and German strain (CP022105). Overall, the findings highlight the potential of the IS900 gene-based molecular technique for MAP detection in small dairy cattle farms. [ABSTRACT FROM AUTHOR]

Simple Summary: The present study aimed to isolate Corynebacterium pseudotuberculosis from caseous lymphadenitis (CLA) in smallholder sheep and goats. Thereafter, antimicrobial resistance determinants, virulence characteristics, and phylogenetic analysis of the isolates were investigated. Our results revealed that 24.54% (54/220) of examined animals showed CLA-compatible lesions from which C. pseudotuberculosis was isolated. Antimicrobial susceptibility testing revealed that all isolates were resistant to bacitracin and florfenicol. Additionally, 92.6% of the isolates displayed resistance to penicillin and erythromycin, while none of the isolates were resistant to norfloxacin. Interestingly, 16.7% of C. pseudotuberculosis isolates recovered from sheep showed vancomycin resistance. Molecular characterization revealed that PLD, PIP, and FagA virulence genes were detected in all examined C. pseudotuberculosis isolates. The bla (β-lactam) resistance gene was present in all isolates. Additionally, 83% of sheep isolates carried the aminoglycoside (aph(3″)-lb), chloramphenicol (cat1), and bacitracin (bcrA) resistance genes. Notably, the glycopeptide (vanA) resistance gene was detected in 8% of the sheep isolates as a first report. Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) genotyping of 10 multi-drug resistant C. pseudotuberculosis isolates showed a high similarity index of 83.6% between isolates from sheep and goats. Nucleotide sequence analysis of partial 16S rRNA sequences of these isolates showed 98.83% similarities with biovar Ovis of globally available reference sequences on the Genbank database. In conclusion, this study emphasized the high isolation rate of C. pseudotuberculosis and its molecular characterization in smallholder sheep and goats in four districts in Egypt. Caseous lymphadenitis (CLA) is a bacterial infection caused by Corynebacterium pseudotuberculosis (C. pseudotuberculosis) that affects sheep and goats, leading to abscess formation in their lymph nodes. The present study aimed to isolate and identify C. pseudotuberculosis from CLA in smallholder sheep and goats, and determine the resistance patterns, virulence, and resistance genes of the isolates. Additionally, genotypic and phylogenetic analysis of the isolates was conducted using ERIC-PCR and DNA sequencing techniques. A cross-sectional study examined 220 animals (130 sheep and 90 goats) from 39 smallholder flocks for clinical signs of CLA. Fifty-four (24.54%) animals showed CLA-compatible lesions, confirmed by C. pseudotuberculosis isolation and PCR identification. Sheep had a lower infection rate of CLA (18.46%) compared with goats (33.3%). Antimicrobial susceptibility testing of 54 C. pseudotuberculosis isolates to 24 antimicrobial drugs revealed that they were 100% resistant to bacitracin and florfenicol, while none of the isolates were resistant to norfloxacin. A high resistance rate was observed for penicillin and erythromycin (92.6% each). Interestingly, 16.7% of C. pseudotuberculosis isolates recovered from sheep showed vancomycin resistance. Molecular characterization of C. pseudotuberculosis isolates revealed that PLD, PIP, and FagA virulence genes were present in all examined isolates. However, the FagB, FagC, and FagD genes were detected in 24 (100%), 20 (83%), and 18 (75%) of the sheep isolates, and 26 (87%), 26 (87%), and 18 (60%) of the goat isolates, respectively. The β-lactam resistance gene was present in all isolates. Furthermore, 83% of the sheep isolates carried the aminoglycoside (aph(3″)-lb), chloramphenicol (cat1), and bacitracin (bcrA) resistance genes. Among the isolates recovered from goats, 73% were found to contain macrolides (ermX), sulfonamide (sul1), and bacitracin (bcrA) resistance genes. It is worrisome that the glycopeptide (vanA) resistance gene was detected in 8% of the sheep isolates as a first report. ERIC-PCR genotyping of 10 multi-drug-resistant C. pseudotuberculosis isolates showed a high similarity index of 83.6% between isolates from sheep and goats. Nucleotide sequence analysis of partial 16S rRNA sequences of C. pseudotuberculosis revealed 98.83% similarity with biovar Ovis of globally available reference sequences on the Genbank database. Overall, our findings might indicate that C. pseudotuberculosis infection in smallholders in Egypt might be underestimated despite the significant financial impact on animal husbandry and potential health hazards it poses. Moreover, this study highlights the importance of implementing a sustainable control strategy and increasing knowledge and awareness among smallholder breeders to mitigate the economic impact of CLA. [ABSTRACT FROM AUTHOR]

Diarrhea caused by Escherichia coli in calves is an important problem in terms of survivability, productivity and treatment costs. In this study, 88 of 150 diarrheic animals tested positive for E. coli. Of these, 54 samples had mixed infection with other bacterial and/or parasitic agents. There are several diarrheagenic E. coli pathotypes including enteropathogenic E. coli (EPEC), Shiga-toxin producing E. coli (STEC), enterotoxigenic E. coli (ETEC) and necrotoxigenic E. coli (NTEC). Molecular detection of virulence factors Stx2, Cdt3, Eae, CNF2, F5, Hly, Stx1, and ST revealed their presence at 39.7, 27.2, 19.3, 15.9, 13.6, 9.0, 3.4, and 3.4 percent, respectively. As many as 13.6% of the isolates lacked virulence genes and none of the isolate had LT or CNF1 toxin gene. The odds of isolating ETEC from male calves was 3.6 times (95% CI: 1.1, 12.4; P value = 0.042) that of female calves, whereas the odds of isolating NTEC from male calves was 72.9% lower (95% CI: 91.3% lower, 15.7% lower; P value = 0.024) than that in females. The odds of isolating STEC in winter was 3.3 times (95% CI: 1.1, 10.3; P value = 0.037) that of spring. Antibiograms showed 48 (54.5%) of the isolates to be multi-drug resistant. The percent resistance to tetracycline, streptomycin, ampicillin, and trimethoprim-sulfamethoxazole was 79.5, 67.0, 54.5, and 43.0, respectively. Ceftazidime (14.8%), amoxicillin-clavulanic acid (13.6%) and aztreonam (11.3%) showed the lowest resistance, and none of the isolates was resistant to imipenem. The results of this study can help improve our understanding of the epidemiological aspects of E. coli infection and to devise strategies for protection against it. The prevalence of E. coli pathotypes can help potential buyers of calves to avoid infected premises. The antibiograms in this study emphasizes the risks associated with the random use of antibiotics. [ABSTRACT FROM AUTHOR]