Multidrug-Resistant Enteropathogenic Escherichia coli Isolated from Diarrhoeic Calves, Milk, and Workers in Dairy Farms: A Potential Public Health Risk.

Enteropathogenic Escherichia coli (EPEC) is a leading cause of diarrhoeagenic diseases in humans and cattle worldwide. The emergence of multidrug-resistant (MDR) EPEC from cattle sources is a public health concern. A total of 240 samples (75 diarrhoeic calves, 150 milk samples, and 15 workers) were examined for prevalence of EPEC in three dairy farms in Egypt. Antimicrobial resistance (AMR) traits were determined by antibiogram and polymerase chain reaction (PCR) detection of ß-lactamase-encoding genes, plasmid-mediated quinolone resistance genes, and carbapenemase-encoding genes. The genetic relatedness of the isolates was assessed using repetitive extragenic palindromic sequence-based PCR (REP-PCR). EPEC isolates were detected in 22.7% (17/75) of diarrhoeic calves, 5.3% (8/150) of milk samples, and 20% (3/15) of worker samples. The detected serovars were O26 (5%), O111 (3.3%), O124 (1.6%), O126 (0.8%), and O55 (0.8%). AMR-EPEC (harbouring any AMR gene) was detected in 9.2% of samples. Among isolates, blaTEM was the most detected gene (39.3%), followed by blaSHV (32.1%) and blaCTX-M-1 (25%). The qnrA, qnrB, and qnrS genes were detected in 21.4%, 10.7%, and 7.1% of isolates, respectively. The blaVIM gene was detected in 14.3% of isolates. All EPEC (100%) isolates were MDR. High resistance rates were reported for ampicillin (100%), tetracycline (89.3%), cefazolin (71%), and ciprofloxacin (64.3%). Three O26 isolates and two O111 isolates showed the highest multiple-antibiotic resistance (MAR) indices (0.85-0.92); these isolates harboured blaSHV-12 and blaCTX-M-15 genes, respectively. REP-PCR genotyping showed high genetic diversity of EPEC, although isolates belonging to the same serotype or farm were clustered together. Two worker isolates (O111 and O26) showed high genetic similarity (80-95%) with diarrhoeic calf isolates of matched serotypes/farms. This may highlight potential inter-species transmission within the farm. This study highlights the potential high risk of cattle (especially diarrhoeic calves) as disseminators of MDR-EPEC and/or their AMR genes in the study area. Prohibition of non-prescribed use of antibiotics in dairy farms in Egypt is strongly warranted.

Molecular Analysis of Antimicrobial Resistance among Enterobacteriaceae Isolated from Diarrhoeic Calves in Egypt.

The present study was designed to investigate the presence of genes that conferred resistance to antimicrobials among Enterobacteriaceae that were isolated from diarrhoeic calves. A total of 120 faecal samples were collected from diarrhoeic calves that were raised in Kafr El-Sheikh governorate, Egypt. The samples were screened for Enterobacteriaceae. A total of 149 isolates of bacteria were recovered and identified; Escherichia coli was found to be the most overwhelming species, followed by Citrobacter diversus, Shigella spp., Serratia spp., Providencia spp., Enterobacter spp., Klebsiella pneumoniae, Proteus spp., Klebsiella oxytoca, and Morganella morganii. All isolates were tested for susceptibility to 12 antimicrobials; resistant and intermediately resistant strains were screened by conventional polymerase chain reaction for the presence of antimicrobial resistance genes. Of the 149 isolates, 37 (24.8%) exhibited multidrug resistant phenotypes. The most prevalent multidrug resistant species were E. coli, C. diversus, Serratia spp., K. pneumoniae, Shigella spp., Providencia spp., and K. oxytoca. Class 1 integrons were detected in 28 (18.8%) isolates. All isolates were negative for class 2 integrons. The blaTEM gene was identified in 37 (24.8%) isolates, whereas no isolates carried the blaCTX-M gene. The florfenicol gene (floR) was detected in two bacterial isolates (1.3%). The findings of this study reveal that calves may act as potential reservoirs of multidrug resistant bacteria that can be easily transmitted to humans.

Leptospirosis as a neglected burden at human-cattle interface in Mid-Delta of Egypt.

INTRODUCTION: Leptospirosis is a neglected zoonosis in developing countries including Egypt where its burden is underestimated. METHODOLOGY: A cross sectional study was carried out to estimate the seroprevalence and associated risk factors of Leptospira interrogans serovar Hardjo infection among cows and leptospirosis among human patients in Mid-Delta of Egypt. RESULTS: Out of 112 examined cows using ELISA, 3.6% were seropositive to L. interrogans serovar Hardjo infection. Seroconversion occurred in 5 animals (1 herd) of all examined animals in convalescent phase testing (5/112, 4.5%). Affected herd suffered acute outbreak with 43.3% within herd prevalence; signs of infection included abortions, bloody urine and sudden death of 2 cows. Highest risk for L. interrogans serovar Hardjo infection in cows was in animals drank from untreated surface water (6.7 times, p = 0.06). The seroprevalence of leptospirosis was 6.2% in all tested humans, 28.6% in nonspecific fever cases and 22.2% in non-viral hepatitis cases. The risk of leptospirosis among patients with nonspecific fever or non-viral hepatitis cases was 4 times higher than those with viral hepatitis (p = 0.01). Additionally, there was a significant association between leptospirosis and patients with livestock contact (Odds 8, p = 0.01). CONCLUSIONS: This is the first report of L. interrogans serovar Hardjo outbreak in cows in Egypt. The study also highlighted the role of leptospirosis as neglected cause of nonspecific fever/non-viral hepatitis in humans in study region.

Antimicrobial and antibiofilm potentials of cinnamon oil and silver nanoparticles against Streptococcus agalactiae isolated from bovine mastitis: new avenues for countering resistance.

BACKGROUND: Streptococcus agalactiae (S. agalactiae) is a contagious pathogen of bovine mastitis. It has financial implications for the dairy cattle industry in certain areas of the world. Since antimicrobial resistance increases in dairy farms, natural antimicrobials from herbal origins and nanoparticles have been given more attention as an alternative therapy. Hence, this study reported the antimicrobial and antibiofilm potentials of cinnamon oil, silver nanoparticles (AgNPs), and their combination against multidrug-resistant (MDR) S. agalactiae recovered from clinical bovine mastitis in Egypt. RESULTS: Our findings revealed that 73% (146/200) of the examined milk samples collected from dairy cows with clinical mastitis were infected with Streptococci species. Of these, 9.59% (14/146) were identified as S. agalactiae and categorized as MDR. S. agalactiae isolates expressed four virulence genes (Hyl, cylE, scpB, and lmb) and demonstrated an ability to produce biofilms. Cinnamon oil showed high antimicrobial (MICs ≤0.063 µg /mL) and antibiofilm (MBIC50 = 4 µg/mL) potentials against planktonic and biofilms of S. agalactiae isolates, respectively. However, AgNPs showed reasonable antimicrobial (MICs ≤16 µg/mL) and relatively low antibiofilm (MBIC50 = 64 µg/mL) activities against screened isolates. Synergistic antimicrobial or additive antibiofilm interactions of cinnamon oil combined with AgNPs were reported for the first time. Scanning electron microscope (SEM) analysis revealed that biofilms of S. agalactiae isolates treated with cinnamon oil were more seriously damaged than observed in AgNPs cinnamon oil combination. Moreover, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) showed that cinnamon oil exerted a remarkable down-regulation of pili biosynthesis genes (pilA and pilB) and their regulator (rogB) against S. agalactiae biofilms, meanwhile the AgNPs cinnamon oil combination demonstrated a lower efficacy. CONCLUSIONS: This is an in vitro preliminary approach that documented the antibiofilm potential of cinnamon oil and the inhibitory activity of cinnamon oil and its combination with AgNPs against MDR S. agalactiae recovered from clinical mastitis. Further in vivo studies should be carried out in animal models to provide evidence of concept for implementing these alternative candidates in the treatment of dairy farms infected by streptococcal mastitis in the future.