Milk from healthy or infected cattle as a source of multi-drug resistant, AmpC β-lactamase-producing Escherichia coli

Abstract

Drinking raw unpasteurized milk or products made from raw milk is a recent trend observed among the people due to their belief in enhancing nutritional properties and better taste. It poses a great risk of microbial transmission along with antibiotic resistance gene pool (extended spectrum beta-lactamase or ampC) among the consumers. The study was conducted to detect the occurrence of AmpC producing E. coli in raw milk samples collected from healthy and mastitic cattle in West Bengal. In total, 450 cow milk samples were collected from all the six agroclimatic zones of West Bengal. The samples were collected from healthy as well as infected cattle suffering with clinical or subclinical mastitis, irrespective of age and breed. The collected milk samples were tested with MAST INDEX reagent for the detection of sub-clinical mastitis. E. coli isolates obtained from cattle milk samples (n=450) were subjected to cefoxitin-cloxacillin double disc synergy test and PCR based confirmation of blaAmpC, class I integron and CITM genes. All the ACBL producing E. coli isolates were subjected for their resistance to other antimicrobials. All the levofloxacin and Co-Trimoxazole resistant ACBL producing isolates were studied for PMQR, sul1, sul2, sul3 genes. E. coli was isolated from 205 collected milk samples (205/450, 45.6%). The phenotypical test detected 231/383 isolates (60.3%) were AMPC producers which were also found to possess bla AmpC. Comparison of ACBL-producing E. coli occurrence in cattle with mastitis, sub-clinical mastitis and healthy cattle revealed significantly higher occurrence rate in infected cattle than the healthy ones (p < 0.05). Phenotypical antibiotic resistance pattern of the isolates showed resistance against penicillin-G (100%), cefoxitin (100%), ampicillin (42%), co-trimoxazole (13.5%) and levofloxacin (5.2%). Out of 30 co-trimoxazole resistant isolates identified, 25 (83%) and seven (30.4%) isolates possessed sul1 and sul2 gene, respectively, whereas, four (13.3%) isolates possessed both sul1 and sul2 genes. Three levofloxacin resistant isolates possessed qnrS gene. This is the first systemic study with substantial numbers of milk samples from healthy and infected cattle depicting occurrence of ACBL-producing E. coli and linkage present in the isolates between the genes encoding ACBL and quinolone or sulphonamide resistance.