Recovery of multi-antibiotic resistant Enterobacterales from retail fresh vegetables and the potential transfer of plasmid-mediated colistin resistance gene, mcr-1, in sprouts

Colistin is a last line therapeutic option in treatment of infections caused by extended spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae. Enterobacterales in fresh vegetables which are usually consumed raw could be reservoirs of multi-drug resistance bacteria. Moreover, the emergence of mobile colistin resistant gene, mcr-1, in Enterobacterales has become a global public health problem. Therefore, this research is to screen the presence of colistin and β-lactam resistance as well as associated genes from retailed fresh vegetables. The factors of effecting transfer frequency of mcr-1 gene in pathogenic Escherichia coli were also evaluated in broth media and food. Colistin resistant bacteria in lettuce and seed sprouts were detected by selective media, and their antibiotic susceptibility profile were determined. A mcr-1 positive bacteria (E. coli NCTC 13846) served as donor and two recipients (E. coli O157:H7 and E. coli O104:H4) were used for conjugative experiments in TSB media at 4, 25 and 37°C for up to 36 hours, as well as in germination process of seed sprouts. The aerobic plate count for lettuce and seed sprouts ranged from 3.88 to 8.47 log CFU/g and the population of bacteria capable of growth on MAC agar supplemented with colistin ranged from 1.55 to 5.76 log CFU/g. Isolates from MAC agar were identified as Serratia marcescens, Providencia, Morganella morganii, Enterobacter spp., Pseudomonas putida, Escherichia coli and Stenotrophomonas maltophilia. Extensively drug resistant bacteria were recovered, 8% of isolates were resistant to >5 antibiotics. The colistin resistance genes, mcr-1 to mcr-4, were not detected, however, blaSHV, blaTEM,and blaCTX-M group genes that encode for beta-lactamase were detected in several colistin tolerant bacteria. The transfer frequency was 10-7 to 10-8 per recipient cell for both two pathogenic E. coli at 25°C and 37°C. No transconjugants were detected at 4°C. However, higher transfer frequency (10⁻³ to 10⁻⁵ per recipient) of mcr-1 was occurred during mung bean sprouts germination. E. coli O104:H4 showed a better conjugative capability on mung bean sprouts than E. coli O157:H7. Susceptibility of several transconjugants to colistin was increased compared with donor cells. The spread of extensively antibiotic resistant bacteria through the global food supply from non-food animal derived products (fresh produce) is a potential human health concern. The horizontal gene transfer in liquid media could be underestimated which cannot represent a real situation. Food matrix is an important habitat for exchanging of the antibiotic resistant gene (ARGs) between microorganisms, potentially facilitating the spread of antibiotic resistance to human.Ph.D.Includes bibliographical reference