Rapid detection and molecular epidemiology of -lactamase producing Enterobacteriaceae isolated from food animals and in-contact humans in Nigeria

The emergence and spread of beta-lactamase-producing Enterobacteriaceae poses a significant threat to public health, necessitating the rapid detection and investigation of the molecular epidemiology of these pathogens. We modified a multiplex real-time (RT)-PCR to concurrently detect beta-lactamase genes (bla(CTX-M), bla(TEM), and bla(SHV)) and Enterobacteriaceae 16S ribosomal RNA. qPCR probes and primers were validated using control isolates, and the sensitivity and specificity assessed. The optimised multiplex qPCR was used to screen 220 non-clinical Enterobacteriaceae from food animals and in-contact humans in Southeast Nigeria selected on cefotaxime-supplemented agar plates. Binary logistic regression was used to explore factors associated with the presence of the bla(TEM) and bla(SHV) genes in these isolates, and a subset of isolates from matched sampling sites and host species were whole genome sequenced, and their antimicrobial resistance (AMR) and plasmid profiles determined. The sensitivity and specificity of the qPCR assay was 100%. All isolates (220/220) were positive for Enterobacteriaceae ribosomal 16S rRNA and bla(CTX-M), while 66.4% (146/220) and 9% (20/220) were positive for bla(TEM) and bla(SHV), respectively. The prevalence of bla(TEM) and bla(SHV) varied across different sampling sites (farm, animal market and abattoirs). Isolates from Abia state were more likely to harbour bla(TEM) (OR = 2.3, p = 0.04) and bla(SHV) (OR = 5.12,p = 0.01) than isolates from Ebonyi state; bla(TEM) was more likely to be detected in isolates from food animals than humans (OR = 2.34, p = 0.03), whereas the reverse was seen for bla(SHV) (OR = 7.23, p = 0.02). Furthermore, Klebsiella and Enterobacter isolates harboured more AMR genes than Escherichia coli, even though they were isolated from the same sample. We also identified pan resistant Klebsiella harbouring resistance to ten classes of antimicrobials and disinfectant. Therefore, we recommend ESKAPE pathogens are included in AMR surveillance in future and suggest qPCRs be utilised for rapid screening of Enterobacteriaceae from human and animal sources.