Functional and genetic characterization of an incF-type multidrug resistance plasmid isolated from fresh spinach.
The presence of antibiotic-resistant bacteria and clinically-relevant antibiotic resistance genes within raw foods is an on-going food safety concern. It is particularly important to be aware of the microbial quality of fresh produce because foods such as leafy greens including lettuce and spinach are minimally processed and often consumed raw therefore they often lack a microbial inactivation step. This study characterizes the genetic and functional aspects of a mobile, multidrug resistance plasmid, pLGP4, isolated from fresh spinach bought from a farmers market. pLGP4 was isolated using a bacterial conjugation approach. The functional characteristics of the plasmid were determined using multidrug resistance profiling and plasmid stability assays. pLGP4 was resistant to six of the eight antibiotics tested and included ciprofloxacin and meropenem. The plasmid was stably maintained within host strains in the absence of an antibiotic selection. The plasmid DNA was sequenced using an Illumina MiSeq high throughput sequencing approach and assembled into contigs using SPAdes. PCR mapping and Sanger DNA sequencing of PCR amplicons was used to complete the plasmid DNA sequence. Comparative sequence analysis determined that the plasmid was similar to plasmids that have been frequently associated with multidrug resistant clinical isolates of Klebsiella spp. DNA sequence analysis showed pLGP4 harboured qnrB1 and several other antibiotic resistance genes including three {beta}-lactamases: blaTEM-1, blaCTX-M-15 and blaOXA-1. The detection of a multidrug-resistant, clinically-relevant plasmid on fresh spinach emphasizes the importance for vegetable producers to implement evidence-based food safety approaches into their production practises to ensure the food safety of leafy greens.