Factors involved in the proliferation of blaCMY-2 plasmid-bearing Escherichia coli in cattle

Abstract

Third-generation cephalosporins are a crucial component of the antibiotic arsenal for human and veterinary medicine. Resistance to third-generation cephalosporins is often conveyed by blaCMY-2 plasmids. The prevalence of blaCMY-2 plasmid-bearing Salmonella and Escherichia coli and ceftiofur (third-generation cephalosporin) use in livestock has been implicated in this process. Nevertheless, injection of ceftiofur results in limited effect on the enteric flora and the majority of drug excreted in the urine. We employed a series of in vitro and in vivo passage studies showing that blaCMY-2 plasmids impose a fitness cost on their bacterial host and this cost is not mitigated by conjugation or segregation systems (Chapter I). Thus, selection is required to maintain blaCMY-2 plasmids in a population. After administration to cattle ceftiofur is quickly converted in to pharmacologically active ceftiofur metabolite (CFM). Because most (~ 70%) CFM is excreted through urine, we hypothesized that selection for cefR E. coli occurs primarily ex vivo rather than in vivo. We first found that when mixed in soil ceftiofur remains bioavailable but some antibiotics (e.g. tetracycline) are not (Chapter II). To further evaluate the fate of CFM ex vivo, we used matrices consisting of soil and feces with urine containing CFM and examined the factors that impact the longevity of CFM. Our findings showed that CFM remains bioavailable in soil for a longer period with higher concentration (as expected) and at colder temperatures. CFM bioavailability is retained much longer in autoclaved soil indicating that the soil microflora is probably responsible for eventual degradation of CFM. Importantly, our findings show that CFM confers a significant numerical (1 to 1.5 log10 cfu per g matrix at 23°C) advantage to ceftiofur resistant bacteria in the soil and this advantage maintained well-beyond the period of CFM bioavailability; exposure to CFM extends survivorship of cefR E. coli an estimated 80 days compared to non-exposed populations. We also showed that exposing dairy calves to bedding with blaCMY-2 positive E. coli is sufficient for transmission (Chapter III). Consequently, the numerical advantage conferred by exposure to CFM in soil is likely to increase the likelihood transmitting cefR E. coli back to livestock.