Mobility of β-lactam resistance under ampicillin treatment in gut microbiota suffering from pre-disturbance

Ingestion of food- or waterborne antibiotic-resistant bacteria may lead to dissemination of antibiotic resistance genes (ARGs) in the gut microbiota. The gut microbiota often suffers from various disturbances. It is not clear whether and how disturbed microbiota may affect ARG mobility under antibiotic treatments. For proof of concept, in the presence or absence of streptomycin pre-treatment, mice were inoculated orally with a β-lactam-susceptible Salmonella enterica serovar Heidelberg clinical isolate (recipient) and a β-lactam resistant Escherichia coli O80:H26 isolate (donor) carrying a bla(CMY-2) gene on an IncI2 plasmid. Immediately following inoculation, mice were treated with or without ampicillin in drinking water for 7 days. Faeces were sampled, donor, recipient and transconjugant were enumerated, bla(CMY-2) abundance was determined by quantitative PCR, faecal microbial community composition was determined by 16S rRNA amplicon sequencing and cecal samples were observed histologically for evidence of inflammation. In faeces of mice that received streptomycin pre-treatment, the donor abundance remained high, and the abundance of S. Heidelberg transconjugant and the relative abundance of Enterobacteriaceae increased significantly during the ampicillin treatment. Co-blooming of the donor, transconjugant and commensal Enterobacteriaceae in the inflamed intestine promoted significantly (P<0.05) higher and possibly wider dissemination of the bla(CMY-2) gene in the gut microbiota of mice that received the combination of streptomycin pre-treatment and ampicillin treatment (Str–Amp) compared to the other mice. Following cessation of the ampicillin treatment, faecal shedding of S. Heidelberg transconjugant persisted much longer from mice in the Str–Amp group compared to the other mice. In addition, only mice in the Str–Amp group shed a commensal E. coli O2:H6 transconjugant, which carries three copies of the bla(CMY-2) gene, one on the IncI2 plasmid and two on the chromosome. The findings highlight the significance of pre-existing gut microbiota for ARG dissemination and persistence during and following antibiotic treatments of infectious diseases.