Abstract
Chicken intestinal Escherichia coli are a reservoir for virulence and antimicrobial resistance (AMR) genes that are often carried on incompatibility group F (IncF) plasmids. The rapid transfer of these plasmids between bacteria in the gut contributes to the emergence of new multidrug-resistant and virulent bacteria that threaten animal agriculture and human health. Thus, the aim of the present study was to determine whether live bacterial prophylactics could affect the distribution of large virulence plasmids and AMR in the intestinal tract and the potential role of smRNA in this process. In this study, we tested ∼100 randomly selected E. coli from pullet feces (n = 3 per group) given no treatment (CON), probiotics (PRO), a live Salmonella vaccine (VAX), or both (P + V). E. coli isolates were evaluated via plasmid profiles and several phenotypic (siderophore production and AMR), and genotypic (PCR for virulence genes and plasmid typing) screens. P + V isolates exhibited markedly attenuated siderophore production, lack of AMR and virulence genes, which are all related to the loss of IncF and ColV plasmids (P < 0.0001). To identify a causal mechanism, we evaluated smRNA levels in the ceca mucus and found a positive association between smRNA concentrations and plasmid content, with both being significantly reduced in P + V birds compared to other groups (P < 0.01). To test this positive association between IncF plasmid transfer and host smRNA concentration, we evenly pooled smRNA per group and treated E. coli mating pairs with serial concentrations of smRNA in vitro. Higher smRNA concentrations resulted in greater rates of IncF plasmid transfer between E. coli donors (APEC O2 or VAX isolate IA-EC-001) and recipient (HS-4) (all groups; P < 0.05). Finally, RNAHybrid predictive analyses detected several chicken miRNAs that hybridize with pilus assembly and plasmid transfer genes on the IncF plasmid pAPEC-O2-R. Overall, we demonstrated P + V treatment reduced smRNA levels in the chicken ceca, which was associated with a reduction in potentially virulent E. coli. Furthermore, we propose a novel mechanism in which intestinal smRNAs signal plasmid exchange between E. coli. Investigations to understand the changes in bacterial gene expression as well as smRNAs responsible for this phenomenon are currently underway.
Highlights
Plasmids are mobile genetic elements that can bolster a host bacterium’s fitness in harsh environments like the gastrointestinal tract by carrying genes encoding antimicrobial resistance (AMR) (Rolain, 2013), iron acquisition factors (Deriu et al, 2013), and antimicrobiallike products colicin (Gordon et al, 1998)
The aim of this study was to determine whether live prophylactics affect the level of E. coli carrying plasmids and genes encoding AMR and extraintestinal pathogenic E. coli (ExPEC) virulence factors in the chicken intestine as well as whether this effect is correlated with small RNA (smRNA)
Proceeding with E. coli isolates from each group for screening, we found a significant decrease in siderophore production via CAS agar in P + V E. coli isolates (P < 0.0001; Figure 1)
Summary
Plasmids are mobile genetic elements that can bolster a host bacterium’s fitness in harsh environments like the gastrointestinal tract by carrying genes encoding AMR (Rolain, 2013), iron acquisition factors (Deriu et al, 2013), and antimicrobiallike products colicin (Gordon et al, 1998). Many of these factors are commonly found in intestinal E. coli and other Enterobacteriaceae on narrow-range plasmids like incompatibility group F (IncF) (Johnson et al, 2005, 2007, 2010; Yang et al, 2015). These factors may have negative consequences for the animal host, as a transfer of these genes can increase bacterial virulence potential and/or generate AMR pathogens (Johnson et al, 2010). Conjugation inhibitors like unsaturated fatty acids (Getino et al, 2015) have been postulated as a means to reduce plasmid transfer. A greater understanding of which host factors influence bacterial conjugation as well as treatments to achieve this are imperative
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