Abstract
The infant gut microbiota develops rapidly during the first 2 years of life, acquiring microorganisms from diverse sources. During this time, significant opportunities exist for the infant to acquire antibiotic resistant bacteria, which can become established and constitute the infant gut resistome. With increased antibiotic resistance limiting our ability to treat bacterial infections, investigations into resistance reservoirs are highly pertinent. This study aimed to explore the nascent resistome in antibiotically-naïve infant gut microbiomes, using a combination of metagenomic approaches. Faecal samples from 22 six-month-old infants without previous antibiotic exposure were used to construct a pooled metagenomic library, which was functionally screened for ampicillin and gentamicin resistance. Our library of ∼220Mb contained 0.45 ampicillin resistant hits/Mb and 0.059 gentamicin resistant hits/Mb. PCR-based analysis of fosmid clones and uncloned metagenomic DNA, revealed a diverse and abundant aminoglycoside and β-lactam resistance reservoir within the infant gut, with resistance determinants exhibiting homology to those found in common gut inhabitants, including Escherichia coli, Enterococcus sp., and Clostridium difficile, as well as to genes from cryptic environmental bacteria. Notably, the genes identified differed from those revealed when a sequence-driven PCR-based screen of metagenomic DNA was employed. Carriage of these antibiotic resistance determinants conferred substantial, but varied (2–512x), increases in antibiotic resistance to their bacterial host. These data provide insights into the infant gut resistome, revealing the presence of a varied aminoglycoside and β-lactam resistance reservoir even in the absence of selective pressure, confirming the infant resistome establishes early in life, perhaps even at birth.
Highlights
There is growing concern that we are rapidly approaching a post-antibiotic era
In a previous study, we have demonstrated that the infant microbiota becomes dominated by Proteobacteria following ampicillin and gentamicin administration in the first 48 hours of life [13], which may be due to the known high prevalence of antibiotic resistant species within this phylum [14]
Identification of gentamicin and ampicillin resistant clones within the infant gut microbiome using a functional metagenomic bank Ten white clones were picked at random from LB agar plates containing 12.5 mg/ml chloramphenicol, IPTG and X-gal and underwent restriction digestion using PstI and NdeI restriction enzymes
Summary
There is growing concern that we are rapidly approaching a post-antibiotic era. As a result every effort is being made to discover and investigate antibiotic resistance reservoirs with the aim of limiting the selection for, or dissemination of, antibiotic resistance genes. In a previous study, we have demonstrated that the infant microbiota becomes dominated by Proteobacteria following ampicillin and gentamicin administration in the first 48 hours of life [13], which may be due to the known high prevalence of antibiotic resistant species within this phylum [14]. Such findings suggest that the infant gut microbiota, though immature and in constant flux, can be a source of resistant bacteria which can become dominant following antibiotic exposure. Given the instability of the infant microbiota in early life, there is considerable opportunity for the infant gut to acquire resistant populations which, if they become established, could have significant effects on shaping the composition of the microbiota later in life [15]
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