Abstract
The aim of this study was to screen for the presence of antimicrobial resistance genes within the saliva and faecal microbiomes of healthy adult human volunteers from five European countries. Two non-culture based approaches were employed to obviate potential bias associated with difficult to culture members of the microbiota. In a gene target-based approach, a microarray was employed to screen for the presence of over 70 clinically important resistance genes in the saliva and faecal microbiomes. A total of 14 different resistance genes were detected encoding resistances to six antibiotic classes (aminoglycosides, β-lactams, macrolides, sulphonamides, tetracyclines and trimethoprim). The most commonly detected genes were erm(B), bla TEM, and sul2. In a functional-based approach, DNA prepared from pooled saliva samples was cloned into Escherichia coli and screened for expression of resistance to ampicillin or sulphonamide, two of the most common resistances found by array. The functional ampicillin resistance screen recovered genes encoding components of a predicted AcrRAB efflux pump. In the functional sulphonamide resistance screen, folP genes were recovered encoding mutant dihydropteroate synthase, the target of sulphonamide action. The genes recovered from the functional screens were from the chromosomes of commensal species that are opportunistically pathogenic and capable of exchanging DNA with related pathogenic species. Genes identified by microarray were not recovered in the activity-based screen, indicating that these two methods can be complementary in facilitating the identification of a range of resistance mechanisms present within the human microbiome. It also provides further evidence of the diverse reservoir of resistance mechanisms present in bacterial populations in the human gut and saliva. In future the methods described in this study can be used to monitor changes in the resistome in response to antibiotic therapy.
Highlights
The human body serves as a host for a diverse range of commensal and symbiotic microorganisms, collectively termed the microbiota
There is evidence that antimicrobial use in humans and animals has had an impact upon the composition of the microbiome [5], antimicrobial resistance (AMR) genes have been detected in humans, animals, and in environments where there is little or no evidence of antibiotic use by man [6,7,8]
The resistome is important in that it acts as a reservoir of AMR genes that can reside in commensals or opportunistic pathogens and can be acquired by pathogens via horizontal gene transfer, and has the potential to interfere with therapeutic options following infection
Summary
The human body serves as a host for a diverse range of commensal and symbiotic microorganisms, collectively termed the microbiota. In a sequenced-based approach, the microbiome is shotgun sequenced and AMR genes identified by homology to known genes in reference databases (reviewed in [13]) These two methods only enable the detection of previously characterised genes and cannot fully explore the capacity of the resistome. The resultant clones are screened for resistance to selected antibiotics This approach enables resistance genes to be identified without prior knowledge of their sequence (reviewed in [14]) and has been used to recover known and novel AMR genes from, for example, soil [15,16], an activated sludge microbial community [17] and the human microbiome [18,19]
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