A reservoir of 'historical' antibiotic resistance genes in remote pristine Antarctic soils

Abstract

Background: Soil bacteria naturally produce antibiotics as a competitive mechanism, with a concomitant evolution, and exchange by horizontal gene transfer, of a range of antibiotic resistance mechanisms. Surveys of bacterial resistance elements in edaphic systems have originated primarily from human-impacted environments, with relatively little information from remote and pristine environments, where the resistome may comprise the ancestral gene diversity.

Methods: We used shotgun metagenomics to assess antibiotic resistance gene (ARG) distribution in 17 pristine and remote Antarctic surface soils within the undisturbed Mackay Glacier region. We also interrogated the phylogenetic placement of ARGs compared to environmental ARG sequences and tested for the presence of horizontal gene transfer elements flanking ARGs.

Results: In total, 177 naturally occurring ARGs were identified, most of which encoded single or multi-drug efflux pumps. Resistance mechanisms for the inactivation of aminoglycosides, chloramphenicol and β-lactam antibiotics were also common. Gram-negative bacteria harboured most ARGs (71%), with fewer genes from Gram-positive Actinobacteria and Bacilli (Firmicutes) (9%), reflecting the taxonomic composition of the soils. Strikingly, the abundance of ARGs per sample had a strong, negative correlation with species richness (r = - 0.49, P < 0.05). This result, coupled with a lack of mobile genetic elements flanking ARGs, suggests that these genes are ancient acquisitions of horizontal transfer events.

Conclusions: ARGs in these remote and uncontaminated soils most likely represent functional efficient historical genes that have since been vertically inherited over generations. The historical ARGs in these pristine environments carry a strong phylogenetic signal and form a monophyletic group relative to ARGs from other similar environments.

Keywords: Antarctica; Antibiotic resistance genes; Metagenomics; Soil resistome.

Fig. 1

Satellite image of the Mackay Glacier ecotone with the 17 sampling sites indicated. Source: Landsat Image Mosaic of Antarctica (LIMA) Digital Database and Google Earth

Fig. 2

ARG frequencies across sampled sites. Number of different ARGs indicated in green, with the number of unique ARGs displayed in red, axis on the left. The relative ARG abundances are shown as a black line, axis on the right

Fig. 3

Co-occurrence network of ARG mechanisms showing resistance mechanisms encoded by diverse soil bacterial phyla. Phyla from all 17 soils that were assigned an ARG are presented here (diamond-shaped nodes), with significant co-occurrences with a specific ARG (circles) indicated (edges)

Fig. 4

ARG family frequencies across sampled sites. The number of different ARG families are indicated in green, with the number of unique ARG families displayed in red, axis on the left. The black line represents the relative ARG family abundance, axis on the right

Fig. 5

The number of ARGs and number of species per site. Linear model indicated in red and lowess in blue (Pearson’s correlation r = − 0.49, P < 0.05)

Fig. 6

Unrooted Bayesian phylogeny of microbial antibiotic resistance gene sequences identified across the 17 Antarctic soil metagenomes. a Dihydrofolate reductase (drfE) genes, b Macrolide transporter ATP-binding permease proteins, and c major facilitator superfamily transporter sequences (rosA) are shown; blue denotes ORFs identified in the Antarctic metagenomes. Reference sequences are provided with accession numbers, and the protein sequence present in noradab is shown in orange