RESUMEN
Antimicrobial resistance (AMR) is a well-recognized, widespread, and growing issue of concern. With increasing incidence of AMR, the ability to respond quickly to infection with or exposure to an AMR pathogen is critical. Approaches that could accurately and more quickly identify whether a pathogen is AMR also are needed to more rapidly respond to existing and emerging biological threats. We examined proteins associated with paired AMR and antimicrobial susceptible (AMS) strains of Yersinia pestis and Francisella tularensis, causative agents of the diseases plague and tularemia, respectively, to identify whether potential existed to use proteins as signatures of AMR. We found that protein expression was significantly impacted by AMR status. Antimicrobial resistance-conferring proteins were expressed even in the absence of antibiotics in growth media, and the abundance of 10-20% of cellular proteins beyond those that directly confer AMR also were significantly changed in both Y. pestis and F. tularensis. Most strikingly, the abundance of proteins involved in specific metabolic pathways and biological functions was altered in all AMR strains examined, independent of species, resistance mechanism, and affected cellular antimicrobial target. We have identified features that distinguish between AMR and AMS strains, including a subset of features shared across species with different resistance mechanisms, which suggest shared biological signatures of resistance. These features could form the basis of novel approaches to identify AMR phenotypes in unknown strains.
RESUMEN
Burkholderia pseudomallei is a soil-dwelling bacterium endemic to Southeast Asia and northern Australia that causes the disease, melioidosis. Although the global genomic diversity of clinical B. pseudomallei isolates has been investigated, there is limited understanding of its genomic diversity across small geographic scales, especially in soil. In this study, we obtained 288 B. pseudomallei isolates from a single soil sample (~100g; intensive site 2, INT2) collected at a depth of 30cm from a site in Ubon Ratchathani Province, Thailand. We sequenced the genomes of 169 of these isolates that represent 7 distinct sequence types (STs), including a new ST (ST1820), based on multi-locus sequence typing (MLST) analysis. A core genome SNP phylogeny demonstrated that all identified STs share a recent common ancestor that diverged an estimated 796-1260 years ago. A pan-genomics analysis demonstrated recombination between clades and intra-MLST phylogenetic and gene differences. To identify potential differential virulence between STs, groups of BALB/c mice (5 mice/isolate) were challenged via subcutaneous injection (500 CFUs) with 30 INT2 isolates representing 5 different STs; over the 21-day experiment, eight isolates killed all mice, 2 isolates killed an intermediate number of mice (1-2), and 20 isolates killed no mice. Although the virulence results were largely stratified by ST, one virulent isolate and six attenuated isolates were from the same ST (ST1005), suggesting that variably conserved genomic regions may contribute to virulence. Genomes from the animal-challenged isolates were subjected to a bacterial genome-wide association study to identify genomic regions associated with differential virulence. One associated region is a unique variant of Hcp1, a component of the type VI secretion system, which may result in attenuation. The results of this study have implications for comprehensive sampling strategies, environmental exposure risk assessment, and understanding recombination and differential virulence in B. pseudomallei.
