Spontaneous and evolutionary changes in the antibiotic resistance of Burkholderia cenocepacia observed by global gene expression analysis

Abstract

BackgroundBurkholderia cenocepacia is a member of the Burkholderia cepacia complex group of bacteria that cause infections in individuals with cystic fibrosis. B. cenocepacia isolate J2315 has been genome sequenced and is representative of a virulent, epidemic CF strain (ET12). Its genome encodes multiple antimicrobial resistance pathways and it is not known which of these is important for intrinsic or spontaneous resistance. To map these pathways, transcriptomic analysis was performed on: (i) strain J2315 exposed to sub-inhibitory concentrations of antibiotics and the antibiotic potentiator chlorpromazine, and (ii) on spontaneous mutants derived from J2315 and with increased resistance to the antibiotics amikacin, meropenem and trimethoprim-sulfamethoxazole. Two pan-resistant ET12 outbreak isolates recovered two decades after J2315 were also compared to identify naturally evolved gene expression changes.ResultsSpontaneous resistance in B. cenocepacia involved more gene expression changes and different subsets of genes than those provoked by exposure to sub inhibitory concentrations of each antibiotic. The phenotype and altered gene expression in the resistant mutants was also stable irrespective of the presence of the priming antibiotic. Both known and novel genes involved in efflux, antibiotic degradation/modification, membrane function, regulation and unknown functions were mapped. A novel role for the phenylacetic acid (PA) degradation pathway genes was identified in relation to spontaneous resistance to meropenem and glucose was found to repress their expression. Subsequently, 20 mM glucose was found to produce greater that 2-fold reductions in the MIC of multiple antibiotics against B. cenocepacia J2315. Mutation of an RND multidrug efflux pump locus (BCAM0925-27) and squalene-hopene cyclase gene (BCAS0167), both upregulated after chlorpromazine exposure, confirmed their role in resistance. The recently isolated outbreak isolates had altered the expression of multiple genes which mirrored changes seen in the antibiotic resistant mutants, corroborating the strategy used to model resistance. Mutation of an ABC transporter gene (BCAS0081) upregulated in both outbreak strains, confirmed its role in B. cenocepacia resistance.ConclusionsGlobal mapping of the genetic pathways which mediate antibiotic resistance in B. cenocepacia has revealed that they are multifactorial, identified potential therapeutic targets and also demonstrated that putative catabolite repression of genes by glucose can improve antibiotic efficacy.

Comparative Metabolic Systems Analysis of Pathogenic Burkholderia

“…Here, we have thoroughly characterized the metabolism of the type strains of B. cenocepacia and B. multivorans. Our network reconstructions can be used to contextualize high-throughput transcriptomic and proteomic data (58,(74)(75)(76)(77) to provide further insight into gene regulation and downstream phenotypes. Additionally, our reconstructions can be used as established starting points to analyze pathogenesis and physiology of the growing list of sequenced Burkholderia strains, including both clinical and environmental isolates (78).…”

Gene Expression Profiling of Burkholderia cenocepacia at the Time of Cepacia Syndrome: Loss of Motility as a Marker of Poor Prognosis?

“…Nevertheless, the expression of QS genes in isolates from patients A and B matched the microarray data. Virulence factors, such as motility, T3SS, and exopolysaccharides, are QS dependent, and their orchestrated regulation in Bcc bacteria was previously reported (19,20). In the context of our test condition (the biological origin of the isolate), we presumed that their significant gene expression changes had important implications for survival in the bloodstream and contributed to bacterial cell defense against the host immune system.…”

Analysis of genomic characters reveals that four distinct gene clusters are correlated with different functions in Burkholderia cenocepacia AU 1054

“…Moreover, the Chi-square independence test indicated that the 73 genes were significantly associated with environment conditions (p <0.001). Higher transcriptional activities of these induced genes under CF-like conditions could indicate that there are adaptive changes for these genes, which facilitates this species to transmit infections among CF patients (Drevinek et al 2008;Sass et al 2011). Taken together, the CF environment transition in B. cenocepacia could give rise to the codon usage bias in these highly expressed genes, and these highly expressed genes showed environmental adaptation to the CF environment.…”

“…Furthermore, we searched the homology of these 826 genes against some functionally confirmed drug-resistance related genes at NCBI (www.ncbi.nlm.nih.gov), and we detected a substantial number of related genes (Supplementary Table S3), e.g., a gene for MFS_1 which belongs to the MFS that facilitates transport across cytoplasmic or internal membranes of a variety of substrates (Holden et al 2009); glnP, which encodes an ABC transporter permease protein that is associated with multidrug resistance (Moussatova et al 2008); emrB, which encodes a member of the drug resistance transporter EmrB/QacA subfamily (Ma et al 1994) and matE, which encodes a MATE efflux family protein that functions in MATE (Holden et al 2009). In responses to treatment with antimicrobial compounds, genes from drug resistance-related systems are under selection pressure in the long term (Cowen and Lindquist 2005;Sass et al 2011). Then a rapid evolution of virulence and drug resistance-related genes will emerge in the infectious pathogens (Cowen and Lindquist 2005).…”