Dataset Information

Molecular evidence of Burkholderia pseudomallei genotypes based on geographical distribution.

ABSTRACT: Background. Central intermediary metabolism (CIM) in bacteria is defined as a set of metabolic biochemical reactions within a cell, which is essential for the cell to survive in response to environmental perturbations. The genes associated with CIM are commonly found in both pathogenic and non-pathogenic strains. As these genes are involved in vital metabolic processes of bacteria, we explored the efficiency of the genes in genotypic characterization of Burkholderia pseudomallei isolates, compared with the established pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) schemes. Methods. Nine previously sequenced B. pseudomallei isolates from Malaysia were characterized by PFGE, MLST and CIM genes. The isolates were later compared to the other 39 B. pseudomallei strains, retrieved from GenBank using both MLST and sequence analysis of CIM genes. UniFrac and hierachical clustering analyses were performed using the results generated by both MLST and sequence analysis of CIM genes. Results. Genetic relatedness of nine Malaysian B. pseudomallei isolates and the other 39 strains was investigated. The nine Malaysian isolates were subtyped into six PFGE profiles, four MLST profiles and five sequence types based on CIM genes alignment. All methods demonstrated the clonality of OB and CB as well as CMS and THE. However, PFGE showed less than 70% similarity between a pair of morphology variants, OS and OB. In contrast, OS was identical to the soil isolate, MARAN. To have a better understanding of the genetic diversity of B. pseudomallei worldwide, we further aligned the sequences of genes used in MLST and genes associated with CIM for the nine Malaysian isolates and 39 B. pseudomallei strains from NCBI database. Overall, based on the CIM genes, the strains were subtyped into 33 profiles where majority of the strains from Asian countries were clustered together. On the other hand, MLST resolved the isolates into 31 profiles which formed three clusters. Hierarchical clustering using UniFrac distance suggested that the isolates from Australia were genetically distinct from the Asian isolates. Nevertheless, statistical significant differences were detected between isolates from Malaysia, Thailand and Australia. Discussion. Overall, PFGE showed higher discriminative power in clustering the nine Malaysian B. pseudomallei isolates and indicated its suitability for localized epidemiological study. Compared to MLST, CIM genes showed higher resolution in distinguishing those non-related strains and better clustering of strains from different geographical regions. A closer genetic relatedness of Malaysian isolates with all Asian strains in comparison to Australian strains was observed. This finding was supported by UniFrac analysis which resulted in geographical segregation between Australia and the Asian countries.

SUBMITTER: Zulkefli NJ

PROVIDER: S-EPMC4797763 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Molecular evidence of Burkholderia pseudomallei genotypes based on geographical distribution.

Zulkefli Noorfatin Jihan NJ Mariappan Vanitha V Vellasamy Kumutha Malar KM Chong Chun Wie CW Thong Kwai Lin KL Ponnampalavanar Sasheela S Vadivelu Jamuna J Teh Cindy Shuan Ju CS

PeerJ 20160315

Background. Central intermediary metabolism (CIM) in bacteria is defined as a set of metabolic biochemical reactions within a cell, which is essential for the cell to survive in response to environmental perturbations. The genes associated with CIM are commonly found in both pathogenic and non-pathogenic strains. As these genes are involved in vital metabolic processes of bacteria, we explored the efficiency of the genes in genotypic characterization of Burkholderia pseudomallei isolates, compar ...[more]

PMID: 26998408

Similar Datasets

Project description:BackgroundBurkholderia pseudomallei is a Gram-negative bacterium found in soil and water in many tropical countries. It causes melioidosis, a potentially fatal infection first described in 1911 in Myanmar. Melioidosis is a common cause of sepsis and death in South and South-east Asia, but it is rarely diagnosed in Myanmar. We conducted a nationwide soil study to identify areas where B. pseudomallei is present.Methodology/principal findingsWe collected soil samples from 387 locations in all 15 states and regions of Myanmar between September 2017 and June 2019. At each site, three samples were taken at each of three different depths (30, 60 and 90 cm) and were cultured for B. pseudomallei separately, along with a pooled sample from each site (i.e. 10 cultures per site). We used a negative binomial regression model to assess associations between isolation of B. pseudomallei and environmental factors (season, soil depth, soil type, land use and climate zones). B. pseudomallei was isolated in 7 of 15 states and regions. Of the 387 sites, 31 (8%) had one or more positive samples and of the 3,870 samples cultured, 103 (2.7%) tested positive for B. pseudomallei. B. pseudomallei was isolated more frequently during the monsoon season [RR-2.28 (95% CI: 0.70-7.38)] and less in the hot dry season [RR-0.70 (95% CI: 0.19-2.56)] compared to the cool dry season, and in the tropical monsoon climate zone [RR-2.26; 95% CI (0.21-6.21)] compared to the tropical dry winter climate zone. However, these associations were not statistically significant. B. pseudomallei was detected at all three depths and from various soil types (clay, silt and sand). Isolation was higher in agricultural land (2.2%), pasture land (8.5%) and disused land (5.8%) than in residential land (0.4%), but these differences were also not significant.Conclusion/significanceThis study confirms a widespread distribution of B. pseudomallei in Myanmar. Clinical studies should follow to obtain a better picture of the burden of melioidosis in Myanmar.

Project description:BACKGROUND: Leucine aminopeptidase (LAP) has been known to be a housekeeping protease, DNA-binding protein and repressor or activator in the operon regulation of virulence-associated genes in several bacterial species. LAP activity was consistently detected in overnight cultures of Burkholderia pseudomallei, the causative agent of melioidosis and this enzyme was partially purified and characterised in this study. The intra- and inter-species nucleotide and deduced amino acid sequence variation of LAP encoding gene (pepA) was determined. A pepA/PCR-RFLP assay was designed to facilitate the identification of major LAP sequence types amongst clinical and environmental isolates of B. pseudomallei. RESULTS: LAP activity was detected in B. pseudomallei culture supernantants by zymographic analysis. Optimum activity was at pH 9 and stable at 50[degree sign]C. Enhanced enzymatic activity was observed in the presence of metallic ions Mg2+, Ca2+, Na+ and K+. LAP activity was inhibited by EDTA, 1,10-phenanthroline, amastatin, Mn2+ and Zn2+. Sequence analysis of the complete nucleotide and deduced amino acid sequences of LAP-encoding (pepA) gene showed close genetic relatedness to B. mallei (similarity 99.7%/99.6%), but not with B. thailandensis (96.4%/96.4%). Eight pepA sequence types were identified by comparison with a 596 bp DNA fragment encompassing central regions of the pepA gene. A pepA/PCR-RFLP was designed to differentiate pepA sequence types. Based on restriction analysis with StuI and HincII enzymes of the amplified pepA gene, clinical and environmental isolates showed different predominant RFLP types. Type I was the most predominant type amongst 71.4% (65/91) of the clinical isolates, while Type II was predominant in 55.6% (5/9) of the environmental isolates. CONCLUSIONS: This study showed that LAP is a secretory product of B. pseudomallei with features similar to LAP of other organisms. Identification of major LAP sequence types of B. pseudomallei was made possible based on RFLP analysis of the pepA gene. The high LAP activity detected in both B. pseudomallei and B. thailandensis, suggests that LAP is probably a housekeeping enzyme rather than a virulence determinant.

Project description:Burkholderia pseudomallei is the etiological agent of melioidosis. Because of the bacterium's intrinsic resistance and propensity to establish latent infections, melioidosis therapy is complicated and prolonged. Newer generation ?-lactams, specifically ceftazidime, are used for acute phase therapy, but resistance to this cephalosporin has been observed. The chromosomally encoded penA gene encodes a putative twin arginine translocase (TAT)-secreted ?-lactamase, and penA mutations have been implicated in ceftazidime resistance in clinical isolates. However, the role of PenA in resistance has not yet been systematically studied in isogenetic B. pseudomallei mutant backgrounds. We investigated the effects of penA deletion, point mutations, and up-regulation, as well as tat operon deletion and PenA TAT-signal sequence mutations. These experiments were made possible by employing a B. pseudomallei strain that is excluded from Select Agent regulations. Deletion of penA significantly (>4-fold) reduced the susceptibility to six of the nine ?-lactams tested and ?16-fold for ampicillin, amoxicillin, and carbenicillin. Overexpression of penA by single-copy, chromosomal expression of the gene under control of the inducible P(tac) promoter, increased resistance levels for all ?-lactams tested 2- to 10-fold. Recreation of the C69Y and P167S PenA amino acid substitutions previously observed in resistant clinical isolates increased resistance to ceftazidime by ?85- and 5- to 8-fold, respectively. Similarly, a S72F substitution resulted in a 4-fold increase in resistance to amoxicillin and clavulanic acid. Susceptibility assays with PenA TAT-signal sequence and ?tatABC mutants, as well as Western blot analysis, confirmed that PenA is a TAT secreted enzyme and not periplasmic but associated with the spheroplastic cell fraction. Lastly, we determined that two LysR-family regulators encoded by genes adjacent to penA do not play a role in transcriptional regulation of penA expression.

Dataset Information

Molecular evidence of Burkholderia pseudomallei genotypes based on geographical distribution.

Publications

Molecular evidence of Burkholderia pseudomallei genotypes based on geographical distribution.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets