Phylogeographic reconstruction of a bacterial species with high levels of lateral gene transfer

Background Phylogeographic reconstruction of some bacterial populations is hindered by low diversity coupled with high levels of lateral gene transfer. A comparison of recombination levels and diversity at seven housekeeping genes for eleven bacterial species, most of which are commonly cited as having high levels of lateral gene transfer shows that the relative contributions of homologous recombination versus mutation for Burkholderia pseudomallei is over two times higher than for Streptococcus pneumoniae and is thus the highest value yet reported in bacteria. Despite the potential for homologous recombination to increase diversity, B. pseudomallei exhibits a relative lack of diversity at these loci. In these situations, whole genome genotyping of orthologous shared single nucleotide polymorphism loci, discovered using next generation sequencing technologies, can provide very large data sets capable of estimating core phylogenetic relationships. We compared and searched 43 whole genome sequences of B. pseudomallei and its closest relatives for single nucleotide polymorphisms in orthologous shared regions to use in phylogenetic reconstruction. Results Bayesian phylogenetic analyses of >14,000 single nucleotide polymorphisms yielded completely resolved trees for these 43 strains with high levels of statistical support. These results enable a better understanding of a separate analysis of population differentiation among >1,700 B. pseudomallei isolates as defined by sequence data from seven housekeeping genes. We analyzed this larger data set for population structure and allele sharing that can be attributed to lateral gene transfer. Our results suggest that despite an almost panmictic population, we can detect two distinct populations of B. pseudomallei that conform to biogeographic patterns found in many plant and animal species. That is, separation along Wallace's Line, a biogeographic boundary between Southeast Asia and Australia. Conclusion We describe an Australian origin for B. pseudomallei, characterized by a single introduction event into Southeast Asia during a recent glacial period, and variable levels of lateral gene transfer within populations. These patterns provide insights into mechanisms of genetic diversification in B. pseudomallei and its closest relatives, and provide a framework for integrating the traditionally separate fields of population genetics and phylogenetics for other bacterial species with high levels of lateral gene transfer

The Core and Accessory Genomes of Burkholderia pseudomallei: Implications for Human Melioidosis

Natural isolates of Burkholderia pseudomallei (Bp), the causative agent of melioidosis, can exhibit significant ecological flexibility that is likely reflective of a dynamic genome. Using whole-genome Bp microarrays, we examined patterns of gene presence and absence across 94 South East Asian strains isolated from a variety of clinical, environmental, or animal sources. 86% of the Bp K96243 reference genome was common to all the strains representing the Bp “core genome”, comprising genes largely involved in essential functions (eg amino acid metabolism, protein translation). In contrast, 14% of the K96243 genome was variably present across the isolates. This Bp accessory genome encompassed multiple genomic islands (GIs), paralogous genes, and insertions/deletions, including three distinct lipopolysaccharide (LPS)-related gene clusters. Strikingly, strains recovered from cases of human melioidosis clustered on a tree based on accessory gene content, and were significantly more likely to harbor certain GIs compared to animal and environmental isolates. Consistent with the inference that the GIs may contribute to pathogenesis, experimental mutation of BPSS2053, a GI gene, reduced microbial adherence to human epithelial cells. Our results suggest that the Bp accessory genome is likely to play an important role in microbial adaptation and virulence

Innate Immune Responses of Pulmonary Epithelial Cells to Burkholderia pseudomallei Infection

10.1371/journal.pone.0007308PLoS ONE410

The study of pulmonary innate immune responses against Burkholderia pseudomallei

Master'sMASTER OF SCIENC

Motor insurance in Singapore : problems and prescriptions

Motor insurance makes up 32% of the total insurance business in Singapore. On statistics , the loss ratio is 62.7%. However, claims is only one aspect of the business expense. Overall, motor insurers are not making enough premium revenue to pay claims, administrative expenses, acquisition expenses. This report seeks to identify and examine the various reasons why motor insurers are suffering from deteriorating results. It emphasizes that immediate action is needed to avoid the erosion of profits from the motor insurance market. Furthermore, likely effects of the current situation are studied and this report attempts to recommend possible solutions to the problems.BUSINES

In vitro quantification of pigment production and transfer in 2D co-cultures and 3D skin organotypic

Summary: Here, we present a protocol to set up and study 2D keratinocyte-melanocyte co-cultures and 3D full-thickness human skin equivalents. We describe steps for culturing of keratinocyte and melanocyte lines and the establishment of both 2D and 3D co-cultures. The cultures are utilized to measure melanin content and investigate mechanisms driving melanin production and transfer, through flow cytometry and immunohistochemistry. Culture conditions are highly amendable to different conditions, and analysis is simple and objective—thus allowing for medium to high throughput.For complete details on the use and execution of this protocol, please refer to Ng et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics

Genomic patterns of pathogen evolution revealed by comparison of Burkholderia pseudomallei, the causative agent of melioidosis, to avirulent Burkholderia thailandensis

Background: The Gram-negative bacterium Burkholderia pseudomallei (Bp) is the causative agent of the human disease melioidosis. To understand the evolutionary mechanisms contributing to Bp virulence, we performed a comparative genomic analysis of Bp K96243 and B. thailandensis (Bt) E264, a closely related but avirulent relative. Results: We found the Bp and Bt genomes to be broadly similar, comprising two highly syntenic chromosomes with comparable numbers of coding regions (CDs), protein family distributions, and horizontally acquired genomic islands, which we experimentally validated to be differentially present in multiple Bt isolates. By examining species-specific genomic regions, we derived molecular explanations for previously-known metabolic differences, discovered potentially new ones, and found that the acquisition of a capsular polysaccharide gene cluster in Bp, a key virulence component, is likely to have occurred non-randomly via replacement of an ancestral polysaccharide cluster. Virulence related genes, in particular members of the Type III secretion needle complex, were collectively more divergent between Bp and Bt compared to the rest of the genome, possibly contributing towards the ability of Bp to infect mammalian hosts. An analysis of pseudogenes between the two species revealed that protein inactivation events were significantly biased towards membrane-associated proteins in Bt and transcription factors in Bp. Conclusion: Our results suggest that a limited number of horizontal-acquisition events, coupled with the fine-scale functional modulation of existing proteins, are likely to be the major drivers underlying Bp virulence. The extensive genomic similarity between Bp and Bt suggests that, in some cases, Bt could be used as a possible model system for studying certain aspects of Bp behavior.Genome Institute of SingaporeSingapore. Agency for Science, Technology and Researc