Towards a Wolbachia Multilocus Sequence Typing system : discrimination of Wolbachia strains present in Drosophila species

Author Posting. © The Author(s), 2006. This is the author's version of the work. It is posted here by permission of Springer for personal use, not for redistribution. The definitive version was published in Current Microbiology 53 (2006): 388-395, doi:10.1007/s00284-006-0054-1.Among the diverse maternally inherited symbionts in arthropods, Wolbachia are the most common and infect over 20% of all species. In a departure from traditional genotyping or phylogenetic methods relying on single Wolbachia genes, the present study represents an initial Multilocus Sequence Typing (MLST) analysis to discriminate closely related Wolbachia pipientis strains, and additional data on sequence diversity in Wolbachia. We report new phylogenetic characterization of four genes (aspC, atpD, sucB and pdhB), and provide an expanded analysis of markers described in previous studies (16S rDNA, ftsZ, groEL, dnaA and gltA). MLST analysis of the bacterial strains present in sixteen different Drosophila-Wolbachia associations detected four distinct clonal complexes that also corresponded to maximum-likelihood identified phylogenetic clades. Among the sixteen associations analyzed, six could not be assigned to MLST clonal complexes and were also shown to be in conflict with relationships predicted by maximum-likelihood phylogenetic inferences. The results demonstrate the discriminatory power of MLST for identifying strains and clonal lineages of Wolbachia and provide a robust foundation for studying the ecology and evolution of this widespread endosymbiont.This work was partially supported by intramural funds of the University of Ioannina to K. Bourtzis, by grants to J.J. Wernegreen from the National Institutes of Health (R01 GM62626-01) and the NASA Astrobiology Institute (NNA04CC04A), and to J.H. Werren and J.J. Wernegreen from the National Science Foundation (EF-0328363)

A Rickettsiella Bacterium from the Hard Tick, Ixodes woodi: Molecular Taxonomy Combining Multilocus Sequence Typing (MLST) with Significance Testing

Hard ticks (Acari: Ixodidae) are known to harbour intracellular bacteria from several phylogenetic groups that can develop both mutualistic and pathogenic relationships to the host. This is of particular importance for public health as tick derived bacteria can potentially be transmitted to mammals, including humans, where e.g. Rickettsia or Coxiella act as severe pathogens. Exact molecular taxonomic identification of tick associated prokaryotes is a necessary prerequisite of the investigation of their relationship to both the tick and possible vertebrate hosts. Previously, an intracellular bacterium had been isolated from a monosexual, parthenogenetically reproducing laboratory colony of females of the hard tick, Ixodes woodi Bishopp, and had preliminarily been characterized as a “Rickettsiella-related bacterium”. In the present molecular taxonomic study that is based on phylogenetic reconstruction from both 16 S ribosomal RNA and protein-encoding marker sequences complemented with likelihood-based significance testing, the bacterium from I. woodi has been identified as a strain of the taxonomic species Rickettsiella grylli. It is the first time that a multilocus sequence typing (MLST) approach based on a four genes comprising MLST scheme has been implemented in order to classify a Rickettsiella-like bacterium to this species. The study demonstrated that MLST holds potential for a better resolution of phylogenetic relationships within the genus Rickettsiella, but requires sequence determination from further Rickettsiella-like bacteria in order to complete the current still fragmentary picture of Rickettsiella systematics

Known and unknown requirements in healthcare

We report experience in requirements elicitation of domain knowledge from experts in clinical and cognitive neurosciences. The elicitation target was a causal model for early signs of dementia indicated by changes in user behaviour and errors apparent in logs of computer activity. A Delphi-style process consisting of workshops with experts followed by a questionnaire was adopted. The paper describes how the elicitation process had to be adapted to deal with problems encountered in terminology and limited consensus among the experts. In spite of the difficulties encountered, a partial causal model of user behavioural pathologies and errors was elicited. This informed requirements for configuring data- and text-mining tools to search for the specific data patterns. Lessons learned for elicitation from experts are presented, and the implications for requirements are discussed as “unknown unknowns”, as well as configuration requirements for directing data-/text-mining tools towards refining awareness requirements in healthcare applications

Gene diversity in grevillea populations introduced in Brazil and its implication on management of genetic resources.

A variabilidade isoenzimática para seis populações de Grevillea robusta, oriundas de um teste de procedências/progenies, implantado no delineamento em blocos casualizados com 5 plantas por parcela, no Sul do Brasil, é descrita. A estrutura genética da população foi analisada utilizando-se marcadores bioquímicos, aos 5 anos de idade, especificamente para os locos MDH-3, PGM-2, DIA-2, PO-1, PO-2, SOD-1, e SKDH-1. As procedências do norte de ocorrência natural (Rathdowney e Woodenbong) apresentaram divergência genética superior, em relação à média das progênies, considerando o número de alelos por locus, (Ap), a riqueza alélica (Rs), a diversidade genética de Nei (H), e o coeficiente de endogamia (f). A endogamia foi detectada em diversos graus. A testemunha comercial apresentou o maior coeficiente de endogamia, (f = 0,4448), comparativamente à média das procedências (f = 0,2306), possivelmente devido à insuficiente amostragem populacional na região de origem (Austrália). Apesar de sua ocorrência natural restrita, observou-se correlação positiva entre divergência genética e distância geográfica entre as populações originais. A distância genética e análise de cluster, baseada no modelo bayesiano, mostrou três grupos de procedências distintos: 1) Rathdowney- QLD e Woodenbong-QLD; 2) Paddy?s Flat-NSW; e 3) Mann River-NSW, Boyd River-NSW e a testemunha comercial (material utilizado no Brasil). O agrupamento da testemunha com as procedências Mann River-NSW e Boyd River-NSW sugere um maior potencial das procedências do norte para o melhoramento genético visando à produção de madeira no Brasil, devido a sua elevada diversidade genética e baixo coeficiente de endogamia

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