Multi-Locus Sequence Analysis Reveals Profound Genetic Diversity among Isolates of the Human Pathogen Bartonella bacilliformis

Bartonella bacilliformis is the aetiological agent of human bartonellosis, a potentially life threatening infection of significant public health concern in the Andean region of South America. Human bartonellosis has long been recognised in the region but a recent upsurge in the number of cases of the disease and an apparent expansion of its geographical distribution have re-emphasized its contemporary medical importance. Here, we describe the development of a multi-locus sequence typing (MLST) scheme for B. bacilliformis and its application to an archive of 43 isolates collected from patients across Peru. MLST identified eight sequence types among these isolates and the delineation of these was generally congruent with those of the previously described typing scheme. Phylogenetic analysis based on concatenated sequence data derived from MLST loci revealed that seven of the eight sequence types were closely related to one another; however, one sequence type, ST8, exhibited profound evolutionary divergence from the others. The extent of this divergence was akin to that observed between other members of the Bartonella genus, suggesting that ST8 strains may be better considered as members of a novel Bartonella genospecies

Negative Regulators of Insulin Signaling Revealed in a Genome-Wide Functional Screen

Type 2 diabetes develops due to a combination of insulin resistance and β-cell failure and current therapeutics aim at both of these underlying causes. Several negative regulators of insulin signaling are known and are the subject of drug discovery efforts. We sought to identify novel contributors to insulin resistance and hence potentially novel targets for therapeutic intervention.An arrayed cDNA library encoding 18,441 human transcripts was screened for inhibitors of insulin signaling and revealed known inhibitors and numerous potential novel regulators. The novel hits included proteins of various functional classes such as kinases, phosphatases, transcription factors, and GTPase associated proteins. A series of secondary assays confirmed the relevance of the primary screen hits to insulin signaling and provided further insight into their modes of action.Among the novel hits was PALD (KIAA1274, paladin), a previously uncharacterized protein that when overexpressed led to inhibition of insulin's ability to down regulate a FOXO1A-driven reporter gene, reduced upstream insulin-stimulated AKT phosphorylation, and decreased insulin receptor (IR) abundance. Conversely, knockdown of PALD gene expression resulted in increased IR abundance, enhanced insulin-stimulated AKT phosphorylation, and an improvement in insulin's ability to suppress FOXO1A-driven reporter gene activity. The present data demonstrate that the application of arrayed genome-wide screening technologies to insulin signaling is fruitful and is likely to reveal novel drug targets for insulin resistance and the metabolic syndrome

Parallel Evolution of a Type IV Secretion System in Radiating Lineages of the Host-Restricted Bacterial Pathogen Bartonella

Adaptive radiation is the rapid origination of multiple species from a single ancestor as the result of concurrent adaptation to disparate environments. This fundamental evolutionary process is considered to be responsible for the genesis of a great portion of the diversity of life. Bacteria have evolved enormous biological diversity by exploiting an exceptional range of environments, yet diversification of bacteria via adaptive radiation has been documented in a few cases only and the underlying molecular mechanisms are largely unknown. Here we show a compelling example of adaptive radiation in pathogenic bacteria and reveal their genetic basis. Our evolutionary genomic analyses of the α-proteobacterial genus Bartonella uncover two parallel adaptive radiations within these host-restricted mammalian pathogens. We identify a horizontally-acquired protein secretion system, which has evolved to target specific bacterial effector proteins into host cells as the evolutionary key innovation triggering these parallel adaptive radiations. We show that the functional versatility and adaptive potential of the VirB type IV secretion system (T4SS), and thereby translocated Bartonella effector proteins (Beps), evolved in parallel in the two lineages prior to their radiations. Independent chromosomal fixation of the virB operon and consecutive rounds of lineage-specific bep gene duplications followed by their functional diversification characterize these parallel evolutionary trajectories. Whereas most Beps maintained their ancestral domain constitution, strikingly, a novel type of effector protein emerged convergently in both lineages. This resulted in similar arrays of host cell-targeted effector proteins in the two lineages of Bartonella as the basis of their independent radiation. The parallel molecular evolution of the VirB/Bep system displays a striking example of a key innovation involved in independent adaptive processes and the emergence of bacterial pathogens. Furthermore, our study highlights the remarkable evolvability of T4SSs and their effector proteins, explaining their broad application in bacterial interactions with the environment

Application of Two-Part Statistics for Comparison of Sequence Variant Counts

Investigation of microbial communities, particularly human associated communities, is significantly enhanced by the vast amounts of sequence data produced by high throughput sequencing technologies. However, these data create high-dimensional complex data sets that consist of a large proportion of zeros, non-negative skewed counts, and frequently, limited number of samples. These features distinguish sequence data from other forms of high-dimensional data, and are not adequately addressed by statistical approaches in common use. Ultimately, medical studies may identify targeted interventions or treatments, but lack of analytic tools for feature selection and identification of taxa responsible for differences between groups, is hindering advancement. The objective of this paper is to examine the application of a two-part statistic to identify taxa that differ between two groups. The advantages of the two-part statistic over common statistical tests applied to sequence count datasets are discussed. Results from the t-test, the Wilcoxon test, and the two-part test are compared using sequence counts from microbial ecology studies in cystic fibrosis and from cenote samples. We show superior performance of the two-part statistic for analysis of sequence data. The improved performance in microbial ecology studies was independent of study type and sequence technology used

Number of addictive substances used related to increased risk of unnatural death: A combined medico-legal and case-record study

<p>Abstract</p> <p>Background</p> <p>Substance use disorders have repeatedly been found to lead to premature death, i.e. drug-related death by disease, fatal intoxications, or trauma (accidents, suicide, undetermined suicide, and homicide). The present study examined the relationship between multi-drug substance use and natural and unnatural death.</p> <p>Methods</p> <p>All consecutive, autopsied patients who had been in contact with the Addiction Centre in Malmö University Hospital from 1993 to 1997 inclusive were investigated. Drug abuse was investigated blindly in the case records and related to the cause of death in 387 subjects.</p> <p>Results</p> <p>Every substance apart from alcohol used previously in life added to the risk of unnatural death in a linear way. There were independent increased risks of fatal heroin overdoses or undetermined suicide. Death by suicide and violent death were unrelated to additional abuse.</p> <p>Conclusion</p> <p>The number of drugs used was related to an increased risk of unnatural death by undetermined suicide (mainly fatal intoxications) and heroin overdose.</p

Sick leave among home-care personnel: a longitudinal study of risk factors

BACKGROUND: Sick leave due to neck, shoulder and back disorders (NSBD) is higher among health-care workers, especially nursing aides/assistant nurses, compared with employees in other occupations. More information is needed about predictors of sick leave among health care workers. The aim of the study was to assess whether self-reported factors related to health, work and leisure time could predict: 1) future certified sick leave due to any cause, in nursing aides/assistant nurses (Study group I) and 2) future self-reported sick leave due to NSBD in nursing aides/assistant nurses (Study group II). METHODS: Study group I, comprised 443 female nursing aides/assistant nurses, not on sick leave at baseline when a questionnaire was completed. Data on certified sick leave were collected after 18 months. Study group II comprised 274 of the women, who at baseline reported no sick leave during the preceding year due to NSBD and who participated at the 18 month follow-up. Data on sick leave due to NSBD were collected from the questionnaire at 18 months. The associations between future sick leave and factors related to health, work and leisure time were tested by logistic regression analyses. RESULTS: Health-related factors such as previous low back disorders (OR: 1.89; 95% CI 1.20–2.97) and previous sick leave (OR 6.40; 95%CI 3.97–10.31), were associated with a higher risk of future sick leave due to any cause. Factors related to health, work and leisure time, i.e. previous low back disorders (OR: 4.45; 95% CI 1.27–15.77) previous sick leave, not due to NSBD (OR 3.30; 95%CI 1.33–8.17), high strain work (OR 2.34; 95%CI 1.05–5.23) and high perceived physical exertion in domestic work (OR 2.56; 95%CI 1.12–5.86) were associated with a higher risk of future sick leave due to NSBD. In the final analyses, previous low back disorders and previous sick leave remained significant in both study groups. CONCLUSION: The results suggest a focus on previous low back disorders and previous sick leave for the design of early prevention programmes aiming at reducing future sick leave due to any cause, as well as due to NSBD, among nursing aides/assistant nurses. A multifactorial approach may be of importance in the early prevention of sick leave due to NSBD

Saturated Fatty Acids and Risk of Coronary Heart Disease: Modulation by Replacement Nutrients

Despite the well-established observation that substitution of saturated fats for carbohydrates or unsaturated fats increases low-density lipoprotein (LDL) cholesterol in humans and animal models, the relationship of saturated fat intake to risk for atherosclerotic cardiovascular disease in humans remains controversial. A critical question is what macronutrient should be used to replace saturated fat. Substituting polyunsaturated fat for saturated fat reduces LDL cholesterol and the total cholesterol to high-density lipoprotein cholesterol ratio. However, replacement of saturated fat by carbohydrates, particularly refined carbohydrates and added sugars, increases levels of triglyceride and small LDL particles and reduces high-density lipoprotein cholesterol, effects that are of particular concern in the context of the increased prevalence of obesity and insulin resistance. Epidemiologic studies and randomized clinical trials have provided consistent evidence that replacing saturated fat with polyunsaturated fat, but not carbohydrates, is beneficial for coronary heart disease. Therefore, dietary recommendations should emphasize substitution of polyunsaturated fat and minimally processed grains for saturated fat

Evidence of Transfer by Conjugation of Type IV Secretion System Genes between Bartonella Species and Rhizobium radiobacter in Amoeba

Background: Bartonella species cospeciate with mammals and live within erythrocytes. Even in these specific niches, it has been recently suggested by bioinformatic analysis of full genome sequences that Lateral Gene Transfer (LGT) may occur but this has never been demonstrated biologically. Here we describe the sequence of the B. rattaustraliani (AUST/NH4 T) circular plasmid (pNH4) that encodes the tra cluster of the Type IV secretion system (T4SS) and we eventually provide evidence that Bartonella species may conjugate and exchange this plasmid inside amoeba. Principal Findings: The T4SS of pNH4 is critical for intracellular viability of bacterial pathogens, exhibits bioinformatic evidence of LGT among bacteria living in phagocytic protists. For instance, 3 out of 4 T4SS encoding genes from pNH4 appear to be closely related to Rhizobiales, suggesting that gene exchange occurs between intracellular bacteria from mammals (bartonellae) and plants (Rhizobiales). We show that B. rattaustraliani and Rhizobium radiobacter both survived within the amoeba Acanthamoeba polyphaga and can conjugate together. Our findings further support the hypothesis that tra genes might also move into and out of bacterial communities by conjugation, which might be the primary means of genomic evolution for intracellular adaptation by cross-talk of interchangeable genes between Bartonella species and plant pathogens. Conclusions: Based on this, we speculate that amoeba favor the transfer of genes as phagocytic protists, which allows fo