Genomic Diversity within the Enterobacter cloacae Complex

Background: Isolates of the Enterobacter cloacae complex have been increasingly isolated as nosocomial pathogens, but phenotypic identification of the E. cloacae complex is unreliable and irreproducible. Identification of species based on currently available genotyping tools is already superior to phenotypic identification, but the taxonomy of isolates belonging to this complex is cumbersome. Methodolgy/Principal Findings: This study shows that multilocus sequence analysis and comparative genomic hybridization based on a mixed genome array is a powerful method for studying species assignment within the E. cloacae complex. The E. cloacae complex is shown to be evolutionarily divided into two clades that are genetically distinct from each other. The younger first clade is genetically more homogenous, contains the Enterobacter hormaechei species and is the most frequently cultured Enterobacter species in hospitals. The second and older clade consists of several (sub)species that are genetically more heterogonous. Genetic markers were identified that could discriminate between the two clades and cluster 1. Conclusions/Significance: Based on genomic differences it is concluded that some previously defined (clonal and heterogenic) (sub)species of the E. cloacae complex have to be redefined because of disagreements with known or proposed nomenclature. However, further improved identification of the redefined species will be possible based on novel markers presented here. © 2008 Paauw et al. Chemicals / CAS: Bacterial Proteins; DNA, Bacteria

Molecular Assessment of Bacterial Vaginosis by Lactobacillus Abundance and Species Diversity

Background To date, women are most often diagnosed with bacterial vaginosis (BV) using microscopy based Nugent scoring or Amsel criteria. However, the accuracy is less than optimal. The aim of the present study was to confirm the identity of known BV-associated composition profiles and evaluate indicators for BV using three molecular methods. Methods Evaluation of indicators for BV was carried out by 16S rRNA amplicon sequencing of the V5-V7 region, a tailor-made 16S rRNA oligonucleotide-based microarray, and a PCR-based profiling technique termed IS-profiling, which is based on fragment variability of the 16S-23S rRNA intergenic spacer region. An inventory of vaginal bacterial species was obtained from 40 females attending a Dutch sexually transmitted infection outpatient clinic, of which 20 diagnosed with BV (Nugent score 7–10), and 20 BV negative (Nugent score 0–3). Results Analysis of the bacterial communities by 16S rRNA amplicon sequencing revealed two clusters in the BV negative women, dominated by either Lactobacillus iners or Lactobacillus crispatus and three distinct clusters in the BV positive women. In the former, there was a virtually complete, negative correlation between L. crispatus and L. iners. BV positive subjects showed cluster profiles that were relatively high in bacterial species diversity and dominated by anaerobic species, including Gardnerella vaginalis, and those belonging to the Families of Lachnospiraceae and Leptotrichiaceae. Accordingly, the Gini-Simpson index of species diversity, and the relative abundance Lactobacillus species appeared consistent indicators for BV. Under the conditions used, only the 16S rRNA amplicon sequencing method was suitable to assess species diversity, while all three molecular composition profiling methods were able to indicate Lactobacillus abundance in the vaginal microbiota. Conclusion An affordable and simple molecular test showing a depletion of the genus Lactobacillus in combination with an increased species diversity of vaginal microbiota could serve as an alternative and practical diagnostic method for the assessment of BV

Deep Sequencing Analyses of Low Density Microbial Communities: Working at the Boundary of Accurate Microbiota Detection

Introduction: Accurate analyses of microbiota composition of low-density communities (10 3 –10 4 bacteria/sample) can be challenging. Background DNA from chemicals and consumables, extraction biases as well as differences in PCR efficiency can significantly interfere with microbiota assessment. This study was aiming to establish protocols for accurate microbiota analysis at low microbial density. Methods: To examine possible effects of bacterial density on microbiota analyses we compared microbiota profiles of seria

Unraveling the Transcriptional Dynamics of NASH Pathogenesis Affecting Atherosclerosis

The prevalence of non-alcoholic steatohepatitis (NASH) is rapidly increasing and associated with cardiovascular disease (CVD), the major cause of mortality in NASH patients. Although sharing common risk factors, the mechanisms by which NASH may directly contribute to the development to CVD remain poorly understood. The aim of this study is to gain insight into key molecular processes of NASH that drive atherosclerosis development. Thereto, a time-course study was performed in Ldlr−/−.Leiden mice fed a high-fat diet to induce NASH and atherosclerosis. The effects on NASH and atherosclerosis were assessed and transcriptome analysis was performed. Ldlr−/−.Leiden mice developed obesity, hyperlipidemia and insulin resistance, with steatosis and hepatic inflammation preceding atherosclerosis development. Transcriptome analysis revealed a time-dependent increase in pathways related to NASH and fibrosis followed by an increase in pro-atherogenic processes in the aorta. Gene regulatory network analysis identified specific liver regulators related to lipid metabolism (SC5D, LCAT and HMGCR), inflammation (IL1A) and fibrosis (PDGF, COL3A1), linked to a set of aorta target genes related to vascular inflammation (TNFA) and atherosclerosis signaling (CCL2 and FDFT1). The present study reveals pathogenic liver processes that precede atherosclerosis development and identifies hepatic key regulators driving the atherogenic pathways and regulators in the aorta

Challenges and advances in systems biology analysis of Bacillus spore physiology; molecular differences between an extreme heat resistant spore forming Bacillus subtilis food isolate and a laboratory strain

Bacterial spore formers are prime organisms of concern in the food industry. Spores from the genus Bacillus are extremely stress resistant, most notably exemplified by high thermotolerance. This sometimes allows surviving spores to germinate and grow out to vegetative cells causing food spoilage and possible intoxication. Similar issues though more pending toward spore toxigenicity are observed for the anaerobic Clostridia. The paper indicates the nature of stress resistance and highlights contemporary molecular approaches to analyze the mechanistic basis of it in Bacilli. A molecular comparison between a laboratory strain and a food borne isolate, very similar at the genomic level to the laboratory strain but generating extremely heat resistant spores, is discussed. The approaches cover genome-wide genotyping, proteomics and genome-wide expression analyses studies. The analyses aim at gathering sufficient molecular information to be able to put together an initial framework for dynamic modelling of spore germination and outgrowth behaviour. Such emerging models should be developed both at the population and at the single spore level. Tools and challenges in achieving the latter are succinctly discussed

Dynamics in Prophage Content of Invasive and Noninvasive M1 and M28 Streptococcus pyogenes Isolates in The Netherlands from 1959 to 1996▿

Invasive group A streptococcal (GAS) disease reemerged in The Netherlands in the late 1980s. To seek an explanation for this resurgence, the genetic compositions of 22 M1 and 19 M28 GAS strains isolated in The Netherlands between 1960s and the mid-1990s were analyzed by using a mixed-genome DNA microarray. During this four-decade period, M1 and especially M28 strains acquired prophages on at least eight occasions. All prophages carried a superantigen (speA2, speC, speK) or a streptodornase (sdaD2, sdn), both associated with invasive GAS disease. Invasive and noninvasive GAS strains did not differ in prophage acquisition, suggesting that there was an overall increase in the pathogenicity of M1 and M28 strains over the last four decades rather than emergence of hypervirulent subclones. The increased overall pathogenic potential may have contributed to the reemergence of invasive GAS disease in The Netherlands

Determination of the Relationship between Group A Streptococcal Genome Content, M Type, and Toxic Shock Syndrome by a Mixed Genome Microarray

Group A streptococci (GAS), or Streptococcus pyogenes, are associated with a remarkable variety of diseases, ranging from superficial infections to life-threatening diseases such as toxic-shock-like syndrome (TSS). GAS strains belonging to M types M1 and M3 are associated with TSS. This study aims to obtain insight into the gene profiles underlying different M types and disease manifestations. Genomic differences between 76 clinically well characterized GAS strains collected in The Netherlands were examined using a mixed-genome microarray. Inter-M-type genomic differences clearly outweighed intra-M-type genome variation. Phages were major contributors to observed genome diversification. We identified four novel genes, including two genes encoding fibronectin-binding-like proteins, which are highly specific to a subset of M types and thus may contribute to M-type-associated disease manifestations. All M12 strains were characterized by the unique absence of the citrate lyase complex and reduced growth under hypoxic, nutrient-deprived conditions. Furthermore, six virulence factors, including genes encoding a complement-inhibiting protein (sic), an exotoxin (speA), iron(III) binding factor, collagen binding factor (cpa), and fibrinogen binding factor (prt2-like), were unique to M1 and/or M3 strains. These virulence factors may contribute to the potential of these strains to cause TSS. Finally, in contrast to M-type-specific virulence profiles, we did not identify a common virulence profile among strains associated with TSS irrespective of their M type

A Novel Personalized Systems Nutrition Program Improves Dietary Patterns, Lifestyle Behaviors and Health-Related Outcomes: Results from the Habit Study

Personalized nutrition may be more effective in changing lifestyle behaviors compared to population-based guidelines. This single-arm exploratory study evaluated the impact of a 10-week personalized systems nutrition (PSN) program on lifestyle behavior and health outcomes. Healthy men and women (n = 82) completed the trial. Individuals were grouped into seven diet types, for which phenotypic, genotypic and behavioral data were used to generate personalized recommendations. Behavior change guidance was also provided. The intervention reduced the intake of calories (−256.2 kcal; p < 0.0001), carbohydrates (−22.1 g; p < 0.0039), sugar (−13.0 g; p < 0.0001), total fat (−17.3 g; p < 0.0001), saturated fat (−5.9 g; p = 0.0003) and PUFA (−2.5 g; p = 0.0065). Additionally, BMI (−0.6 kg/m2; p < 0.0001), body fat (−1.2%; p = 0.0192) and hip circumference (−5.8 cm; p < 0.0001) were decreased after the intervention. In the subgroup with the lowest phenotypic flexibility, a measure of the body’s ability to adapt to environmental stressors, LDL (−0.44 mmol/L; p = 0.002) and total cholesterol (−0.49 mmol/L; p < 0.0001) were reduced after the intervention. This study shows that a PSN program in a workforce improves lifestyle habits and reduces body weight, BMI and other health-related outcomes. Health improvement was most pronounced in the compromised phenotypic flexibility subgroup, which indicates that a PSN program may be effective in targeting behavior change in health-compromised target groups