Is l-arabinose important for the endophytic lifestyle of Pseudomonas spp.?

Twenty endophytic bacteria were isolated from surface-sterilized stems and roots of cucumber plants. After removal of potential siblings and human pathogens, the remaining seven strains were identified based on their 16S rDNA as Pseudomonas fluorescens (2 strains) and P. putida (5 strains). Three strains, namely P. fluorescens CS1, P. fluorescens CR2 and P. putida CR3, were able to suppress tomato foot and root rot (TFRR). Special attention was paid to the characterization of the BIOLOG carbon oxidation profiles of the isolated pseudomonads in order to identify nutrients which might be important for their endophytic lifestyle. Comparative analysis of the profiles of these seven strains with those of seven rhizospheric Pseudomonas spp. revealed that endophytes were able to oxidize l-arabinose and 2,3-butanediol significantly more often than the rhizospheric group. An independent growth experiment performed in tubes using l-arabinose and 2,3-butanediol as sole carbon sources showed the same results as seen using BIOLOG for l-arabinose, but not for 2,3-butanediol. Since l-arabinose is one of the most abundant sugars in xylem of cucumber plants and was not detected in their rhizosphere, our data suggest that utilization of l-arabinose might be a trait contributing to the endophytic lifestyle of the isolated Pseudomonas endophytes.     

Assessing the evolutionary impact of amino acid mutations in the human genome

Quantifying the distribution of fitness effects among newly arising mutations in the human genome is key to resolving important debates in medical and evolutionary genetics. Here, we present a method for inferring this distribution using Single Nucleotide Polymorphism (SNP) data from a population with non-stationary demographic history (such as that of modern humans). Application of our method to 47,576 coding SNPs found by direct resequencing of 11,404 protein coding-genes in 35 individuals (20 European Americans and 15 African Americans) allows us to assess the relative contribution of demographic and selective effects to patterning amino acid variation in the human genome. We find evidence of an ancient population expansion in the sample with African ancestry and a relatively recent bottleneck in the sample with European ancestry. After accounting for these demographic effects, we find strong evidence for great variability in the selective effects of new amino acid replacing mutations. In both populations, the patterns of variation are consistent with a leptokurtic distribution of selection coefficients (e.g., gamma or log-normal) peaked near neutrality. Specifically, we predict 27–29% of amino acid changing (nonsynonymous) mutations are neutral or nearly neutral (|s|<0.01%), 30–42% are moderately deleterious (0.01%<|s|<1%), and nearly all the remainder are highly deleterious or lethal (|s|>1%). Our results are consistent with 10–20% of amino acid differences between humans and chimpanzees having been fixed by positive selection with the remainder of differences being neutral or nearly neutral. Our analysis also predicts that many of the alleles identified via whole-genome association mapping may be selectively neutral or (formerly) positively selected, implying that deleterious genetic variation affecting disease phenotype may be missed by this widely used approach for mapping genes underlying complex traits.     

Medical sequencing at the extremes of human body mass

Body weight is a quantitative trait with significant heritability in humans. To identify potential genetic contributors to this phenotype, we resequenced the coding exons and splice junctions of 58 genes in 379 obese and 378 lean individuals. Our 96-Mb survey included 21 genes associated with monogenic forms of obesity in humans or mice, as well as 37 genes that function in body weight-related pathways. We found that the monogenic obesity-associated gene group was enriched for rare nonsynonymous variants unique to the obese population compared with the lean population. In addition, computational analysis predicted a greater fraction of deleterious variants within the obese cohort. Together, these data suggest that multiple rare alleles contribute to obesity in the population and provide a medical sequencing-based approach to detect them.     

Regional trends and drivers of the global methane budget

The ongoing development of the Global Carbon Project (GCP) global methane (CH₄) budget shows a continuation of increasing CH₄ emissions and CH₄ accumulation in the atmosphere during 2000–2017. Here, we decompose the global budget into 19 regions (18 land and 1 oceanic) and five key source sectors to spatially attribute the observed global trends. A comparison of top-down (TD) (atmospheric and transport model-based) and bottom-up (BU) (inventory- and process model-based) CH₄ emission estimates demonstrates robust temporal trends with CH₄ emissions increasing in 16 of the 19 regions. Five regions—China, Southeast Asia, USA, South Asia, and Brazil—account for >40% of the global total emissions (their anthropogenic and natural sources together totaling >270 Tg CH₄ yr^?1 in 2008–2017). Two of these regions, China and South Asia, emit predominantly anthropogenic emissions (>75%) and together emit more than 25% of global anthropogenic emissions. China and the Middle East show the largest increases in total emission rates over the 2000 to 2017 period with regional emissions increasing by >20%. In contrast, Europe and Korea and Japan show a steady decline in CH₄ emission rates, with total emissions decreasing by ~10% between 2000 and 2017. Coal mining, waste (predominantly solid waste disposal) and livestock (especially enteric fermentation) are dominant drivers of observed emissions increases while declines appear driven by a combination of waste and fossil emission reductions. As such, together these sectors present the greatest risks of further increasing the atmospheric CH₄ burden and the greatest opportunities for greenhouse gas abatement.     

SNP2RFLP: a computational tool to facilitate genetic mapping using benchtop analysis of SNPs

Genome-wide analysis of single nucleotide polymorphism (SNP) markers is an extremely efficient means for genetic mapping of mutations or traits in mice. However, this approach often defines a relatively large recombinant interval. To facilitate the refinement of this interval, we developed the program SNP2RFLP. This program can be used to identify region-specific SNPs in which the polymorphic nucleotide creates a restriction fragment length polymorphism (RFLP) that can be readily assayed at the benchtop using restriction enzyme digestion of SNP-containing PCR products. The program permits user-defined queries that maximize the informative markers for a particular application. This facilitates fine-mapping in a region containing a mutation of interest, which should prove valuable to the mouse genetics community. SNP2RFLP and further details are publicly available at http://genetics.bwh.harvard.edu/snp2rflp/.

     

A cognitive behavioural intervention to reduce sexually transmitted infections among gay men: randomised trial

ObjectiveTo determine the effectiveness of a brief cognitive behavioural intervention in reducing the incidence of sexually transmitted infections among gay men.DesignRandomised controlled trial with 12 months'' follow up.SettingSexual health clinic in London.Participants343 gay men with an acute sexually transmitted infection or who reported having had unprotected anal intercourse in the past year.Results72% (361/499) of men invited to enter the study did so. 90% (308/343) of participants returned at least one follow up questionnaire or re-attended the clinic and requested a check up for sexually transmitted infections during follow up. At baseline, 37% (63/172) of the intervention group and 30% (50/166) of the control group reported having had unprotected anal intercourse in the past month. At 12 months, the proportions were 27% (31/114) and 32% ( 39/124) respectively (P=0.56). However, 31% (38/123) of the intervention group and 21% (35/168) of controls had had at least one new infection diagnosed at the clinic (adjusted odds ratio 1.66, 95% confidence interval 1.00 to 2.74). Considering only men who requested a check up for sexually transmitted infections, the proportion diagnosed with a new infection was 58% (53/91) for men in the intervention group and 43% (35/81) for men in the control group (adjusted odds ratio 1.84, 0.99 to 3.40). Using a regional database that includes information from 23 sexual health clinics in London, we determined that few participants had attended other sexual health clinics.ConclusionsThis behavioural intervention was acceptable and feasible to deliver, but it did not reduce the risk of acquiring a new sexually transmitted infection among these gay men at high risk. Even carefully designed interventions should not be assumed to bring benefit. It is important to evaluate their effects in randomised trials with objective clinical end points.

What is already known on this topic

The need for effective HIV prevention strategies based on reducing sexual risk behaviour remains importantFew interventions to reduce sexual risk behaviour have been rigorously evaluated using randomised controlled trials

What this study adds

This is the first randomised controlled trial of an intervention addressing sexual behaviour in homosexual men that uses sexually transmitted infections and self reported behaviour as end pointsThe intervention was brief and feasible to use in a busy clinic, but it did not reduce the risk of participants acquiring new infectionsThe potential for behavioural interventions to do more harm than good needs to be taken seriously     

Construction of a fluorescent biosensor family

Bacterial periplasmic binding proteins (bPBPs) are specific for a wide variety of small molecule ligands. bPBPs undergo a large, ligand-mediated conformational change that can be linked to reporter functions to monitor ligand concentrations. This mechanism provides the basis of a general system for engineering families of reagentless biosensors that share a common physical signal transduction functionality and detect many different analytes. We demonstrate the facility of designing optical biosensors based on fluorophore conjugates using 8 environmentally sensitive fluorophores and 11 bPBPs specific for diverse ligands, including sugars, amino acids, anions, cations, and dipeptides. Construction of reagentless fluorescent biosensors relies on identification of sites that undergo a local conformational change in concert with the global, ligand-mediated hinge-bending motion. Construction of cysteine mutations at these locations then permits site-specific coupling of environmentally sensitive fluorophores that report ligand binding as changes in fluorescence intensity. For 10 of the bPBPs presented in this study, the three-dimensional receptor structure was used to predict the location of reporter sites. In one case, a bPBP sensor specific for glutamic and aspartic acid was designed starting from genome sequence information and illustrates the potential for discovering novel binding functions in the microbial genosphere using bioinformatics.     

High incidence of plant growth-stimulating bacteria associated with the rhizosphere of wheat grown on salinated soil in Uzbekistan

Soil salinization is increasing steadily in many parts of the world and causes major problems for plant productivity. Under these stress conditions, root-associated beneficial bacteria can help improve plant growth and nutrition. In this study, salt-tolerant bacteria from the rhizosphere of Uzbek wheat with potentially beneficial traits were isolated and characterized. Eight strains which initially positively affect the growth of wheat plants in vitro were investigated in detail. All eight strains are salt tolerant and have some of the following plant growth-beneficial properties: production of auxin, HCN, lipase or protease and wheat growth promotion. Using sequencing of part of the 16S rDNA, the eight new isolates were identified as Acinetobacter (two strains), Pseudomonas aeruginosa , Staphylococcus saprophyticus , Bacillus cereus , Enterobacter hormaechei , Pantoae agglomerans and Alcaligenes faecalis . All these strains are potential human pathogens. Possible reasons for why these bacteria present in the rhizosphere and establish there are discussed.     

Most rare missense alleles are deleterious in humans: implications for complex disease and association studies

The accumulation of mildly deleterious missense mutations in individual human genomes has been proposed to be a genetic basis for complex diseases. The plausibility of this hypothesis depends on quantitative estimates of the prevalence of mildly deleterious de novo mutations and polymorphic variants in humans and on the intensity of selective pressure against them. We combined analysis of mutations causing human Mendelian diseases, of human-chimpanzee divergence, and of systematic data on human genetic variation and found that ~20% of new missense mutations in humans result in a loss of function, whereas ~27% are effectively neutral. Thus, the remaining 53% of new missense mutations have mildly deleterious effects. These mutations give rise to many low-frequency deleterious allelic variants in the human population, as is evident from a new data set of 37 genes sequenced in >1,500 individual human chromosomes. Surprisingly, up to 70% of low-frequency missense alleles are mildly deleterious and are associated with a heterozygous fitness loss in the range 0.001-0.003. Thus, the low allele frequency of an amino acid variant can, by itself, serve as a predictor of its functional significance. Several recent studies have reported a significant excess of rare missense variants in candidate genes or pathways in individuals with extreme values of quantitative phenotypes. These studies would be unlikely to yield results if most rare variants were neutral or if rare variants were not a significant contributor to the genetic component of phenotypic inheritance. Our results provide a justification for these types of candidate-gene (pathway) association studies and imply that mutation-selection balance may be a feasible evolutionary mechanism underlying some common diseases.     

Common single-nucleotide polymorphisms act in concert to affect plasma levels of high-density lipoprotein cholesterol

The identification of DNA sequence variants underlying human complex phenotypes remains a significant challenge for several reasons: individual variants can have small phenotypic effects or low population frequencies, and multiple allelic variants may act in concert to affect a trait. We evaluated the combined effect of allelic variants in seven genes involved in high-density lipoprotein (HDL) metabolism, using forward stepwise regression. Analysis of all known common single-nucleotide polymorphisms (SNPs) in the seven candidate genes revealed four variants that were associated with incremental changes in HDL cholesterol levels in three independent samples. Conversely, analysis of 660 polymorphisms in eight genes that do not appear to be involved in HDL metabolism did not identify any associations with plasma HDL-cholesterol levels. These data indicate that several common SNPs act in concert to influence plasma levels of HDL cholesterol.