ShoRAH: estimating the genetic diversity of a mixed sample from next-generation sequencing data

<p>Abstract</p> <p>Background</p> <p>With next-generation sequencing technologies, experiments that were considered prohibitive only a few years ago are now possible. However, while these technologies have the ability to produce enormous volumes of data, the sequence reads are prone to error. This poses fundamental hurdles when genetic diversity is investigated.</p> <p>Results</p> <p>We developed ShoRAH, a computational method for quantifying genetic diversity in a mixed sample and for identifying the individual clones in the population, while accounting for sequencing errors. The software was run on simulated data and on real data obtained in wet lab experiments to assess its reliability.</p> <p>Conclusions</p> <p>ShoRAH is implemented in C++, Python, and Perl and has been tested under Linux and Mac OS X. Source code is available under the GNU General Public License at <url>http://www.cbg.ethz.ch/software/shorah</url>.</p

Cocrystal structure of a class-I preQ1 riboswitch reveals a pseudoknot recognizing an essential hypermodified nucleobase

Riboswitches are mRNA domains that bind metabolites and modulate gene expression in cis. We report cocrystal structures of a remarkably compact riboswitch (34 nucleotides suffice for ligand recognition) from Bacillus subtilis selective for the essential nucleobase preQ1 (7-aminomethyl-7-deazaguanine). These reveal a previously unrecognized pseudoknot fold, and suggest a conserved gene-regulatory mechanism whereby ligand binding promotes sequestration of an RNA segment that otherwise assembles into a transcriptional anti-terminator

Dynamics of mitochondrial heteroplasmy in three families investigated via a repeatable re-sequencing study

Background: Originally believed to be a rare phenomenon, heteroplasmy - the presence of more than one mitochondrial DNA (mtDNA) variant within a cell, tissue, or individual - is emerging as an important component of eukaryotic genetic diversity. Heteroplasmies can be used as genetic markers in applications ranging from forensics to cancer diagnostics. Yet the frequency of heteroplasmic alleles may vary from generation to generation due to the bottleneck occurring during oogenesis. Therefore, to understand the alterations in allele frequencies at heteroplasmic sites, it is of critical importance to investigate the dynamics of maternal mtDNA transmission. Results: Here we sequenced, at high coverage, mtDNA from blood and buccal tissues of nine individuals from three families with a total of six maternal transmission events. Using simulations and re-sequencing of clonal DNA, we devised a set of criteria for detecting polymorphic sites in heterogeneous genetic samples that is resistant to the noise originating from massively parallel sequencing technologies. Application of these criteria to nine human mtDNA samples revealed four heteroplasmic sites. Conclusions: Our results suggest that the incidence of heteroplasmy may be lower than estimated in some other recent re-sequencing studies, and that mtDNA allelic frequencies differ significantly both between tissues of the same individual and between a mother and her offspring. We designed our study in such a way that the complete analysis described here can be repeated by anyone either at our site or directly on the Amazon Cloud. Our computational pipeline can be easily modified to accommodate other applications, such as viral re-sequencing

Design principles for riboswitch function

Scientific and technological advances that enable the tuning of integrated regulatory components to match network and system requirements are critical to reliably control the function of biological systems. RNA provides a promising building block for the construction of tunable regulatory components based on its rich regulatory capacity and our current understanding of the sequence–function relationship. One prominent example of RNA-based regulatory components is riboswitches, genetic elements that mediate ligand control of gene expression through diverse regulatory mechanisms. While characterization of natural and synthetic riboswitches has revealed that riboswitch function can be modulated through sequence alteration, no quantitative frameworks exist to investigate or guide riboswitch tuning. Here, we combined mathematical modeling and experimental approaches to investigate the relationship between riboswitch function and performance. Model results demonstrated that the competition between reversible and irreversible rate constants dictates performance for different regulatory mechanisms. We also found that practical system restrictions, such as an upper limit on ligand concentration, can significantly alter the requirements for riboswitch performance, necessitating alternative tuning strategies. Previous experimental data for natural and synthetic riboswitches as well as experiments conducted in this work support model predictions. From our results, we developed a set of general design principles for synthetic riboswitches. Our results also provide a foundation from which to investigate how natural riboswitches are tuned to meet systems-level regulatory demands

Feedback as intervention for team learning in virtual teams: the role of team cohesion and personality

Scholars and practitioners agree that virtual teams (VTs) have become commonplace in today's digital workplace. Relevant literature argues that learning constitutes a significant contributor to team member satisfaction and performance, and that, at least in face-to-face teams, team cohesion fosters team learning. Given the additional challenges VTs face, e.g. geographical dispersion, which are likely have a negative influence on cohesion, in this paper we shed light on the relationship between team cohesion and team learning. We adopted a quantitative approach and studied 54 VTs in our quest to understand the role of feedback in mediating this relationship and, more specifically, the role of personality traits in moderating the indirect effect of team feedback and guided reflection intervention on TL through team cohesion within the VT context. Our findings highlight the importance of considering aspects related to the team composition when devising intervention strategies for VTs, and provide empirical support for an interactionist model between personality and emergent states such as cohesion. Implications for theory and practice are also discussed

Genetic Basis of Growth Adaptation of Escherichia coli after Deletion of pgi, a Major Metabolic Gene

Bacterial survival requires adaptation to different environmental perturbations such as exposure to antibiotics, changes in temperature or oxygen levels, DNA damage, and alternative nutrient sources. During adaptation, bacteria often develop beneficial mutations that confer increased fitness in the new environment. Adaptation to the loss of a major non-essential gene product that cripples growth, however, has not been studied at the whole-genome level. We investigated the ability of Escherichia coli K-12 MG1655 to overcome the loss of phosphoglucose isomerase (pgi) by adaptively evolving ten replicates of E. coli lacking pgi for 50 days in glucose M9 minimal medium and by characterizing endpoint clones through whole-genome re-sequencing and phenotype profiling. We found that 1) the growth rates for all ten endpoint clones increased approximately 3-fold over the 50-day period; 2) two to five mutations arose during adaptation, most frequently in the NADH/NADPH transhydrogenases udhA and pntAB and in the stress-associated sigma factor rpoS; and 3) despite similar growth rates, at least three distinct endpoint phenotypes developed as defined by different rates of acetate and formate secretion. These results demonstrate that E. coli can adapt to the loss of a major metabolic gene product with only a handful of mutations and that adaptation can result in multiple, alternative phenotypes

Personality, personnel selection, and job performance

Job Performance: The term job performance can either refer to the objective or subjective outcomes one achieves in a specific job (e.g., the profit of a sales persons, the number of publications of a scientist, the number of successful operations of a surgeon) or to work-related activities (e.g., writing an article, conducting specific surgical acts). In the majority of research on this topic, job performance as an outcome is used. Personnel selection: Personnel selection refers to the process of selecting the best employees for specific jobs. Introduction One major application of personality research is in the area of personnel selection. The key question in this area is to which extent personality can predict how well a candidate will perform on the job he or she is applying for. Most scholars in this area acknowledge that personality has predictive validity for job performance. In line with this, personality assessment is part of the selection procedure in many organizations

Decrease of virulence for BALB/c mice produced by continuous subculturing of Nocardia brasiliensis

<p>Abstract</p> <p>Background</p> <p>Subculturing has been extensively used to attenuate human pathogens. In this work we studied the effect of continuous subculturing of <it>Nocardia brasiliensis </it>HUJEG-1 on virulence in a murine model.</p> <p>Methods</p> <p><it>Nocardia brasiliensis </it>HUJEG-1 was subcultured up to 130 times on brain heart infusion over four years. BALB/c mice were inoculated in the right foot pad with the bacteria subcultured 0, 40, 80, 100 and 130 times (T₀, T₄₀, T₈₀T₁₀₀and T₁₃₀). The induction of resistance was tested by using T₁₃₀to inoculate a group of mice followed by challenge with T0 12 weeks later. Biopsies were taken from the newly infected foot-pad and immunostained with antibodies against CD4, CD8 and CD14 in order to analyze the in situ immunological changes.</p> <p>Results</p> <p>When using T₄₀, T₈₀T₁₀₀and T₁₃₀as inoculums we observed lesions in 10, 5, 0 and 0 percent of the animals, respectively, at the end of 12 weeks. In contrast, their controls produced mycetoma in 80, 80, 70 and 60% of the inoculated animals. When studying the protection of T₁₃₀, we observed a partial resistance to the infection. Immunostaining revealed an intense CD4+ lymphocytic and macrophage infiltrate in healing lesions.</p> <p>Conclusions</p> <p>After 130 in vitro passages of <it>N. brasiliensis </it>HUJEG-1 a severe decrease in its virulence was observed. Immunization of BALB/c mice, with these attenuated cells, produced a state of partial resistance to infection with the non-subcultured isolate.</p

To be or not to be a pseudogene: a molecular epidemiological approach to the mclx genes and its impact in tuberculosis

Tuberculosis presents a myriad of symptoms, progression routes and propagation patterns not yet fully understood. Whereas for a long time research has focused solely on the patient immunity and overall susceptibility, it is nowadays widely accepted that the genetic diversity of its causative agent, Mycobacterium tuberculosis, plays a key role in this dynamic. This study focuses on a particular family of genes, the mclxs (Mycobacterium cyclase/LuxR-like genes), which codify for a particular and nearly mycobacterial-exclusive combination of protein domains. mclxs genes were found to be pseudogenized by frameshift-causing insertion(s)/deletion(s) in a considerable number of M. tuberculosis complex strains and clinical isolates. To discern the functional implications of the pseudogenization, we have analysed the pattern of frameshift-causing mutations in a group of M. tuberculosis isolates while taking into account their microbial-, patient- and disease-related traits. Our logistic regression-based analyses have revealed disparate effects associated with the transcriptional inactivation of two mclx genes. In fact, mclx2 (Rv1358) pseudogenization appears to be primarily driven by the microbial phylogenetic background, being mainly related to the Euro-American (EAm) lineage; on the other hand, mclx3 (Rv2488c) presents a higher tendency for pseudogenization among isolates from patients born on the Western Pacific area, and from isolates causing extra-pulmonary infections. These results contribute to the overall knowledge on the biology of M. tuberculosis infection, whereas at the same time launch the necessary basis for the functional assessment of these so far overlooked genes.This work was supported by Fundacao para a Ciencia e Tecnologia (FCT), Portugal, and cofunded by Programa Operacional Regional do Norte (ON.2-O Novo Norte), Quadro de Referencia Estrategico Nacional (QREN), through the Fundo Europeu de Desenvolvimento Regional (FEDER), and from Projeto Estrategico - LA 26 - 2013-2014 (PEst-C/SAU/LA0026/2013). H.N.-G. received a personal FCT Grant (SFRH/BD/33902/2209). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Rates of Mutation and Host Transmission for an Escherichia coli Clone over 3 Years

Although over 50 complete Escherichia coli/Shigella genome sequences are available, it is only for closely related strains, for example the O55:H7 and O157:H7 clones of E. coli, that we can assign differences to individual evolutionary events along specific lineages. Here we sequence the genomes of 14 isolates of a uropathogenic E. coli clone that persisted for 3 years within a household, including a dog, causing a urinary tract infection (UTI) in the dog after 2 years. The 20 mutations observed fit a single tree that allows us to estimate the mutation rate to be about 1.1 per genome per year, with minimal evidence for adaptive change, including in relation to the UTI episode. The host data also imply at least 6 host transfer events over the 3 years, with 2 lineages present over much of that period. To our knowledge, these are the first direct measurements for a clone in a well-defined host community that includes rates of mutation and host transmission. There is a concentration of non-synonymous mutations associated with 2 transfers to the dog, suggesting some selection pressure from the change of host. However, there are no changes to which we can attribute the UTI event in the dog, which suggests that this occurrence after 2 years of the clone being in the household may have been due to chance, or some unknown change in the host or environment. The ability of a UTI strain to persist for 2 years and also to transfer readily within a household has implications for epidemiology, diagnosis, and clinical intervention