Comparative genomic analysis of three Leishmania species that cause diverse human disease.

Leishmania parasites cause a broad spectrum of clinical disease. Here we report the sequencing of the genomes of two species of Leishmania: Leishmania infantum and Leishmania braziliensis. The comparison of these sequences with the published genome of Leishmania major reveals marked conservation of synteny and identifies only approximately 200 genes with a differential distribution between the three species. L. braziliensis, contrary to Leishmania species examined so far, possesses components of a putative RNA-mediated interference pathway, telomere-associated transposable elements and spliced leader-associated SLACS retrotransposons. We show that pseudogene formation and gene loss are the principal forces shaping the different genomes. Genes that are differentially distributed between the species encode proteins implicated in host-pathogen interactions and parasite survival in the macrophage.

Delayed detection of a ventricular septal defect following penetrating trauma.

This is a case report of a 27-year-old male who sustained a stab wound to the chest with a resulting penetrating cardiac injury and subsequent emergency thoracotomy. The patient survived his injury and on post-op day two, a holosystolic murmur was noted during physical exam, but he remained hemodynamically stable and intubated. A transthoracic echocardiogram revealed the presence of a ventricular septal defect (VSD), with Doppler flow revealing shunting from the left ventricular cavity into the right ventricular cavity. Ultimately, the clinicians decided upon a trial of extubation with a plan for delayed closure of the VSD. Upon extubation,the patient became hypoxemic with evidence of pulmonary edema; thus, he was re-intubated. The defect was surgically repaired, and the patient had an uneventful recovery thereafter. The purpose of this case report is to present an example of delayed detection of a ventricular septal defect after a penetrating cardiac injury.

Developing a strategy to scale up place-based arts initiatives that support mental health and wellbeing: A realist evaluation of 'Arts for the Blues'.

Place-based arts initiatives are regarded as rooted in local need and as having capacity to engage local assets. However, many place-based arts initiatives remain poorly funded and short-lived, receiving little attention on how to scale up and sustain their activities. In this study we make a unique contribution to knowledge about scaling up place-based arts initiatives that support mental health and wellbeing through focusing on the example of 'Arts for the Blues', an arts-based psychological group intervention designed to reduce depression and improve wellbeing amongst primary care mental health service users in deprived communities. Methodologically, we used realist evaluation to refine the study's theoretical assumptions about scaling up, drawing on the lived and professional experiences of 225 diverse stakeholders' and frontline staff through a series of focus groups and evaluation questions at two stakeholders' events and four training days. Based on our findings, we recommend that to scale up place-based arts initiatives which support mental health and wellbeing: (i) the initiative needs to be adaptable, clear, collaborative, evidence-based, personalised and transformative; (ii) the organisation has to have a relevant need, have an understanding of the arts, has to have resources, inspiration and commitment from staff members, relevant skillsets and help from outside the organisation; (iii) at a policy level it is important to pay attention to attitude shifts towards the arts, meet rules, guidelines and standards expected from services, highlight gaps in provision, seek out early intervention and treatment options, and consider service delivery changes. The presence of champions at a local level and buy-in from managers, local leaders and policy makers are also needed alongside actively seeking to implement arts initiatives in different settings across geographical spread. Our theoretically-based and experientially-refined study provides the first ever scaling up framework developed for place-based arts initiatives that support the mental health and wellbeing, offering opportunities for spread and adoption of such projects in different organisational contexts, locally, nationally and internationally.

Evolutionary dynamics of Clostridium difficile over short and long time scales.

Clostridium difficile has rapidly emerged as the leading cause of antibiotic-associated diarrheal disease, with the transcontinental spread of various PCR ribotypes, including 001, 017, 027 and 078. However, the genetic basis for the emergence of C. difficile as a human pathogen is unclear. Whole genome sequencing was used to analyze genetic variation and virulence of a diverse collection of thirty C. difficile isolates, to determine both macro and microevolution of the species. Horizontal gene transfer and large-scale recombination of core genes has shaped the C. difficile genome over both short and long time scales. Phylogenetic analysis demonstrates C. difficile is a genetically diverse species, which has evolved within the last 1.1-85 million years. By contrast, the disease-causing isolates have arisen from multiple lineages, suggesting that virulence evolved independently in the highly epidemic lineages.

Complete genome sequence and comparative genome analysis of enteropathogenic Escherichia coli O127:H6 strain E2348/69.

Enteropathogenic Escherichia coli (EPEC) was the first pathovar of E. coli to be implicated in human disease; however, no EPEC strain has been fully sequenced until now. Strain E2348/69 (serotype O127:H6 belonging to E. coli phylogroup B2) has been used worldwide as a prototype strain to study EPEC biology, genetics, and virulence. Studies of E2348/69 led to the discovery of the locus of enterocyte effacement-encoded type III secretion system (T3SS) and its cognate effectors, which play a vital role in attaching and effacing lesion formation on gut epithelial cells. In this study, we determined the complete genomic sequence of E2348/69 and performed genomic comparisons with other important E. coli strains. We identified 424 E2348/69-specific genes, most of which are carried on mobile genetic elements, and a number of genetic traits specifically conserved in phylogroup B2 strains irrespective of their pathotypes, including the absence of the ETT2-related T3SS, which is present in E. coli strains belonging to all other phylogroups. The genome analysis revealed the entire gene repertoire related to E2348/69 virulence. Interestingly, E2348/69 contains only 21 intact T3SS effector genes, all of which are carried on prophages and integrative elements, compared to over 50 effector genes in enterohemorrhagic E. coli O157. As E2348/69 is the most-studied pathogenic E. coli strain, this study provides a genomic context for the vast amount of existing experimental data. The unexpected simplicity of the E2348/69 T3SS provides the first opportunity to fully dissect the entire virulence strategy of attaching and effacing pathogens in the genomic context.

Longitudinal monitoring for the emergence of epidermal growth factor C797S resistance mutations in non-small cell lung cancer using blood-based droplet digital PCR.

Validation of assays for the C797S mutation as a biomarker for osimertinib resistance is promising in guiding treatment decision-making for multidrug resistant non-small cell lung cancer. A newly developed droplet digital PCR (ddPCR) assay was used to retrospectively evaluate the emergence of the C797S mutation in six remnant plasma samples in this case report. It was found that the detected emergence of C797S clearly correlated with clinical signs of treatment resistance. Had these data been available to aid treatment selection in real time, there would have been hope for recaptured disease response and control instead of treatment cessation. The results of this study show that highly sensitive ddPCR methods can be used for the monitoring of emergent epidermal growth factor somatic variant mutations in circulation.

The genome sequence of the fish pathogen Aliivibrio salmonicida strain LFI1238 shows extensive evidence of gene decay.

BACKGROUND: The fish pathogen Aliivibrio salmonicida is the causative agent of cold-water vibriosis in marine aquaculture. The Gram-negative bacterium causes tissue degradation, hemolysis and sepsis in vivo. RESULTS: In total, 4 286 protein coding sequences were identified, and the 4.6 Mb genome of A. salmonicida has a six partite architecture with two chromosomes and four plasmids. Sequence analysis revealed a highly fragmented genome structure caused by the insertion of an extensive number of insertion sequence (IS) elements. The IS elements can be related to important evolutionary events such as gene acquisition, gene loss and chromosomal rearrangements. New A. salmonicida functional capabilities that may have been aquired through horizontal DNA transfer include genes involved in iron-acquisition, and protein secretion and play potential roles in pathogenicity. On the other hand, the degeneration of 370 genes and consequent loss of specific functions suggest that A. salmonicida has a reduced metabolic and physiological capacity in comparison to related Vibrionaceae species. CONCLUSION: Most prominent is the loss of several genes involved in the utilisation of the polysaccharide chitin. In particular, the disruption of three extracellular chitinases responsible for enzymatic breakdown of chitin makes A. salmonicida unable to grow on the polymer form of chitin. These, and other losses could restrict the variety of carrier organisms A. salmonicida can attach to, and associate with. Gene acquisition and gene loss may be related to the emergence of A. salmonicida as a fish pathogen.

Development and Clinical Utility of a Blood-Based Test Service for the Rapid Identification of Actionable Mutations in Non-Small Cell Lung Carcinoma.

Nearly 80% of cancer patients do not have genetic mutation results available at initial oncology consultation; up to 25% of patients begin treatment before receiving their results. These factors hinder the ability to pursue optimal treatment strategies. This study validates a blood-based genome-testing service that provides accurate results within 72 hours. We focused on targetable variants in advanced non-small cell lung carcinoma-epidermal growth factor receptor gene (EGFR) variant L858R, exon 19 deletion (ΔE746-A750), and T790M; GTPase Kirsten ras gene (KRAS) variants G12C/D/V; and echinoderm microtubule associated protein like and 4 anaplastic lymphoma receptor tyrosine kinase fusion (EML4-ALK) transcripts 1/2/3. Test development included method and clinical validation using samples from donors with (n = 219) or without (n = 30) cancer. Clinical sensitivity and specificity for each variant ranged from 78.6% to 100% and 94.2% to 100%, respectively. We also report on 1643 non-small cell lung carcinoma samples processed in our CLIA-certified laboratory. Mutation results were available within 72 hours for 94% of the tests evaluated. We detected 10.5% mutations for EGFR sensitizing (n = 2801 samples tested), 13.8% mutations for EGFR resistance (n = 1055), 13.2% mutations in KRAS (n = 3477), and 2% mutations for EML4-ALK fusion (n = 304). This rapid, highly sensitive, and actionable blood-based assay service expands testing options and supports faster treatment decisions.

The genome of Mycobacterium africanum West African 2 reveals a lineage-specific locus and genome erosion common to the M. tuberculosis complex.

BACKGROUND: M. africanum West African 2 constitutes an ancient lineage of the M. tuberculosis complex that commonly causes human tuberculosis in West Africa and has an attenuated phenotype relative to M. tuberculosis. METHODOLOGY/PRINCIPAL FINDINGS: In search of candidate genes underlying these differences, the genome of M. africanum West African 2 was sequenced using classical capillary sequencing techniques. Our findings reveal a unique sequence, RD900, that was independently lost during the evolution of two important lineages within the complex: the "modern" M. tuberculosis group and the lineage leading to M. bovis. Closely related to M. bovis and other animal strains within the M. tuberculosis complex, M. africanum West African 2 shares an abundance of pseudogenes with M. bovis but also with M. africanum West African clade 1. Comparison with other strains of the M. tuberculosis complex revealed pseudogenes events in all the known lineages pointing toward ongoing genome erosion likely due to increased genetic drift and relaxed selection linked to serial transmission-bottlenecks and an intracellular lifestyle. CONCLUSIONS/SIGNIFICANCE: The genomic differences identified between M. africanum West African 2 and the other strains of the Mycobacterium tuberculosis complex may explain its attenuated phenotype, and pave the way for targeted experiments to elucidate the phenotypic characteristic of M. africanum. Moreover, availability of the whole genome data allows for verification of conservation of targets used for the next generation of diagnostics and vaccines, in order to ensure similar efficacy in West Africa.