Microreact: visualizing and sharing data for genomic epidemiology and phylogeography

Visualization is frequently used to aid our interpretation of complex datasets. Within microbial genomics, visualizing the relationships between multiple genomes as a tree provides a framework onto which associated data (geographical, temporal, phenotypic and epidemiological) are added to generate hypotheses and to explore the dynamics of the system under investigation. Selected static images are then used within publications to highlight the key findings to a wider audience. However, these images are a very inadequate way of exploring and interpreting the richness of the data. There is, therefore, a need for flexible, interactive software that presents the population genomic outputs and associated data in a user-friendly manner for a wide range of end users, from trained bioinformaticians to front-line epidemiologists and health workers. Here, we present Microreact, a web application for the easy visualization of datasets consisting of any combination of trees, geographical, temporal and associated metadata. Data files can be uploaded to Microreact directly via the web browser or by linking to their location (e.g. from Google Drive/Dropbox or via API), and an integrated visualization via trees, maps, timelines and tables provides interactive querying of the data. The visualization can be shared as a permanent web link among collaborators, or embedded within publications to enable readers to explore and download the data. Microreact can act as an end point for any tool or bioinformatic pipeline that ultimately generates a tree, and provides a simple, yet powerful, visualization method that will aid research and discovery and the open sharing of datasets

The widespread use of topical antimicrobials enriches for resistance in Staphylococcus aureus isolated from patients with atopic dermatitis.

BACKGROUND: Carriage rates of Staphylococcus aureus on affected skin in atopic dermatitis (AD) are approximately 70%. Increasing disease severity during flares and overall disease severity correlate with increased burden of S. aureus. Treatment in AD therefore often targets S. aureus with topical and systemic antimicrobials. OBJECTIVES: To determine whether antimicrobial sensitivities and genetic determinants of resistance differed in S. aureus isolates from the skin of children with AD and healthy child nasal carriers. METHODS: In this case-control study, we compared S. aureus isolates from children with AD (n = 50) attending a hospital dermatology department against nasal carriage isolates from children without skin disease (n = 49) attending a hospital emergency department for noninfective conditions. Using whole genome sequencing we generated a phylogenetic framework for the isolates based on variation in the core genome, then compared antimicrobial resistance phenotypes and genotypes between disease groups. RESULTS: Staphylococcus aureus from cases and controls had on average similar numbers of phenotypic resistances per isolate. Case isolates differed in their resistance patterns, with fusidic acid resistance (FusR ) being significantly more frequent in AD (P = 0·009). The genetic basis of FusR also differentiated the populations, with chromosomal mutations in fusA predominating in AD (P = 0·049). Analysis revealed that FusR evolved multiple times and via multiple mechanism in the population. Carriage of plasmid-derived qac genes, which have been associated with reduced susceptibility to antiseptics, was eight times more frequent in AD (P = 0·016). CONCLUSIONS: The results suggest that strong selective pressure drives the emergence and maintenance of specific resistances in AD

Genomic surveillance of Acinetobacter baumannii in the Philippines, 2013-2014

This work was supported by a Newton Fund award from the Medical Research Council (UK) MR/N019296/1 and the Philippine Council for Health Research and Development. This work was also partially supported by research grant U01CA207167 from the U.S. National Institutes of Health. S.A. and D.M.A. were additionally supported by the National Institute for Health Research (UK) Global Health Research Unit on genomic Surveillance of AMR (16_136_111) and by the Centre for Genomic Pathogen Surveillance.Objective: Acinetobacter baumannii is an opportunistic nosocomial pathogen that has increasingly become resistant to carbapenems worldwide. In the Philippines, rates of carbapenem resistance and multidrug resistance are above 50%. We undertook a genomic study of carbapenem-resistant A. baumannii in the Philippines to characterize the population diversity and antimicrobial resistance mechanisms. Methods: We sequenced the whole genomes of 117 A. baumannii isolates recovered by 16 hospitals in the Philippines between 2013 and 2014. From the genome sequences, we determined the multilocus sequence type, presence of acquired determinants of antimicrobial resistance and relatedness between isolates. We also compared the phenotypic and genotypic resistance results. Results: Carbapenem resistance was mainly explained by acquisition of the class-D beta-lactamase gene bla(OXA-23). The concordance between phenotypic and genotypic resistance to imipenem was 98.15%, and it was 94.97% overall for the seven antibiotics analysed. Twenty-two different sequence types were identified, including 7 novel types. The population was dominated by the high-risk international clone 2 (i.e. clonal complex 92), in particular by ST195 and ST208 and their single locus variants. Using whole-genome sequencing, we identified local clusters representing potentially undetected nosocomial outbreaks, as well as multi-hospital clusters that indicated interhospital dissemination. Comparison with global genomes suggested that the establishment of carbapenem-resistant international clone 2 in the Philippines is likely the result of clonal expansion and geographical dissemination, and at least partly explained by inadequate hospital infection control and prevention. Discussion: This is the first extensive genomic study of carbapenem-resistant A. baumannii in the Philippines, and it underscores the importance of hospital infection control and prevention measures to contain high-risk clones.Publisher PDFPeer reviewe

Dvegrajski zbornik, br. 4, zbornik radova znanstvenih skupova "Crtice uz povijesti Kanfanarštine" 2016. i 2017., gl. urednik Marko Jelenić, Kanfanar: Općina Kanfanar / Udruga za očuvanje i promociju nasljeđa - Dvegrajci, 2018., 287 str.

Antibiotic resistance forms a serious threat to the health of hospitalised patients, rendering otherwise treatable bacterial infections potentially life-threatening. A thorough understanding of the mechanisms by which resistance spreads between patients in different hospitals is required in order to design effective control strategies. We measured the differences between bacterial populations of 52 hospitals in the United Kingdom and Ireland, using whole-genome sequences from 1085 MRSA clonal complex 22 isolates collected between 1998 and 2012. The genetic differences between bacterial populations were compared with the number of patients transferred between hospitals and their regional structure. The MRSA populations within single hospitals, regions and countries were genetically distinct from the rest of the bacterial population at each of these levels. Hospitals from the same patient referral regions showed more similar MRSA populations, as did hospitals sharing many patients. Furthermore, the bacterial populations from different time-periods within the same hospital were generally more similar to each other than contemporaneous bacterial populations from different hospitals. We conclude that, while a large part of the dispersal and expansion of MRSA takes place among patients seeking care in single hospitals, inter-hospital spread of resistant bacteria is by no means a rare occurrence. Hospitals are exposed to constant introductions of MRSA on a number of levels: (1) most MRSA is received from hospitals that directly transfer large numbers of patients, while (2) fewer introductions happen between regions or (3) across national borders, reflecting lower numbers of transferred patients. A joint coordinated control effort between hospitals, is therefore paramount for the national control of MRSA, antibiotic-resistant bacteria and other hospital-associated pathogens

Bacterial microevolution and the Pangenome

The comparison of multiple genome sequences sampled from a bacterial population reveals considerable diversity in both the core and the accessory parts of the pangenome. This diversity can be analysed in terms of microevolutionary events that took place since the genomes shared a common ancestor, especially deletion, duplication, and recombination. We review the basic modelling ingredients used implicitly or explicitly when performing such a pangenome analysis. In particular, we describe a basic neutral phylogenetic framework of bacterial pangenome microevolution, which is not incompatible with evaluating the role of natural selection. We survey the different ways in which pangenome data is summarised in order to be included in microevolutionary models, as well as the main methodological approaches that have been proposed to reconstruct pangenome microevolutionary history

Genome sequencing and molecular characterisation of Staphylococcus aureus ST772-MRSA-V, “Bengal Bay Clone”

Background: The PVL-positive ST772-MRSA-V is an emerging community-associated (CA-) MRSA clone that has been named Bengal Bay Clone since most patients have epidemiological connections to the Indian subcontinent. It is found increasingly common in other areas of the world. Methods: One isolate of ST772-MRSA-V was sequenced using the Illumina Genome Analyzer System. After initial assembling the multiple sequence contigs were analysed using different in-house annotation scripts. Results were compared to microarray hybridisation results of clinical isolates of ST772-MRSA-V, of related strains and to another ST772-MRSA-V genome sequence. Results: According to MLST e-burst analysis, ST772-MRSA-V belongs to Clonal Complex (CC)1, differing from ST1 only in one MLST allele (pta-22). However, there are several additional differences including agr alleles (group II rather than III), capsule type (5 rather than 8), the presence of the egc enterotoxin gene cluster and of the enterotoxin homologue ORF CM14 as well as the absence of the enterotoxin H gene seh. Enterotoxin genes sec and sel are present. ST772-MRSA-V harbours the genes encoding enterotoxin A (sea) and PVL (lukS/F-PV). Both are located on the same prophage. Conclusions: ST772-MRSA-V may have emerged from the same lineage as globally spread CC1 and CC5 strains. It has acquired a variety of virulence factors, and for a CA-MRSA strain it has an unusually high number of genes associated with antibiotic resistance

Potential mechanisms underlying the acute lung dysfunction and bacterial extrapulmonary dissemination during Burkholderia cenocepacia respiratory infection

<p>Abstract</p> <p>Background</p> <p><it>Burkholderia cenocepacia</it>, an opportunistic pathogen that causes lung infections in cystic fibrosis (CF) patients, is associated with rapid and usually fatal lung deterioration due to necrotizing pneumonia and sepsis, a condition known as cepacia syndrome. The key bacterial determinants associated with this poor clinical outcome in CF patients are not clear. In this study, the cytotoxicity and procoagulant activity of <it>B. cenocepacia </it>from the ET-12 lineage, that has been linked to the cepacia syndrome, and four clinical isolates recovered from CF patients with mild clinical courses were analysed in both <it>in vitro </it>and <it>in vivo </it>assays.</p> <p>Methods</p> <p><it>B. cenocepacia-</it>infected BEAS-2B epithelial respiratory cells were used to investigate the bacterial cytotoxicity assessed by the flow cytometric detection of cell staining with propidium iodide. Bacteria-induced procoagulant activity in cell cultures was assessed by a colorimetric assay and by the flow cytometric detection of tissue factor (TF)-bearing microparticles in cell culture supernatants. Bronchoalveolar lavage fluids (BALF) from intratracheally infected mice were assessed for bacterial proinflammatory and procoagulant activities as well as for bacterial cytotoxicity, by the detection of released lactate dehydrogenase.</p> <p>Results</p> <p>ET-12 was significantly more cytotoxic to cell cultures but clinical isolates Cl-2, Cl-3 and Cl-4 exhibited also a cytotoxic profile. ET-12 and CI-2 were similarly able to generate a TF-dependent procoagulant environment in cell culture supernatant and to enhance the release of TF-bearing microparticles from infected cells. In the <it>in vivo </it>assay, all bacterial isolates disseminated from the mice lungs, but Cl-2 and Cl-4 exhibited the highest rates of recovery from mice livers. Interestingly, Cl-2 and Cl-4, together with ET-12, exhibited the highest cytotoxicity. All bacteria were similarly capable of generating a procoagulant and inflammatory environment in animal lungs.</p> <p>Conclusion</p> <p><it>B. cenocepacia </it>were shown to exhibit cytotoxic and procoagulant activities potentially implicated in bacterial dissemination into the circulation and acute pulmonary decline detected in susceptible CF patients. Improved understanding of the mechanisms accounting for <it>B. cenocepacia</it>-induced clinical decline has the potential to indicate novel therapeutic strategies to be included in the care <it>B. cenocepacia</it>-infected patients.</p

Adaptive Evolution of Staphylococcus aureus during Chronic Endobronchial Infection of a Cystic Fibrosis Patient

The molecular adaptation of Staphylococcus aureus to its host during chronic infection is not well understood. Comparative genome sequencing of 3 S. aureus isolates obtained sequentially over 26 months from the airways of a cystic fibrosis patient, revealed variation in phage content, and genetic polymorphisms in genes which influence antibiotic resistance, and global regulation of virulence. The majority of polymorphisms were isolate-specific suggesting the existence of an heterogeneous infecting population that evolved from a single infecting strain of S. aureus. The genetic variation identified correlated with differences in growth rate, hemolytic activity, and antibiotic sensitivity, implying a profound effect on the ecology of S. aureus. In particular, a high frequency of mutations in loci associated with the alternate transcription factor SigB, were observed. The identification of genes under diversifying selection during long-term infection may inform the design of novel therapeutics for the control of refractory chronic infections

Complete Killing of Caenorhabditis elegans by Burkholderia pseudomallei Is Dependent on Prolonged Direct Association with the Viable Pathogen

Background: Burkholderia pseudomallei is the causative agent of melioidosis, a disease of significant morbidity and mortality in both human and animals in endemic areas. Much remains to be known about the contributions of genotypic variations within the bacteria and the host, and environmental factors that lead to the manifestation of the clinical symptoms of melioidosis. Methodology/Principal Findings: In this study, we showed that different isolates of B. pseudomallei have divergent ability to kill the soil nematode Caenorhabditis elegans. The rate of nematode killing was also dependent on growth media: B. pseudomallei grown on peptone-glucose media killed C. elegans more rapidly than bacteria grown on the nematode growth media. Filter and bacteria cell-free culture filtrate assays demonstrated that the extent of killing observed is significantly less than that observed in the direct killing assay. Additionally, we showed that B. pseudomallei does not persistently accumulate within the C. elegans gut as brief exposure to B. pseudomallei is not sufficient for C. elegans infection. Conclusions/Significance: A combination of genetic and environmental factors affects virulence. In addition, we have also demonstrated that a Burkholderia-specific mechanism mediating the pathogenic effect in C. elegans requires proliferating B

Horizontal DNA transfer mechanisms of bacteria as weapons of intragenomic conflict

Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell-cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing "arms race." Reduced rates of transformation have also been observed in cells infected by MGEs that reduce the concentration of extracellular DNA through secretion of DNases. Simulations predicted that either mechanism of limiting transformation would benefit individual MGEs, but also that this tactic's effectiveness was limited by competition with other MGEs coinfecting the same cell. A further observed behaviour we hypothesised to reduce elimination by transformation was MGE activation when cells become competent. Our model predicted that this response was effective at counteracting transformation independently of competing MGEs. Therefore, this framework is able to explain both common properties of MGEs, and the seemingly paradoxical bacterial behaviours of transformation and cell-cell killing within clonally related populations, as the consequences of intragenomic conflict between self-replicating chromosomes and parasitic MGEs. The antagonistic nature of the different mechanisms of HDT over short timescales means their contribution to bacterial evolution is likely to be substantially greater than previously appreciated