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BACKGROUND: Multigene panel tests for breast cancer predisposition routinely include ATM as it is now a well-established breast cancer predisposition gene. METHODS: We included ATM in a multigene panel test applied to the Australian Breast Cancer Family Registry (ABCFR), a population-based case-control-family study of breast cancer, with the purpose of estimating the prevalence and penetrance of heterozygous ATM pathogenic variants from the family data, using segregation analysis. RESULTS: The estimated breast cancer hazard ratio for carriers of pathogenic ATM variants in the ABCFR was 1.32 (95% confidence interval 0.45-3.87; P = 0.6). The estimated cumulative risk of breast cancer to age 80 years for heterozygous ATM pathogenic variant carriers was estimated to be 13% (95% CI 4.6-30). CONCLUSIONS: Although ATM has been definitively identified as a breast cancer predisposition gene, further evidence, such as variant-specific penetrance estimates, are needed to inform risk management strategies for carriers of pathogenic variants to increase the clinical utility of population testing of this gene.
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Neoplasias da Mama , Fatores Etários , Idoso de 80 Anos ou mais , Proteínas Mutadas de Ataxia Telangiectasia/genética , Austrália/epidemiologia , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Feminino , Predisposição Genética para Doença , Humanos , Proteínas Supressoras de Tumor/genéticaRESUMO
Population-based estimates of breast cancer risk for carriers of pathogenic variants identified by gene-panel testing are urgently required. Most prior research has been based on women selected for high-risk features and more data is needed to make inference about breast cancer risk for women unselected for family history, an important consideration of population screening. We tested 1464 women diagnosed with breast cancer and 862 age-matched controls participating in the Australian Breast Cancer Family Study (ABCFS), and 6549 healthy, older Australian women enroled in the ASPirin in Reducing Events in the Elderly (ASPREE) study for rare germline variants using a 24-gene-panel. Odds ratios (ORs) were estimated using unconditional logistic regression adjusted for age and other potential confounders. We identified pathogenic variants in 11.1% of the ABCFS cases, 3.7% of the ABCFS controls and 2.2% of the ASPREE (control) participants. The estimated breast cancer OR [95% confidence interval] was 5.3 [2.1-16.2] for BRCA1, 4.0 [1.9-9.1] for BRCA2, 3.4 [1.4-8.4] for ATM and 4.3 [1.0-17.0] for PALB2. Our findings provide a population-based perspective to gene-panel testing for breast cancer predisposition and opportunities to improve predictors for identifying women who carry pathogenic variants in breast cancer predisposition genes.
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BACKGROUND: The androgen receptor (AR) pathway-associated gene nuclear receptor coactivator 2 (NCOA2) has an established oncogenic role in early prostate cancer and likewise is a driver of metastatic disease and castration-resistant prostate cancer. However, its significance as a biomarker in metastatic castration-resistant prostate cancer (mCRPC), both alone and in conjunction with co-occurring AR alterations using a liquid biopsy approach has not been investigated. METHODS: Ninety-one patients were included in this study, (n = 68 receiving an androgen receptor pathway inhibitor and n = 23 receiving taxane chemotherapy). Up to 30 ml of peripheral blood was collected before commencing treatment from each patient. Plasma cell-free DNA, along with a matched germline sample, underwent targeted next-generation sequencing using a validated, highly sensitive in-house prostate cancer panel. Variants in AR and NCOA2 were identified and correlated with clinical outcomes. RESULTS: Plasma AR and NCOA2 aberrations were identified in 35% and 13% of the cohort, respectively, whilst 8% had concurrent AR and NCOA2 alterations. NCOA2 copy number gain and any NCOA2 aberration predicted for lower prostate-specific antigen (PSA) response rates. Likewise, median overall survival was shorter for NCOA2 gain (10.1 vs. 18.3 months; p = .004), remaining significant after adjusting for covariates including circulating tumor DNA fraction and tumor suppressor gene alterations. Importantly, dual AR and NCOA2 aberrations were also associated with inferior outcomes, including no PSA responses in patients treated with AR pathway inhibitors (0% vs. 64%; p = .02). CONCLUSIONS: These data highlight the importance of identifying multiple markers of AR pathway modulation in mCRPC and represent the first instance of the assessment of plasma NCOA2 status as a prognostic biomarker for standard-of-care therapies. Further assessment is warranted to determine if NCOA2 aberrations are a marker of primary resistance to AR pathway inhibitors.
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Coativador 2 de Receptor Nuclear/sangue , Neoplasias de Próstata Resistentes à Castração/sangue , Receptores Androgênicos/sangue , Idoso , Idoso de 80 Anos ou mais , Antagonistas de Receptores de Andrógenos/uso terapêutico , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/sangue , Hidrocarbonetos Aromáticos com Pontes/uso terapêutico , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/mortalidade , Neoplasias de Próstata Resistentes à Castração/patologia , Taxa de Sobrevida , Taxoides/uso terapêuticoRESUMO
VTRNA2-1 is a metastable epiallele with accumulating evidence that methylation at this region is heritable, modifiable and associated with disease including risk and progression of cancer. This study investigated the influence of genetic variation and other factors such as age and adult lifestyle on blood DNA methylation in this region. We first sequenced the VTRNA2-1 gene region in multiple-case breast cancer families in which VTRNA2-1 methylation was identified as heritable and associated with breast cancer risk. Methylation quantitative trait loci (mQTL) were investigated using a prospective cohort study (4500 participants with genotyping and methylation data). The cis-mQTL analysis (334 variants ± 50 kb of the most heritable CpG site) identified 43 variants associated with VTRNA2-1 methylation (p < 1.5 × 10-4); however, these explained little of the methylation variation (R2 < 0.5% for each of these variants). No genetic variants elsewhere in the genome were found to strongly influence VTRNA2-1 methylation. SNP-based heritability estimates were consistent with the mQTL findings (h2 = 0, 95%CI: -0.14 to 0.14). We found no evidence that age, sex, country of birth, smoking, body mass index, alcohol consumption or diet influenced blood DNA methylation at VTRNA2-1. Genetic factors and adult lifestyle play a minimal role in explaining methylation variability at the heritable VTRNA2-1 cluster.
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Metilação de DNA/genética , MicroRNAs/genética , Polimorfismo de Nucleotídeo Único/genética , Idoso , Neoplasias da Mama/genética , Estudos de Casos e Controles , Ilhas de CpG/genética , Feminino , Estudo de Associação Genômica Ampla/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Locos de Características Quantitativas/genéticaRESUMO
Case-control studies of breast cancer have consistently shown that pathogenic variants in CHEK2 are associated with about a 3-fold increased risk of breast cancer. Information about the recurrent protein-truncating variant CHEK2 c.1100delC dominates this estimate. There have been no formal estimates of age-specific cumulative risk of breast cancer for all CHEK2 pathogenic (including likely pathogenic) variants combined. We conducted a population-based case-control-family study of pathogenic CHEK2 variants (26 families, 1071 relatives) and estimated the age-specific cumulative risk of breast cancer using segregation analysis. The estimated hazard ratio for carriers of pathogenic CHEK2 variants (combined) was 4.9 (95% CI 2.5-9.5) relative to non-carriers. The HR for carriers of the CHEK2 c.1100delC variant was estimated to be 3.5 (95% CI 1.02-11.6) and the HR for carriers of all other CHEK2 variants combined was estimated to be 5.7 (95% CI 2.5-12.9). The age-specific cumulative risk of breast cancer was estimated to be 18% (95% CI 11-30%) and 33% (95% CI 21-48%) to age 60 and 80 years, respectively. These findings provide important information for the clinical management of breast cancer risk for women carrying pathogenic variants in CHEK2.
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While gene panel sequencing is becoming widely used for cancer risk prediction, its clinical utility with respect to predicting aggressive prostate cancer (PrCa) is limited by our current understanding of the genetic risk factors associated with predisposition to this potentially lethal disease phenotype. This study included 837 men diagnosed with aggressive PrCa and 7261 controls (unaffected men and men who did not meet criteria for aggressive PrCa). Rare germline pathogenic variants (including likely pathogenic variants) were identified by targeted sequencing of 26 known or putative cancer predisposition genes. We found that 85 (10%) men with aggressive PrCa and 265 (4%) controls carried a pathogenic variant (p < 0.0001). Aggressive PrCa odds ratios (ORs) were estimated using unconditional logistic regression. Increased risk of aggressive PrCa (OR (95% confidence interval)) was identified for pathogenic variants in BRCA2 (5.8 (2.7-12.4)), BRCA1 (5.5 (1.8-16.6)), and ATM (3.8 (1.6-9.1)). Our study provides further evidence that rare germline pathogenic variants in these genes are associated with increased risk of this aggressive, clinically relevant subset of PrCa. These rare genetic variants could be incorporated into risk prediction models to improve their precision to identify men at highest risk of aggressive prostate cancer and be used to identify men with newly diagnosed prostate cancer who require urgent treatment.
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BACKGROUND: Diphtheria is a potentially fatal respiratory disease caused by toxigenic Corynebacterium diphtheriae. Although resistance to erythromycin has been recognized, ß-lactam resistance in toxigenic diphtheria has not been described. Here, we report a case of fatal respiratory diphtheria caused by toxigenic C. diphtheriae resistant to penicillin and all other ß-lactam antibiotics, and describe a novel mechanism of inducible carbapenem resistance associated with the acquisition of a mobile resistance element. METHODS: Long-read whole-genome sequencing was performed using Pacific Biosciences Single Molecule Real-Time sequencing to determine the genome sequence of C. diphtheriae BQ11 and the mechanism of ß-lactam resistance. To investigate the phenotypic inducibility of meropenem resistance, short-read sequencing was performed using an Illumina NextSeq500 sequencer on the strain both with and without exposure to meropenem. RESULTS: BQ11 demonstrated high-level resistance to penicillin (benzylpenicillin minimum inhibitory concentration [MIC]â ≥â 256 µg/ml), ß-lactam/ß-lactamase inhibitors and cephalosporins (amoxicillin/clavulanic acid MICâ ≥â 256 µg/mL; ceftriaxone MICâ ≥â 8 µg/L). Genomic analysis of BQ11 identified acquisition of a novel transposon carrying the penicillin-binding protein (PBP) Pbp2c, responsible for resistance to penicillin and cephalosporins. When strain BQ11 was exposed to meropenem, selective pressure drove amplification of the transposon in a tandem array and led to a corresponding change from a low-level to a high-level meropenem-resistant phenotype. CONCLUSIONS: We have identified a novel mechanism of inducible antibiotic resistance whereby isolates that appear to be carbapenem susceptible on initial testing can develop in vivo resistance to carbapenems with repeated exposure. This phenomenon could have significant implications for the treatment of C. diphtheriae infection, and may lead to clinical failure.
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Corynebacterium diphtheriae , Difteria , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Corynebacterium diphtheriae/genética , Difteria/tratamento farmacológico , Humanos , Lactamas/uso terapêutico , Testes de Sensibilidade Microbiana , Penicilinas/uso terapêuticoRESUMO
PURPOSE: To characterize the spectrum of BRCA1 and BRCA2 pathogenic germline variants in women from south-west Poland and west Ukraine affected with breast or ovarian cancer. Testing in women at high risk of breast and ovarian cancer in these regions is currently mainly limited to founder mutations. METHODS: Unrelated women affected with breast and/or ovarian cancer from Poland (n = 337) and Ukraine (n = 123) were screened by targeted sequencing. Excluded from targeted sequencing were 34 Polish women who had previously been identified as carrying a founder mutation in BRCA1. No prior testing had been conducted among the Ukrainian women. Thus, this study screened BRCA1 and BRCA2 in the germline DNA of 426 women in total. RESULTS: We identified 31 and 18 women as carriers of pathogenic/likely pathogenic (P/LP) genetic variants in BRCA1 and BRCA2, respectively. We observed five BRCA1 and eight BRCA2 P/LP variants (13/337, 3.9%) in the Polish women. Combined with the 34/337 (10.1%) founder variants identified prior to this study, the overall P/LP variant frequency in the Polish women was thus 14% (47/337). Among the Ukrainian women, 16/123 (13%) women were identified as carrying a founder mutation and 20/123 (16.3%) were found to carry non-founder P/LP variants (10 in BRCA1 and 10 in BRCA2). CONCLUSIONS: These results indicate that genetic testing in women at high risk of breast and ovarian cancer in Poland and Ukraine should not be limited to founder mutations. Extended testing will enhance risk stratification and management for these women and their families.
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Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença , Testes Genéticos/métodos , Mutação em Linhagem Germinativa , Neoplasias Ovarianas/genética , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/patologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Neoplasias Ovarianas/epidemiologia , Neoplasias Ovarianas/patologia , Polônia/epidemiologia , Ucrânia/epidemiologiaRESUMO
Plasma circulating tumor DNA (ctDNA) analysis has emerged as a minimally invasive means to perform molecular tumor typing. Here we developed a custom ultra-sensitive ctDNA next-generation sequencing assay using molecular barcoding technology and off-the-shelf reagents combined with bioinformatics tools for enhanced ctDNA analysis. Assay performance was assessed via a spike-in experiment and the technique was applied to analyze 41 plasma samples from men with advanced prostate cancer. Orthogonal validation was performed using a commercial assay. Sensitivity and specificity of 93 and 99.5% were recorded for ultra-rare somatic variants (<1%), with high concordance observed between the in-house and commercial assays. The optimized protocol dramatically improved the efficiency of the assay and enabled the detection of low-frequency somatic variants from plasma cell-free DNA (cfDNA).
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DNA Tumoral Circulante/sangue , Sequenciamento de Nucleotídeos em Larga Escala , Análise de Sequência de DNA , Ácidos Nucleicos Livres/sangue , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento de Nucleotídeos em Larga Escala/normas , Humanos , Biópsia Líquida/métodos , Biópsia Líquida/normas , Masculino , Neoplasias da Próstata/sangue , Neoplasias da Próstata/patologia , Sensibilidade e Especificidade , Análise de Sequência de DNA/métodos , Análise de Sequência de DNA/normasRESUMO
Limisphaera ngatamarikiensis NGM72.4T is a thermophilic representative of the class Verrucomicrobiae Isolated from geothermally heated subaqueous clay sediments from a Ngatamariki hotspring in Aotearoa New Zealand, the 3,908,748-bp genome was sequenced using the Illumina HiSeq 2500 platform. Annotation revealed 3,083 coding sequences, including 3,031 proteins, 3 rRNA genes, and 46 tRNA genes.
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Few genetic risk factors have been demonstrated to be specifically associated with aggressive prostate cancer (PrCa). Here, we report a case-case study of PrCa comparing the prevalence of germline pathogenic/likely pathogenic (P/LP) genetic variants in 787 men with aggressive disease and 769 with nonaggressive disease. Overall, we observed P/LP variants in 11.4% of men with aggressive PrCa and 9.8% of men with nonaggressive PrCa (two-tailed Fisher's exact tests, P = .28). The proportion of BRCA2 and ATM P/LP variant carriers in men with aggressive PrCa exceeded that observed in men with nonaggressive PrCa; 18/787 carriers (2.3%) and 4/769 carriers (0.5%), P = .004, and 14/787 carriers (0.02%) and 5/769 carriers (0.01%), P = .06, respectively. Our findings contribute to the extensive international effort to interpret the genetic variation identified in genes included on gene-panel tests, for which there is currently an insufficient evidence-base for clinical translation in the context of PrCa risk.
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Células Germinativas/metabolismo , Mutação em Linhagem Germinativa/genética , Neoplasias da Próstata/genética , Idoso , Proteína BRCA2/genética , Estudos de Coortes , Predisposição Genética para Doença/genética , Testes Genéticos/métodos , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Próstata/patologia , Antígeno Prostático Específico/genética , Neoplasias da Próstata/patologiaRESUMO
The advent of gene panel testing is challenging the previous practice of using clinically defined cancer family syndromes to inform single-gene genetic screening. Individual and family cancer histories that would have previously indicated testing of a single gene or a small number of related genes are now, increasingly, leading to screening across gene panels that contain larger numbers of genes. We have applied a gene panel test that included four DNA mismatch repair (MMR) genes (MLH1, MSH2, MSH6 and PMS2) to an Australian population-based case-control-family study of breast cancer. Altogether, eight pathogenic variants in MMR genes were identified: six in 1421 case-families (0.4%, 4 MSH6 and 2 PMS2) and two in 833 control-families (0.2%, one each of MLH1 and MSH2). This testing highlights the current and future challenges for clinical genetics in the context of anticipated gene panel-based population-based screening that includes the MMR genes. This testing is likely to provide additional opportunities for cancer prevention via cascade testing for Lynch syndrome and precision medicine for breast cancer treatment.
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Neoplasias da Mama/genética , Reparo de Erro de Pareamento de DNA/genética , Mutação em Linhagem Germinativa , Estudos de Casos e Controles , Estudos de Coortes , Neoplasias Colorretais Hereditárias sem Polipose/genética , Proteínas de Ligação a DNA/genética , Feminino , Perfilação da Expressão Gênica/métodos , Predisposição Genética para Doença , Humanos , Masculino , Endonuclease PMS2 de Reparo de Erro de Pareamento/genética , Proteína 1 Homóloga a MutL/genética , Proteína 2 Homóloga a MutS/genética , New South Wales , Linhagem , Sistema de Registros/estatística & dados numéricos , VitóriaRESUMO
Cultivation-independent surveys have shown that the desert soils of Antarctica harbour surprisingly rich microbial communities. Given that phototroph abundance varies across these Antarctic soils, an enduring question is what supports life in those communities with low photosynthetic capacity. Here we provide evidence that atmospheric trace gases are the primary energy sources of two Antarctic surface soil communities. We reconstructed 23 draft genomes from metagenomic reads, including genomes from the candidate bacterial phyla WPS-2 and AD3. The dominant community members encoded and expressed high-affinity hydrogenases, carbon monoxide dehydrogenases, and a RuBisCO lineage known to support chemosynthetic carbon fixation. Soil microcosms aerobically scavenged atmospheric H2 and CO at rates sufficient to sustain their theoretical maintenance energy and mediated substantial levels of chemosynthetic but not photosynthetic CO2 fixation. We propose that atmospheric H2, CO2 and CO provide dependable sources of energy and carbon to support these communities, which suggests that atmospheric energy sources can provide an alternative basis for ecosystem function to solar or geological energy sources. Although more extensive sampling is required to verify whether this process is widespread in terrestrial Antarctica and other oligotrophic habitats, our results provide new understanding of the minimal nutritional requirements for life and open the possibility that atmospheric gases support life on other planets.
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Atmosfera/química , Ciclo do Carbono , Monóxido de Carbono/metabolismo , Clima Desértico , Hidrogênio/metabolismo , Microbiologia do Solo , Solo/química , Regiões Antárticas , Dióxido de Carbono/análise , Dióxido de Carbono/metabolismo , Monóxido de Carbono/análise , Ecossistema , Exobiologia , Genoma/genética , Hidrogênio/análise , Metagenômica , Oxirredução , Fotossíntese , FilogeniaRESUMO
Pasteurella multocida is an important veterinary pathogen that produces a wide range of lipopolysaccharide (LPS) structures, many of which mimic host glycoproteins. In this study, we complete our analysis of the LPS produced by the P. multocida Heddleston serovars by reporting the LPS structure and the LPS outer core biosynthesis loci of the type strains representing Heddleston serovars 6, 7 and 16. Genetic analysis revealed that the type strains representing serovars 6 and 7 share the same LPS outer core biosynthesis locus which we have designated LPS genotype L4. Comparative bioinformatic analysis revealed that although the serovar 16 type strain contained a different LPS locus, L8, there was a significant degree of nucleotide identity between the L4 and L8 loci. Structural analysis revealed that the LPS glycoforms produced by the L4 and L8 strains all contained the highly conserved inner core produced by all other P. multocida strains examined to date. The residues within the LPS outer core produced by the L4 and L8 strains were either Gal or derivatives of Gal; unlike all other P. multocida Heddleston type strains examined there are no heptosyltransferases encoded in the L4 and L8 outer core biosynthesis loci. The structure of the L4 LPS outer core produced by the serovar 6 type strain consisted of ß-Gal-(1-3)-ß-N-acetylgalactosamine (GalNAc)-(1-4)-ß-GalNAc3OAc-(1-4)-α-GalNAc3OAc-(1-3)-ß-Gal, whereas the serovar 7 type strain produced a highly truncated LPS outer core containing only a single ß-Gal residue. The structure of the L8 LPS outer core produced by the serovar 16 type strain consisted of ß-Gal-(1-3)-ß-GalNAc-(1-4)-(α-GalNAc-(1-3)-)-α-GalNAc.
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Gammaproteobacteria/genética , Genótipo , Lipopolissacarídeos/química , Sorogrupo , Gammaproteobacteria/metabolismo , Lipopolissacarídeos/biossínteseRESUMO
Mammals express the sialic acids N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) on cell surfaces, where they act as receptors for pathogens, including influenza A virus (IAV). Neu5Gc is synthesized from Neu5Ac by the enzyme cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH). In humans, this enzyme is inactive and only Neu5Ac is produced. Ferrets are susceptible to human-adapted IAV strains and have been the dominant animal model for IAV studies. Here we show that ferrets, like humans, do not synthesize Neu5Gc. Genomic analysis reveals an ancient, nine-exon deletion in the ferret CMAH gene that is shared by the Pinnipedia and Musteloidia members of the Carnivora. Interactions between two human strains of IAV with the sialyllactose receptor (sialic acid--α2,6Gal) confirm that the type of terminal sialic acid contributes significantly to IAV receptor specificity. Our results indicate that exclusive expression of Neu5Ac contributes to the susceptibility of ferrets to human-adapted IAV strains.
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Sequência de Bases , Furões/virologia , Oxigenases de Função Mista/química , Ácido N-Acetilneuramínico/metabolismo , Receptores Virais/metabolismo , Deleção de Sequência , Animais , Caniformia/genética , Caniformia/imunologia , Caniformia/virologia , Sequência de Carboidratos , Éxons , Furões/genética , Furões/imunologia , Expressão Gênica , Humanos , Vírus da Influenza A/genética , Vírus da Influenza A/metabolismo , Oxigenases de Função Mista/deficiência , Oxigenases de Função Mista/genética , Dados de Sequência Molecular , Ácido N-Acetilneuramínico/química , Ácidos Neuramínicos/química , Ácidos Neuramínicos/metabolismo , Infecções por Orthomyxoviridae/virologia , Receptores Virais/química , Receptores Virais/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tropismo ViralAssuntos
Escherichia coli/genética , Escherichia coli/isolamento & purificação , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Plasmídeos/genética , Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Humanos , Klebsiella pneumoniae/efeitos dos fármacos , Tailândia/epidemiologiaRESUMO
Molecular surveys of aphotic habitats have indicated the presence of major uncultured lineages phylogenetically classified as members of the Cyanobacteria. One of these lineages has recently been proposed as a nonphotosynthetic sister phylum to the Cyanobacteria, the Melainabacteria, based on recovery of population genomes from human gut and groundwater samples. Here, we expand the phylogenomic representation of the Melainabacteria through sequencing of six diverse population genomes from gut and bioreactor samples supporting the inference that this lineage is nonphotosynthetic, but not the assertion that they are strictly fermentative. We propose that the Melainabacteria is a class within the phylogenetically defined Cyanobacteria based on robust monophyly and shared ancestral traits with photosynthetic representatives. Our findings are consistent with theories that photosynthesis occurred late in the Cyanobacteria and involved extensive lateral gene transfer and extends the recognized functionality of members of this phylum.
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Reatores Biológicos/microbiologia , Cianobactérias/genética , Genoma Bacteriano , Phascolarctidae/microbiologia , Filogenia , Animais , Evolução Biológica , Cianobactérias/classificação , Cianobactérias/isolamento & purificação , Cianobactérias/metabolismo , Fezes/microbiologia , Genética Populacional , Masculino , Fotossíntese , RNA Ribossômico 16S , Eliminação de Resíduos Líquidos/instrumentação , Eliminação de Resíduos Líquidos/métodosRESUMO
Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrheal disease around the globe, causing an estimated 380,000 deaths annually. The disease is caused by a wide variety of strains. Here, we report the genome sequence of ETEC strain B2C, which was isolated from an American soldier in Vietnam.
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Urinary tract infections (UTIs) are one of the most commonly acquired bacterial infections in humans, and uropathogenic Escherichia coli strains are responsible for over 80% of all cases. The standard method for identification of uropathogens in clinical laboratories is cultivation, primarily using solid growth media under aerobic conditions, coupled with morphological and biochemical tests of typically a single isolate colony. However, these methods detect only culturable microorganisms, and characterization is phenotypic in nature. Here, we explored the genotypic identity of communities in acute uncomplicated UTIs from 50 individuals by using culture-independent amplicon pyrosequencing and whole-genome and metagenomic shotgun sequencing. Genus-level characterization of the UTI communities was achieved using the 16S rRNA gene (V8 region). Overall UTI community richness was very low in comparison to other human microbiomes. We strain-typed Escherichia-dominated UTIs using amplicon pyrosequencing of the fimbrial adhesin gene, fimH. There were nine highly abundant fimH types, and each UTI sample was dominated by a single type. Molecular analysis of the corresponding clinical isolates revealed that in the majority of cases the isolate was representative of the dominant taxon in the community at both the genus and the strain level. Shotgun sequencing was performed on a subset of eight E. coli urine UTI and isolate pairs. The majority of UTI microbial metagenomic sequences mapped to isolate genomes, confirming the results obtained using phylogenetic markers. We conclude that for the majority of acute uncomplicated E. coli-mediated UTIs, single cultured isolates are diagnostic of the infection. IMPORTANCE In clinical practice, the diagnosis and treatment of acute uncomplicated urinary tract infection (UTI) are based on analysis of a single bacterial isolate cultured from urine, and it is assumed that this isolate represents the dominant UTI pathogen. However, these methods detect only culturable bacteria, and the existence of multiple pathogens as well as strain diversity within a single infection is not examined. Here, we explored bacteria present in acute uncomplicated UTIs using culture-independent sequence-based methods. Escherichia coli was the most common organism identified, and analysis of E. coli dominant UTI samples and their paired clinical isolates revealed that in the majority of infections the cultured isolate was representative of the dominant taxon at both the genus and the strain level. Our data demonstrate that in most cases single cultured isolates are diagnostic of UTI and are consistent with the notion of bottlenecks that limit strain diversity during UTI pathogenesis.
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Infecções por Escherichia coli/microbiologia , Metagenoma , Infecções Urinárias/microbiologia , Escherichia coli Uropatogênica/classificação , Escherichia coli Uropatogênica/genética , Adesinas de Escherichia coli/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Proteínas de Fímbrias/genética , Genes de RNAr , Genótipo , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Tipagem Molecular , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Escherichia coli Uropatogênica/isolamento & purificação , Adulto JovemRESUMO
OBJECTIVES: To obtain an expanded understanding of antibiotic resistance evolution in vivo, particularly in the context of vancomycin exposure. METHODS: The whole genomes of six consecutive methicillin-resistant Staphylococcus aureus blood culture isolates (ST239-MRSA-III) from a single patient exposed to various antimicrobials (over a 77 day period) were sequenced and analysed. RESULTS: Variant analysis revealed the existence of non-susceptible sub-populations derived from a common susceptible ancestor, with the predominant circulating clone(s) selected for by type and duration of antimicrobial exposure. CONCLUSIONS: This study highlights the dynamic nature of bacterial evolution and that non-susceptible sub-populations can emerge from clouds of variation upon antimicrobial exposure. Diagnostically, this has direct implications for sample selection when using whole-genome sequencing as a tool to guide clinical therapy. In the context of bacteraemia, deep sequencing of bacterial DNA directly from patient blood samples would avoid culture 'bias' and identify mutations associated with circulating non-susceptible sub-populations, some of which may confer cross-resistance to alternate therapies.