RESUMO
BACKGROUND: Hospital outbreaks of carbapenemase-producing organisms, such as blaIMP-4-containing organisms, are an increasing threat to patient safety. OBJECTIVES: To investigate the genomic dynamics of a 10 year (2006-15) outbreak of blaIMP-4-containing organisms in a burns unit in a hospital in Sydney, Australia. METHODS: All carbapenem-non-susceptible or MDR clinical isolates (2006-15) and a random selection of equivalent or ESBL-producing environmental isolates (2012-15) were sequenced [short-read (Illumina), long-read (Oxford Nanopore Technology)]. Sequence data were used to assess genetic relatedness of isolates (Mash; mapping and recombination-adjusted phylogenies), perform in silico typing (MLST, resistance genes and plasmid replicons) and reconstruct a subset of blaIMP plasmids for comparative plasmid genomics. RESULTS: A total of 46/58 clinical and 67/96 environmental isolates contained blaIMP-4. All blaIMP-4-positive organisms contained five or more other resistance genes. Enterobacter cloacae was the predominant organism, with 12 other species mainly found in either the environment or patients, some persisting despite several cleaning methods. On phylogenetic analysis there were three genetic clusters of E. cloacae containing both clinical and environmental isolates, and an additional four clusters restricted to either reservoir. blaIMP-4 was mostly found as part of a cassette array (blaIMP-4-qacG2-aacA4-catB3) in a class 1 integron within a previously described IncM2 plasmid (pEl1573), with almost complete conservation of this cassette across the species over the 10 years. Several other plasmids were also implicated, including an IncF plasmid backbone not previously widely described in association with blaIMP-4. CONCLUSIONS: Genetic backgrounds disseminating blaIMP-4 can persist, diversify and evolve amongst both human and environmental reservoirs during a prolonged outbreak despite intensive prevention efforts.
Assuntos
Proteínas de Bactérias , beta-Lactamases , Antibacterianos/farmacologia , Austrália/epidemiologia , Proteínas de Bactérias/genética , Surtos de Doenças , Genômica , Hospitais , Humanos , Integrons , Testes de Sensibilidade Microbiana , Tipagem de Sequências Multilocus , Filogenia , Plasmídeos/genética , beta-Lactamases/genética , beta-Lactamases/metabolismoRESUMO
BACKGROUND: Hospital antimicrobial stewardship strategies, such as 'Start Smart, Then Focus' in the UK, balance the need for prompt, effective antibiotic treatment with the need to limit antibiotic overuse using 'review and revise'. However, only a minority of review decisions are to stop antibiotics. Research suggests that this is due to both behavioural and organizational factors. OBJECTIVES: To develop and optimize the Antibiotic Review Kit (ARK) intervention. ARK is a complex digital, organizational and behavioural intervention that supports implementation of 'review and revise' to help healthcare professionals safely stop unnecessary antibiotics. METHODS: A theory-, evidence- and person-based approach was used to develop and optimize ARK and its implementation. This was done through iterative stakeholder consultation and in-depth qualitative research with doctors, nurses and pharmacists in UK hospitals. Barriers to and facilitators of the intervention and its implementation, and ways to address them, were identified and then used to inform the intervention's development. RESULTS: A key barrier to stopping antibiotics was reportedly a lack of information about the original prescriber's rationale for and their degree of certainty about the need for antibiotics. An integral component of ARK was the development and optimization of a Decision Aid and its implementation to increase transparency around initial prescribing decisions. CONCLUSIONS: The key output of this research is a digital and behavioural intervention targeting important barriers to stopping antibiotics at review (see http://bsac-vle.com/ark-the-antibiotic-review-kit/ and http://antibioticreviewkit.org.uk/). ARK will be evaluated in a feasibility study and, if successful, a stepped-wedge cluster-randomized controlled trial at acute hospitals across the NHS.
Assuntos
Antibacterianos/administração & dosagem , Gestão de Antimicrobianos/métodos , Prescrições de Medicamentos/estatística & dados numéricos , Medicina Geral/métodos , Pessoal de Saúde/educação , Antibacterianos/normas , Prescrições de Medicamentos/normas , Medicina Geral/educação , Medicina Geral/normas , Implementação de Plano de Saúde/métodos , Implementação de Plano de Saúde/normas , Hospitais/estatística & dados numéricos , Humanos , Pesquisa Qualitativa , Participação dos Interessados , Reino UnidoRESUMO
BACKGROUND: Extended-spectrum cephalosporin resistance (ESC-R) in Escherichia coli and Klebsiella pneumoniae is a healthcare threat; high gastrointestinal carriage rates are reported from South-east Asia. Colonisation prevalence data in Cambodia are lacking. The aim of this study was to determine gastrointestinal colonisation prevalence of ESC-resistant E. coli (ESC-R-EC) and K. pneumoniae (ESC-R-KP) in Cambodian children/adolescents and associated socio-demographic risk factors; and to characterise relevant resistance genes, their genetic contexts, and the genetic relatedness of ESC-R strains using whole genome sequencing (WGS). RESULTS: Faeces and questionnaire data were obtained from individuals < 16 years in north-western Cambodia, 2012. WGS of cultured ESC-R-EC/KP was performed (Illumina). Maximum likelihood phylogenies were used to characterise relatedness of isolates; ESC-R-associated resistance genes and their genetic contexts were identified from de novo assemblies using BLASTn and automated/manual annotation. 82/148 (55%) of children/adolescents were ESC-R-EC/KP colonised; 12/148 (8%) were co-colonised with both species. Independent risk factors for colonisation were hospitalisation (OR: 3.12, 95% CI [1.52-6.38]) and intestinal parasites (OR: 3.11 [1.29-7.51]); school attendance conferred decreased risk (OR: 0.44 [0.21-0.92]. ESC-R strains were diverse; the commonest ESC-R mechanisms were blaCTX-M 1 and 9 sub-family variants. Structures flanking these genes were highly variable, and for blaCTX-M-15, - 55 and - 27 frequently involved IS26. Chromosomal blaCTX-M integration was common in E. coli. CONCLUSIONS: Gastrointestinal ESC-R-EC/KP colonisation is widespread in Cambodian children/adolescents; hospital admission and intestinal parasites are independent risk factors. The genetic contexts of blaCTX-M are highly mosaic, consistent with rapid horizontal exchange. Chromosomal integration of blaCTX-M may result in stable propagation in these community-associated pathogens.
Assuntos
Portador Sadio/epidemiologia , Cefalosporinas/farmacologia , Farmacorresistência Bacteriana , Infecções por Escherichia coli/epidemiologia , Trato Gastrointestinal/microbiologia , Infecções por Klebsiella/epidemiologia , Adolescente , Antibacterianos/farmacologia , Camboja/epidemiologia , Portador Sadio/microbiologia , Criança , Pré-Escolar , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/patogenicidade , Feminino , Trato Gastrointestinal/parasitologia , Hospitalização , Humanos , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/patogenicidade , Masculino , Doenças Parasitárias/epidemiologia , Doenças Parasitárias/microbiologia , Prevalência , Fatores de Risco , Inquéritos e Questionários , Sequenciamento Completo do GenomaRESUMO
Background and Objectives: Serratia marcescens is an emerging nosocomial pathogen, and the carbapenemase blaNDM has been reported in several surveys in Romania. We aimed to investigate the molecular epidemiology of S. marcescens in two Romanian hospitals over 2010-15, including a neonatal NDM-1 S. marcescens outbreak. Methods: Isolates were sequenced using Illumina technology together with carbapenem-non-susceptible NDM-1-positive and NDM-1-negative Klebsiella pneumoniae and Enterobacter cloacae to provide genomic context. A subset was sequenced with MinION to fully resolve NDM-1 plasmid structures. Resistance genes, plasmid replicons and ISs were identified in silico for all isolates; an annotated phylogeny was reconstructed for S. marcescens. Fully resolved study NDM-1 plasmid sequences were compared with the most closely related publicly available NDM-1 plasmid reference. Results: 44/45 isolates were successfully sequenced (S. marcescens, n = 33; K. pneumoniae, n = 7; E. cloacae, n = 4); 10 with MinION. The S. marcescens phylogeny demonstrated several discrete clusters of NDM-1-positive and -negative isolates. All NDM-1-positive isolates across species harboured a pKOX_NDM1-like plasmid; more detailed comparisons of the plasmid structures demonstrated a number of differences, but highlighted the largely conserved plasmid backbones across species and hospital sites. Conclusions: The molecular epidemiology is most consistent with the importation of a pKOX_NDM1-like plasmid into Romania and its dissemination amongst K. pneumoniae/E. cloacae and subsequently S. marcescens across hospitals. The data suggested multiple acquisitions of this plasmid by S. marcescens in the two hospitals studied; transmission events within centres, including a large outbreak on the Targu Mures neonatal unit; and sharing of the pKOX_NDM1-like plasmid between species within outbreaks.
Assuntos
Genoma Bacteriano , Infecções por Serratia/epidemiologia , Serratia marcescens/genética , beta-Lactamases/genética , DNA Bacteriano/genética , Surtos de Doenças , Enterobacteriaceae/genética , Infecções por Enterobacteriaceae/epidemiologia , Fezes/microbiologia , Transferência Genética Horizontal , Hospitais , Humanos , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , Plasmídeos/genética , Romênia/epidemiologia , Análise de Sequência de DNA , Serratia marcescens/enzimologia , Sequenciamento Completo do Genoma/métodos , beta-Lactamases/biossínteseRESUMO
Whole-genome sequencing (WGS) makes it possible to determine the relatedness of bacterial isolates at a high resolution, thereby helping to characterize outbreaks. However, for Staphylococcus aureus, the accumulation of within-host diversity during carriage might limit the interpretation of sequencing data. In this study, we hypothesized the converse, namely, that within-host diversity can in fact be exploited to reveal the involvement of long-term carriers (LTCs) in outbreaks. We analyzed WGS data from 20 historical outbreaks and applied phylogenetic methods to assess genetic relatedness and to estimate the time to most recent common ancestor (TMRCA). The findings were compared with the routine investigation results and epidemiological evidence. Outbreaks with epidemiological evidence for an LTC source had a mean estimated TMRCA (adjusted for outbreak duration) of 243 days (95% highest posterior density interval [HPD], 143 to 343 days) compared with 55 days (95% HPD, 28 to 81 days) for outbreaks lacking epidemiological evidence for an LTC (P = 0.004). A threshold of 156 days predicted LTC involvement with a sensitivity of 0.875 and a specificity of 1. We also found 6/20 outbreaks included isolates with differing antimicrobial susceptibility profiles; however, these had only modestly increased pairwise diversity (mean 17.5 single nucleotide variants [SNVs] [95% confidence interval {CI}, 17.3 to 17.8]) compared with isolates with identical antibiograms (12.7 SNVs [95% CI, 12.5 to 12.8]) (P < 0.0001). Additionally, for 2 outbreaks, WGS identified 1 or more isolates that were genetically distinct despite having the outbreak pulsed-field gel electrophoresis (PFGE) pulsotype. The duration-adjusted TMRCA allowed the involvement of LTCs in outbreaks to be identified and could be used to decide whether screening for long-term carriage (e.g., in health care workers) is warranted. Requiring identical antibiograms to trigger investigation could miss important contributors to outbreaks.
Assuntos
Portador Sadio/epidemiologia , Surtos de Doenças , Tipagem Molecular , Infecções Estafilocócicas/epidemiologia , Staphylococcus aureus/classificação , Staphylococcus aureus/genética , Sequenciamento Completo do Genoma , Adulto , Portador Sadio/microbiologia , Eletroforese em Gel de Campo Pulsado , Genótipo , Humanos , Testes de Sensibilidade Microbiana , Filogenia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/isolamento & purificaçãoRESUMO
Studies of the transmission epidemiology of antimicrobial-resistant Escherichia coli, such as strains harboring extended-spectrum beta-lactamase (ESBL) genes, frequently use selective culture of rectal surveillance swabs to identify isolates for molecular epidemiological investigation. Typically, only single colonies are evaluated, which risks underestimating species diversity and transmission events. We sequenced the genomes of 16 E. coli colonies from each of eight fecal samples (n = 127 genomes; one failure), taken from different individuals in Cambodia, a region of high ESBL-producing E. coli prevalence. Sequence data were used to characterize both the core chromosomal diversity of E. coli isolates and their resistance/virulence gene content as a proxy measure of accessory genome diversity. The 127 E. coli genomes represented 31 distinct sequence types (STs). Seven (88%) of eight subjects carried ESBL-positive isolates, all containing blaCTX-M variants. Diversity was substantial, with a median of four STs/individual (range, 1 to 10) and wide genetic divergence at the nucleotide level within some STs. In 2/8 (25%) individuals, the same blaCTX-M variant occurred in different clones, and/or different blaCTX-M variants occurred in the same clone. Patterns of other resistance genes and common virulence factors, representing differences in the accessory genome, were also diverse within and between clones. The substantial diversity among intestinally carried ESBL-positive E. coli bacteria suggests that fecal surveillance, particularly if based on single-colony subcultures, will likely underestimate transmission events, especially in high-prevalence settings.
Assuntos
Escherichia coli/classificação , Escherichia coli/enzimologia , Fezes/microbiologia , Variação Genética , beta-Lactamases/metabolismo , Adolescente , Camboja , DNA Bacteriano/química , DNA Bacteriano/genética , Farmacorresistência Bacteriana , Escherichia coli/isolamento & purificação , Feminino , Genes Bacterianos , Genoma Bacteriano , Genótipo , Humanos , Masculino , Análise de Sequência de DNA , Fatores de Virulência/genéticaRESUMO
OBJECTIVES: There are limited data on Enterobacter cloacae outbreaks and fewer describing these in association with NDM-1. With whole-genome sequencing, we tested the hypothesis that a cluster of 16 E. cloacae bacteraemia cases in a Nepali neonatal unit represented a single clonal outbreak, using a wider set of epidemiologically unrelated clinical E. cloacae isolates for comparison. METHODS: Forty-three isolates were analysed, including 23 E. cloacae and 3 Citrobacter sp. isolates obtained from blood cultures from 16 neonates over a 3 month period. These were compared with two contemporaneous community-associated drug-resistant isolates from adults, a unit soap dispenser isolate and a set of historical invasive isolates (n=14) from the same geographical locality. RESULTS: There were two clear neonatal outbreaks and one isolated case in the unit. One outbreak was associated with an NDM-1 plasmid also identified in a historical community-associated strain. The smaller, second outbreak was likely associated with a contaminated soap dispenser. The two community-acquired adult cases and three sets of historical hospital-associated neonatal isolates represented four additional genetic clusters. CONCLUSIONS: E. cloacae infections in this context represent several different transmission networks, operating at the community/hospital and host strain/plasmid levels. Wide sampling frames and high-resolution typing methods are needed to describe the complex molecular epidemiology of E. cloacae outbreaks, which is not appropriately reflected by routine susceptibility phenotypes. Soap dispensers may represent a reservoir for E. cloacae and bacterial strains and plasmids may persist in hospitals and in the community for long periods, sporadically being involved in outbreaks of disease.
Assuntos
Bacteriemia/epidemiologia , Surtos de Doenças , Enterobacter cloacae/isolamento & purificação , Infecções por Enterobacteriaceae/epidemiologia , Adulto , Bacteriemia/microbiologia , Bacteriemia/transmissão , Sangue/microbiologia , Análise por Conglomerados , DNA Bacteriano/química , DNA Bacteriano/genética , Transmissão de Doença Infecciosa , Enterobacter cloacae/classificação , Enterobacter cloacae/genética , Infecções por Enterobacteriaceae/microbiologia , Infecções por Enterobacteriaceae/transmissão , Microbiologia Ambiental , Genoma Bacteriano , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Recém-Nascido , Unidades de Terapia Intensiva Neonatal , Epidemiologia Molecular , Dados de Sequência Molecular , Nepal/epidemiologia , Estudos RetrospectivosRESUMO
NDM-producing Klebsiella pneumoniae strains represent major clinical and infection control challenges, particularly in resource-limited settings with high rates of antimicrobial resistance. Determining whether transmission occurs at a gene, plasmid, or bacterial strain level and within hospital and/or the community has implications for monitoring and controlling spread. Whole-genome sequencing (WGS) is the highest-resolution typing method available for transmission epidemiology. We sequenced carbapenem-resistant K. pneumoniae isolates from 26 individuals involved in several infection case clusters in a Nepali neonatal unit and 68 other clinical Gram-negative isolates from a similar time frame, using Illumina and PacBio technologies. Within-outbreak chromosomal and closed-plasmid structures were generated and used as data set-specific references. Three temporally separated case clusters were caused by a single NDM K. pneumoniae strain with a conserved set of four plasmids, one being a 304,526-bp plasmid carrying bla(NDM-1). The plasmids contained a large number of antimicrobial/heavy metal resistance and plasmid maintenance genes, which may have explained their persistence. No obvious environmental/human reservoir was found. There was no evidence of transmission of outbreak plasmids to other Gram-negative clinical isolates, although bla(NDM) variants were present in other isolates in different genetic contexts. WGS can effectively define complex antimicrobial resistance epidemiology. Wider sampling frames are required to contextualize outbreaks. Infection control may be effective in terminating outbreaks caused by particular strains, even in areas with widespread resistance, although this study could not demonstrate evidence supporting specific interventions. Larger, detailed studies are needed to characterize resistance genes, vectors, and host strains involved in disease, to enable effective intervention.
Assuntos
Cromossomos Bacterianos/química , Infecção Hospitalar/epidemiologia , Doenças Endêmicas , Genoma Bacteriano , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , beta-Lactamases/genética , Antibacterianos/uso terapêutico , Mapeamento Cromossômico , Cromossomos Bacterianos/metabolismo , Infecções Comunitárias Adquiridas/tratamento farmacológico , Infecções Comunitárias Adquiridas/epidemiologia , Infecções Comunitárias Adquiridas/microbiologia , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/microbiologia , Farmacorresistência Bacteriana , Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Hospitais , Humanos , Lactente , Recém-Nascido , Unidades de Terapia Intensiva Neonatal , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/isolamento & purificação , Nepal/epidemiologia , Plasmídeos/química , Plasmídeos/metabolismo , beta-Lactamases/metabolismoRESUMO
Staphylococcus aureus is a commensal that can also cause invasive infection. Reports suggest that nasal cocolonization occurs rarely, but the resources required to sequence multiple colonies have precluded its large-scale investigation. A staged protocol was developed to maximize detection of mixed-spa-type colonization while minimizing laboratory resources using 3,197 S. aureus-positive samples from a longitudinal study of healthy individuals in Oxfordshire, United Kingdom. Initial typing of pooled material from each sample identified a single unambiguous strain in 89.6% of samples. Twelve single-colony isolates were typed from samples producing ambiguous initial results. All samples could be resolved into one or more spa types using the protocol. Cocolonization point prevalence was 3.4 to 5.8% over 24 months of follow-up in 360 recruitment-positives. However, 18% were cocolonized at least once, most only transiently. Cocolonizing spa types were completely unrelated in 56% of samples. Of 272 recruitment-positives returning ≥12 swabs, 166 (61%) carried S. aureus continuously but only 106 (39%) carried the same single spa type without any cocolonization; 31 (11%) switched spa type and 29 (11%) had transient cocarriage. S. aureus colonization is dynamic even in long-term carriers. New unrelated cocolonizing strains could increase invasive disease risk, and ongoing within-host evolution could increase invasive potential, possibilities that future studies should explore.
Assuntos
Portador Sadio/microbiologia , Coinfecção/microbiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/classificação , Staphylococcus aureus/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Portador Sadio/epidemiologia , Coinfecção/epidemiologia , Feminino , Voluntários Saudáveis , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Tipagem Molecular , Prevalência , Infecções Estafilocócicas/epidemiologia , Staphylococcus aureus/genética , Reino Unido , Adulto JovemRESUMO
Whole-genome sequencing (WGS) could potentially provide a single platform for extracting all the information required to predict an organism's phenotype. However, its ability to provide accurate predictions has not yet been demonstrated in large independent studies of specific organisms. In this study, we aimed to develop a genotypic prediction method for antimicrobial susceptibilities. The whole genomes of 501 unrelated Staphylococcus aureus isolates were sequenced, and the assembled genomes were interrogated using BLASTn for a panel of known resistance determinants (chromosomal mutations and genes carried on plasmids). Results were compared with phenotypic susceptibility testing for 12 commonly used antimicrobial agents (penicillin, methicillin, erythromycin, clindamycin, tetracycline, ciprofloxacin, vancomycin, trimethoprim, gentamicin, fusidic acid, rifampin, and mupirocin) performed by the routine clinical laboratory. We investigated discrepancies by repeat susceptibility testing and manual inspection of the sequences and used this information to optimize the resistance determinant panel and BLASTn algorithm. We then tested performance of the optimized tool in an independent validation set of 491 unrelated isolates, with phenotypic results obtained in duplicate by automated broth dilution (BD Phoenix) and disc diffusion. In the validation set, the overall sensitivity and specificity of the genomic prediction method were 0.97 (95% confidence interval [95% CI], 0.95 to 0.98) and 0.99 (95% CI, 0.99 to 1), respectively, compared to standard susceptibility testing methods. The very major error rate was 0.5%, and the major error rate was 0.7%. WGS was as sensitive and specific as routine antimicrobial susceptibility testing methods. WGS is a promising alternative to culture methods for resistance prediction in S. aureus and ultimately other major bacterial pathogens.
Assuntos
Biologia Computacional/métodos , Farmacorresistência Bacteriana , Genoma Bacteriano , Análise de Sequência de DNA/métodos , Staphylococcus aureus/genética , Antibacterianos/farmacologia , Humanos , Sensibilidade e Especificidade , Staphylococcus aureus/efeitos dos fármacosRESUMO
No study to date has compared multilocus variable-number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS) in an investigation of the transmission of Clostridium difficile infection. Isolates from 61 adults with ongoing and/or recurrent C. difficile infections and 17 asymptomatic carriage episodes in children (201 samples), as well as from 61 suspected outbreaks affecting 2 to 41 patients in 31 hospitals in the United Kingdom (300 samples), underwent 7-locus MLVA and WGS in parallel. When the first and last samples from the same individual taken for a median (interquartile range [IQR]) of 63 days (43 to 105 days) apart were compared, the estimated rates of the evolution of single nucleotide variants (SNVs), summed tandem-repeat differences (STRDs), and locus variants (LVs) were 0.79 (95% confidence interval [CI], 0.00 to 1.75), 1.63 (95% CI, 0.00 to 3.59), and 1.21 (95% CI, 0.00 to 2.67)/called genome/year, respectively. Differences of >2 SNVs and >10 STRDs have been used to exclude direct case-to-case transmission. With the first serial sample per individual being used to assess discriminatory power, across all pairs of samples sharing a PCR ribotype, 192/283 (68%) differed by >10 STRDs and 217/283 (77%) by >2 SNVs. Among all pairs of cases from the same suspected outbreak, 1,190/1,488 (80%) pairs had concordant results using >2 SNVs and >10 STRDs to exclude transmission. For the discordant pairs, 229 (15%) had ≥2 SNVs but ≤10 STRDs, and 69 (5%) had ≤2 SNVs but ≥10 STRDs. Discordant pairs had higher numbers of LVs than concordant pairs, supporting the more diverse measure in each type of discordant pair. Conclusions on whether the potential outbreaks were confirmed were concordant in 58/61 (95%) investigations. Overall findings using MLVA and WGS were very similar despite the fact that they analyzed different parts of the bacterial genome. With improvements in WGS technology, it is likely that MLVA locus data will be available from WGS in the near future.
Assuntos
Clostridioides difficile/classificação , Clostridioides difficile/genética , Infecções por Clostridium/epidemiologia , Repetições Minissatélites , Tipagem Molecular/métodos , Análise de Sequência de DNA , Adulto , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/microbiologia , Infecções por Clostridium/transmissão , Surtos de Doenças , Humanos , Lactente , Epidemiologia Molecular/métodos , Reino UnidoRESUMO
OBJECTIVES: Whole-genome sequencing potentially represents a single, rapid and cost-effective approach to defining resistance mechanisms and predicting phenotype, and strain type, for both clinical and epidemiological purposes. This retrospective study aimed to determine the efficacy of whole genome-based antimicrobial resistance prediction in clinical isolates of Escherichia coli and Klebsiella pneumoniae. METHODS: Seventy-four E. coli and 69 K. pneumoniae bacteraemia isolates from Oxfordshire, UK, were sequenced (Illumina HiSeq 2000). Resistance phenotypes were predicted from genomic sequences using BLASTn-based comparisons of de novo-assembled contigs with a study database of >100 known resistance-associated loci, including plasmid-associated and chromosomal genes. Predictions were made for seven commonly used antimicrobials: amoxicillin, co-amoxiclav, ceftriaxone, ceftazidime, ciprofloxacin, gentamicin and meropenem. Comparisons were made with phenotypic results obtained in duplicate by broth dilution (BD Phoenix). Discrepancies, either between duplicate BD Phoenix results or between genotype and phenotype, were resolved with gradient diffusion analyses. RESULTS: A wide variety of antimicrobial resistance genes were identified, including blaCTX-M, blaLEN, blaOKP, blaOXA, blaSHV, blaTEM, aac(3')-Ia, aac-(3')-IId, aac-(3')-IIe, aac(6')-Ib-cr, aadA1a, aadA4, aadA5, aadA16, aph(6')-Id, aph(3')-Ia, qnrB and qnrS, as well as resistance-associated mutations in chromosomal gyrA and parC genes. The sensitivity of genome-based resistance prediction across all antibiotics for both species was 0.96 (95% CI: 0.94-0.98) and the specificity was 0.97 (95% CI: 0.95-0.98). Very major and major error rates were 1.2% and 2.1%, respectively. CONCLUSIONS: Our method was as sensitive and specific as routinely deployed phenotypic methods. Validation against larger datasets and formal assessments of cost and turnaround time in a routine laboratory setting are warranted.
Assuntos
DNA Bacteriano/genética , Farmacorresistência Bacteriana , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Genoma Bacteriano , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/genética , Antibacterianos/farmacologia , Bacteriemia/microbiologia , Escherichia coli/isolamento & purificação , Humanos , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade Microbiana/métodos , Sensibilidade e Especificidade , Análise de Sequência de DNA , Reino UnidoRESUMO
In many Asian populations, the commonest form of severe thalassemia results from the coinheritance of HbE and beta thalassemia. The management of this disease is particularly difficult because of its extreme clinical diversity; although some genetic and adaptive factors have been identified as phenotypic modifiers, the reasons remain unclear. Because the role of the environment in the course of severe thalassemia has been neglected completely and because malaria due to both Plasmodium falciparum and Plasmodium vivax has been prevalent in Sri Lanka, we carried out a pilot study of patients with HbE beta thalassemia that showed high frequencies of antibodies to both parasite species and that 28.6% of the children had DNA-based evidence of current infection with P. vivax. Malarial antibodies then were assessed in patients with HbE beta thalassemia compared with those in age-matched controls. There was a significant increase in the frequency of antibodies in the thalassemic patients, particularly against P. vivax and in young children. There was also a higher frequency in those who had been splenectomized compared with those with intact spleens, although in the latter it was still higher than that in the controls. The thalassemic patients showed significant correlations between malaria antibody status and phenotype. Patients with HbE beta thalassemia may be more prone to malaria, particularly P. vivax, which is reflected in their clinical severity. Because P. vivax malaria is widespread in Asia, further studies of its interaction with HbE beta thalassemia and related diseases are required urgently as a part of ongoing thalassemia control programs.
Assuntos
Povo Asiático , Malária/complicações , Talassemia beta/complicações , Talassemia beta/patologia , Adolescente , Adulto , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Estudos de Casos e Controles , Criança , Exposição Ambiental , Humanos , Malária/epidemiologia , Malária/imunologia , Fenótipo , Projetos Piloto , Prevalência , Esplenectomia , Sri Lanka/epidemiologia , Talassemia beta/imunologiaRESUMO
BACKGROUND: Antimicrobial stewardship initiatives in secondary care depend on clinicians undertaking antibiotic prescription reviews but decisions to limit antibiotic treatment at review are complex. AIM: To assess the feasibility and acceptability of implementing ARK (Antibiotic Review Kit), a behaviour change intervention made up of four components (brief online tool, prescribing decision aid, regular data collection and feedback process, and patient leaflet) to support stopping antibiotic treatment when it is safe to do so among hospitalized patients; before definitive evaluation through a stepped-wedge cluster-randomized controlled trial. METHODS: Acceptability of the different intervention elements was assessed for a period of 12 weeks by uptake of the online tool, adoption of the decision aid into prescribing practice, and rates of decisions to stop antibiotics at review (assessed through repeated point-prevalence surveys). Patient perceptions of the information leaflet were assessed through a brief questionnaire. FINDINGS: All elements of the intervention were successfully introduced into practice. A total of 132 staff encompassing a broad range of prescribers and non-prescribers completed the online tool (19.4 per 100 acute beds), including 97% (32/33) of the pre-specified essential clinical staff. Among 588 prescription charts evaluated in seven point-prevalence surveys over the 12-week implementation period, 82% overall (76-90% at each survey) used the decision aid. The median antibiotic stop rate post implementation was 36% (range: 29-40% at each survey) compared with 9% pre implementation (P < 0.001). CONCLUSION: ARK provides a feasible and acceptable mechanism to support stopping antibiotics safely at post-prescription reviews in an acute hospital setting.
Assuntos
Antibacterianos/uso terapêutico , Gestão de Antimicrobianos/métodos , Terapia Comportamental/métodos , Aceitação pelo Paciente de Cuidados de Saúde , Atitude do Pessoal de Saúde , Estudos de Viabilidade , Hospitais , HumanosRESUMO
Despite the importance of healthcare-associated infection, few studies have quantified the association between severe infection and hospital exposure in UK populations. Our aim was to estimate the proportion of the population with recent hospital admission, together with rates of infection in hospital-exposed and hospital-naïve populations. We studied bacteraemia as a marker of severe infection in a population of 550,000, served by two hospitals, between 1 April 2000 and 31 March 2005. Hospital-exposed persons accounted for 8.3% of the population, defined as having been resident in a hospital in the last year. The hospital-exposed population accounted for 55% of all admissions, and 42% of emergency admissions to medical, paediatric or surgery departments. After adjustment for age, the hospital-exposed group had much higher rates of admission bacteraemia. Age-standardised incidence rate ratios relative to hospital-naïve patients were 43 [95% confidence interval (CI): 22-85] for meticillin-resistant Staphylococcus aureus (MRSA), 20 (15-27) for S. aureus other than MRSA, 7.3 (5.2-10) for Streptococcus pneumoniae, and 14 (11-18) for E. coli. MRSA was common among hospital-exposed admissions, including emergencies in hospital-exposed men, rates of admission MRSA bacteraemia (31 per 100,000 per annum) and S. pneumoniae bacteraemia (33 per 100,000 per annum) were similar. This quantitative analysis confirms that prior hospital admission is a major risk factor for bacteraemia on hospital admission; it is unclear whether acquisition of pathogens in hospital, co-morbidity or other factors explain this.
Assuntos
Bacteriemia/epidemiologia , Infecção Hospitalar/epidemiologia , Infecções por Escherichia coli/epidemiologia , Infecções Pneumocócicas/epidemiologia , Infecções Estafilocócicas/epidemiologia , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Infecções Comunitárias Adquiridas/epidemiologia , Feminino , Hospitalização/estatística & dados numéricos , Humanos , Incidência , Lactente , Recém-Nascido , Masculino , Resistência a Meticilina , Pessoa de Meia-Idade , Reino Unido/epidemiologiaRESUMO
Whole genome sequencing (WGS) can help to relate Mycobacterium tuberculosis genomes to one another to assess genetic relatedness and infer the likelihood of transmission between cases. The same sequence data are now increasingly being used to predict drug resistance and susceptibility. Controlling the spread of tuberculosis and providing patients with the correct treatment are central to the World Health Organization's target to 'End TB' by 2035, for which the global prevalence of drug-resistant tuberculosis remains one of the main obstacles to success. So far, WGS has been applied largely to drug-susceptible strains for the purposes of understanding transmission, leaving a number of analytical considerations before transferring what has been learnt from drug-susceptible disease to drug-resistant tuberculosis. We discuss these potential problems here, alongside some of the challenges to characterizing the Mycobacterium tuberculosis 'resistome'-the optimal knowledge-base required for WGS-based assays to successfully direct individualized treatment regimens through the prediction of drug resistance and susceptibility in the future.
Assuntos
Técnicas de Tipagem Bacteriana/métodos , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/efeitos dos fármacos , Análise de Sequência de DNA/métodos , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Genoma Bacteriano , Genótipo , Humanos , Epidemiologia Molecular/métodos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificaçãoRESUMO
BACKGROUND: Carriage of Staphylococcus aureus is a risk for infections. Targeted decolonization reduces postoperative infections but depends on accurate screening. AIM: To compare detection of S. aureus carriage in healthy individuals between anatomical sites and nurse- versus self-swabbing; also to determine whether a single nasal swab predicted carriage over four weeks. METHODS: Healthy individuals were recruited via general practices. After consent, nurses performed multi-site swabbing (nose, throat, and axilla). Participants performed nasal swabbing twice-weekly for four weeks. Swabs were returned by mail and cultured for S. aureus. All S. aureus isolates underwent spa typing. Persistent carriage in individuals returning more than three self-swabs was defined as culture of S. aureus from all or all but one self-swabs. FINDINGS: In all, 102 individuals underwent multi-site swabbing; S. aureus carriage was detected from at least one site from 40 individuals (39%). There was no difference between nose (29/102, 28%) and throat (28/102, 27%) isolation rates: the combination increased total detection rate by 10%. Ninety-nine patients returned any self-swab, and 96 returned more than three. Nasal carriage detection was not significantly different on nurse or self-swab [28/99 (74%) vs 26/99 (72%); χ2: P=0.75]. Twenty-two out of 25 participants with first self-swab positive were persistent carriers and 69/71 with first self-swab negative were not, giving high positive predictive value (88%), and very high negative predictive value (97%). CONCLUSION: Nasal swabs detected the majority of carriage; throat swabs increased detection by 10%. Self-taken nasal swabs were equivalent to nurse-taken swabs and predicted persistent nasal carriage over four weeks.
Assuntos
Portador Sadio/diagnóstico , Manejo de Espécimes/métodos , Infecções Estafilocócicas/diagnóstico , Staphylococcus aureus/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Tipagem Molecular , Staphylococcus aureus/classificação , Staphylococcus aureus/genética , Adulto JovemRESUMO
The dissemination of carbapenem resistance in Escherichia coli has major implications for the management of common infections. bla KPC, encoding a transmissible carbapenemase (KPC), has historically largely been associated with Klebsiella pneumoniae, a predominant plasmid (pKpQIL), and a specific transposable element (Tn4401, ~10 kb). Here we characterize the genetic features of bla KPC emergence in global E. coli, 2008-2013, using both long- and short-read whole-genome sequencing. Amongst 43/45 successfully sequenced bla KPC-E. coli strains, we identified substantial strain diversity (n = 21 sequence types, 18% of annotated genes in the core genome); substantial plasmid diversity (≥9 replicon types); and substantial bla KPC-associated, mobile genetic element (MGE) diversity (50% not within complete Tn4401 elements). We also found evidence of inter-species, regional and international plasmid spread. In several cases bla KPC was found on high copy number, small Col-like plasmids, previously associated with horizontal transmission of resistance genes in the absence of antimicrobial selection pressures. E. coli is a common human pathogen, but also a commensal in multiple environmental and animal reservoirs, and easily transmissible. The association of bla KPC with a range of MGEs previously linked to the successful spread of widely endemic resistance mechanisms (e.g. bla TEM, bla CTX-M) suggests that it may become similarly prevalent.
Assuntos
Proteínas de Bactérias/genética , Escherichia coli/metabolismo , Klebsiella pneumoniae/enzimologia , beta-Lactamases/genética , Sequência de Bases , Farmacorresistência Bacteriana Múltipla/genética , Dosagem de Genes , Filogenia , Plasmídeos/metabolismo , Replicon/genéticaRESUMO
Haemoglobin E beta thalassaemia is the commonest form of severe thalassaemia in many Asian countries, but little is known about its natural history, the reasons for clinical diversity, or its management. We studied 109 Sri Lankan patients with the disorder over 5 years. 25 patients were not receiving transfusion; transfusion was stopped with no deleterious effect in a further 37. We identified several genetic and environmental factors that might contribute to the phenotypic diversity of the disorder, including modifiers of haemoglobin F production, malaria, and age-related changes in adaptive function. Our findings suggest that haemoglobin E beta thalassaemia can be managed without transfusion in many patients, even with low haemoglobin levels. Age-related changes in the pattern of adaptation to anaemia suggest that different and more cost-effective approaches to management should be explored.