RESUMEN
Shiga toxin-producing Escherichia coli (STEC) is a significant cause of gastrointestinal infection and the haemolytic-uremic syndrome (HUS). STEC outbreaks are commonly associated with food but animal contact is increasingly being implicated in its transmission. We report an outbreak of STEC affecting young infants at a nursery in a rural community (three HUS cases, one definite case, one probable case, three possible cases and five carriers, based on the combination of clinical, epidemiological and laboratory data) identified using culture-based and molecular techniques. The investigation identified repeated animal contact (animal farming and petting) as a likely source of STEC introduction followed by horizontal transmission. Whole genome sequencing (WGS) was used for real-time investigation of the incident and revealed a unique strain of STEC O26:H11 carrying stx2a and intimin. Following a public health intervention, no additional cases have occurred. This is the first STEC outbreak reported from Israel. WGS proved as a useful tool for rapid laboratory characterization and typing of the outbreak strain and informed the public health response at an early stage of this unusual outbreak.
Asunto(s)
Brotes de Enfermedades , Infecciones por Escherichia coli/epidemiología , Síndrome Hemolítico-Urémico/epidemiología , Escherichia coli Shiga-Toxigénica/fisiología , Infecciones por Escherichia coli/microbiología , Proteínas de Escherichia coli/genética , Genómica , Síndrome Hemolítico-Urémico/microbiología , Humanos , Lactante , Israel/epidemiología , Casas Cuna , Filogenia , Salud Pública , Análisis de Secuencia de ADN , Escherichia coli Shiga-Toxigénica/genéticaRESUMEN
General Practitioner consultation rates for influenza-like illness (ILI) are monitored through several geographically distinct schemes in the UK, providing early warning to government and health services of community circulation and intensity of activity each winter. Following on from the 2009 pandemic, there has been a harmonization initiative to allow comparison across the distinct existing surveillance schemes each season. The moving epidemic method (MEM), proposed by the European Centre for Disease Prevention and Control for standardizing reporting of ILI rates, was piloted in 2011/12 and 2012/13 along with the previously proposed UK method of empirical percentiles. The MEM resulted in thresholds that were lower than traditional thresholds but more appropriate as indicators of the start of influenza virus circulation. The intensity of the influenza season assessed with the MEM was similar to that reported through the percentile approach. The MEM pre-epidemic threshold has now been adopted for reporting by each country of the UK. Further work will continue to assess intensity of activity and apply standardized methods to other influenza-related data sources.
Asunto(s)
Notificación de Enfermedades/métodos , Monitoreo Epidemiológico , Gripe Humana/diagnóstico , Gripe Humana/epidemiología , Atención Primaria de Salud/métodos , Humanos , Reino Unido/epidemiologíaRESUMEN
Sequence-based typing (SBT) for Legionella pneumophila (Lp) has dramatically improved Legionnaires' disease (LD) cluster investigation. Microbial whole genome sequencing (WGS) is a promising modality for investigation but sequence analysis methods are neither standardised, nor agreed. We sought to develop a WGS-based typing scheme for Lp using de novo assembly and a genome-wide gene-by-gene approach (core genome multilocus sequence typing, cgMLST). We analysed 17 publicly available Lp genomes covering the whole species variation to define a core genome (1,521 gene targets) which was validated using 21 additional published genomes. The genomes of 12 Lp strains implicated in three independent cases of paediatric humidifier-associated LD were subject to cgMLST together with three 'outgroup' strains. cgMLST was able to resolve clustered strains and clearly identify related and unrelated strains. Thus, a cgMLST scheme was readily achievable and provided high-resolution analysis of Lp strains. cgMLST appears to have satisfactory discriminatory power for LD cluster analysis and is advantageous over mapping followed by single nucleotide polymorphism (SNP) calling as it is portable and easier to standardise. cgMLST thus has the potential for becoming a gold standard tool for LD investigation. Humidifiers pose an ongoing risk as vehicles for LD and should be considered in cluster investigation and control efforts.
Asunto(s)
Variación Genética , Legionella pneumophila/clasificación , Legionella pneumophila/genética , Tipificación Molecular/métodos , Tipificación de Secuencias Multilocus/métodos , Análisis de Secuencia de ADN , ADN Bacteriano/química , ADN Bacteriano/genética , Genoma Bacteriano , Humanos , Legionella pneumophila/aislamiento & purificación , Epidemiología Molecular/métodos , Datos de Secuencia Molecular , Polimorfismo de Nucleótido SimpleRESUMEN
The optimal method for surveillance of carbapenem-resistant Acinetobacter spp. (CRAB) is unknown. A collection of CRAB strains (n = 42), carbapenem-susceptible strains (CSAB), and non-Acinetobacter strains (n = 18) was used to evaluate six laboratory surveillance methods: MacConkey (MAC), MAC + 1 µg/ml imipenem (MAC-IPM), minimal salts agar + 1 % acetate (MSA), MSA with IPM disk (MSA-IPM), CHROMagarKPC, and CHROMagar Acinetobacter with CR102 (CHROMAcineto). CHROMAcineto was 100 % sensitive and specific. CHROMagarKPC and MAC-IPM were highly sensitive (>95 %), but their specificity was substantially hampered by the breakthrough growth of CSAB. MSA was unsuitable for CRAB detection. CHROMAcineto is a promising medium for CRAB detection and warrants further clinical evaluation.
Asunto(s)
Infecciones por Acinetobacter/diagnóstico , Acinetobacter/aislamiento & purificación , Antibacterianos/farmacología , Técnicas Bacteriológicas/métodos , Carbapenémicos/farmacología , Medios de Cultivo/química , Resistencia betalactámica , Agar , Humanos , Sensibilidad y EspecificidadRESUMEN
In February 2013, wild poliovirus type 1 (WPV1) was reintroduced into southern Israel and resulted in continuous silent circulation in the highly immune population. As a part of the public health emergency response, a novel real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay was developed, to allow for the sensitive and specific detection of the circulatingWPV1-South Asian (SOAS) strain. Specific primers and probes derived from the VP-1 region were designed, based on sequenced sewage isolates, and used to simultaneously amplify this WPV1-SOAS sequence together with bacteriophage MS-2 as internal control. High titre WPV1-SOAS stock virus was used for assay optimisation and 50 processed sewage samples collected from southern Israel and tested by reference culture based methods were used for analytical validation of the assay's performance. The limit of detection of the multiplex qRT-PCR (SOAS/MS-2) assay was 0.1 plaque-forming unit (pfu)/reaction (20 pfu/mL) for WPV1-SOAS RNA with 100% sensitivity, specificity, positive and negative predictive values when compared to the culture based method. The turnaround time was rapid, providing results for environmental samples within 24 to 48 hours from completion of sewage processing, instead of five to seven days by culture-based analysis. Direct sewage testing by qRT-PCR assay proved to be a useful tool for rapid detection and environmental surveillance of WPV1-SOAS circulating strain during emergency response. Application of the approach for detection of WPV1-SOAS in stool samples obtained during acute flaccid paralysis (AFP) surveillance or field surveys should be further evaluated.
Asunto(s)
Poliovirus/genética , Poliovirus/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Enfermedades Transmisibles Emergentes/epidemiología , Enfermedades Transmisibles Emergentes/virología , Cartilla de ADN/genética , ARN Polimerasas Dirigidas por ADN , Heces/virología , Humanos , Israel/epidemiología , Poliomielitis , Poliovirus/clasificación , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Análisis de Secuencia , Aguas del Alcantarillado/virologíaRESUMEN
An emergency response was triggered by recovery of wild poliovirus type 1 (WPV1) of the South Asia (SOAS) lineage from sewage in southern Israel in April 2013 during routine environmental surveillance. Public health risk assessment necessitated intensification of environmental surveillance in order to facilitate countrywide monitoring of WPV1-SOAS circulation. This involved increasing sampling frequency and broadening the geographical area, for better coverage of the population at risk, as well as modifying sewage testing algorithms to accommodate a newly developed WPV1-SOAS-specific quantitative real-time RT-PCR assay for screening of RNA extracted directly from sewage concentrates, in addition to standard virus isolation. Intensified surveillance in 74 sites across Israel between 1 February and 31 August 2013 documented a sustained high viral load of WPV1-SOAS in sewage samples from six Bedouin settlements and two cities with Jewish and Arab populations in the South district. Lower viral loads and intermittent detection were documented in sampling sites representing 14 mixed communities in three of the five health districts in central and northern Israel. Environmental surveillance plays a fundamental role in routine monitoring of WPV circulation in polio-free countries. The rapid assay specific for the circulating strain facilitated implementation of intensified surveillance and informed the public health response and decision-making.
Asunto(s)
Monitoreo del Ambiente , Poliomielitis/epidemiología , Poliovirus/aislamiento & purificación , Aguas del Alcantarillado/virología , Humanos , Israel/epidemiología , Poliomielitis/diagnóstico , Poliomielitis/virología , Poliovirus/genética , Vigilancia de la Población , Salud Pública , Reacción en Cadena en Tiempo Real de la Polimerasa , Medición de RiesgoRESUMEN
Israel has been certified as polio-free by the World Health Organization and its routine immunisation schedule consists of inactivated poliovirus vaccine (IPV) only. At the end of May 2013, the Israeli Ministry of Health (MOH) has confirmed the reintroduction of wild-type poliovirus 1 into the country. Documented ongoing human-to-human transmission necessitated a thorough risk assessment followed by a supplemental immunisation campaign using oral polio vaccine (OPV). The unusual situation in which ongoing poliovirus transmission was picked up through an early warning system of sewage monitoring without active polio cases, brought about significant challenges in risk communication. This paper reviews the challenges faced by the MOH and the communication strategy devised, in order to facilitate and optimise the various components of the public health response, particularly vaccination. Lessons learned from our recent experience may inform risk communication approaches in other countries that may face a similar situation as global polio eradication moves towards the 'End game'.
Asunto(s)
Comunicación , Brotes de Enfermedades/prevención & control , Monitoreo del Ambiente , Poliomielitis/virología , Poliovirus/aislamiento & purificación , Aguas del Alcantarillado/virología , Enfermedades Transmisibles Emergentes/epidemiología , Humanos , Israel/epidemiología , Tamizaje Masivo , Poliomielitis/diagnóstico , Poliomielitis/transmisión , Poliovirus/clasificación , Poliovirus/inmunología , Vacuna Antipolio Oral/inmunología , Vigilancia de la Población , Medición de RiesgoRESUMEN
Poliovirus vaccine coverage in Israel is over 90%. The last nine birth cohorts have been vaccinated exclusively with inactivated polio vaccine (IPV). However, between February and July 2013 type 1 wild poliovirus (WPV1) was detected persistently in 10 and intermittently in 8 of 47 environmental surveillance sites in southern and central Israel and in 30 stool samples collected during July from healthy individuals in southern Israel. We report results of sequence and phylogenetic analyses of genes encoding capsid proteins to determine the source and transmission mode of the virus. WPV1 capsid protein 1 nucleotide sequences were most closely related to South Asia (SOAS) cluster R3A polioviruses circulating in Pakistan in 2012 and isolated from Egyptian sewage in December 2012. There was no noticeable geographical clustering within WPV1-positive sites. Uniform codon usage among isolates from Pakistan, Egypt and Israel showed no signs of optimisation or deoptimisation. Bayesian phylogenetic time clock analysis of the entire capsid coding region (2,643 nt) with a 1.1% evolutionary rate indicated that Israeli and Egyptian WPV1-SOAS lineages diverged in September 2012, while Israeli isolates split into two sub-branches after January 2013. This suggests one or more introduction events into Israel with subsequent silent circulation despite high population immunity.
Asunto(s)
Epidemiología Molecular/métodos , Poliomielitis/epidemiología , Poliomielitis/transmisión , Poliovirus/genética , Poliovirus/aislamiento & purificación , Teorema de Bayes , Monitoreo del Ambiente/métodos , Heces/virología , Humanos , Israel/epidemiología , Cadenas de Markov , Método de Montecarlo , Filogenia , Poliomielitis/diagnóstico , Poliomielitis/virología , Poliovirus/clasificación , Vigilancia de la Población , Análisis de Secuencia , Aguas del Alcantarillado/virologíaRESUMEN
Israel was certified as polio-free country in June 2002, along with the rest of the World Health Organization European Region. Some 11 years later, wild-type polio virus 1 (WPV1) was isolated initially from routine sewage samples collected between 7 and 13 April 2013 in two cities in the Southern district. WPV1-specific analysis of samples indicated WPV1 introduction into that area in early February 2013. National supplementary immunisation with oral polio vaccine has been ongoing since August 2013.
Asunto(s)
Brotes de Enfermedades/prevención & control , Poliomielitis/virología , Poliovirus/aislamiento & purificación , Vigilancia de la Población/métodos , Aguas del Alcantarillado/virología , Enfermedades Transmisibles Emergentes/epidemiología , Humanos , Israel/epidemiología , Vacunación Masiva , Poliomielitis/diagnóstico , Poliovirus/inmunología , Vacuna Antipolio Oral/inmunologíaRESUMEN
Microbiological surveillance for detection of carbapenem-resistant A. baumannii is important, but recovery of A. baumannii is inadequate. We studied A. baumannii recovery by a particular transport system that is possibly superior over standard swabs, using reference and clinical strains. First, the recovery rates relating to the various swabs were compared with regard to various combinations of transport times (0 h, 1 h, 24 h, 48 h), storage times (0 weeks, 1 week, 2 weeks, 4 weeks) and storage temperatures (4°c,-80°c) using live counts. Second, the recovery of different inocula of strains mixed with fecal microbiota was evaluated by plating on selective medium. The new transport system exhibited a decline of <3log10 under almost all conditions studied and performed better than standard swabs under several conditions. If plated on selective media, the new transport system performed well, even after prolonged transport or with a low inoculum, and its processing could be delayed by up to 2 weeks, especially if refrigerated. The new transport system may thus enhance A. baumannii surveillance.
Asunto(s)
Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/aislamiento & purificación , Técnicas Bacteriológicas/métodos , Carbapenémicos/farmacología , Manejo de Especímenes/métodos , Resistencia betalactámica , Antibacterianos/farmacología , HumanosRESUMEN
Although communicable diseases have hitherto played a small part in illness associated with Olympic Games, an outbreak of infection in a national team, Games venue or visiting spectators has the potential to disrupt a global sporting event and distract from the international celebration of athletic excellence. Preparation for hosting the Olympic Games includes implementation of early warning systems for detecting emerging infection problems. Ensuring capability for rapid microbiological diagnoses to inform situational risk assessments underpins the ability to dispel rumours. These are a prelude to control measures to minimize impact of any outbreak of infectious disease at a time of intense public scrutiny. Complex multidisciplinary teamwork combined with laboratory technical innovation and efficient information flows underlie the Health Protection Agency's preparation for the London 2012 Olympic and Paralympic Games. These will deliver durable legacies for clinical and public health microbiology, outbreak investigation and control in the coming years.
Asunto(s)
Control de Enfermedades Transmisibles/métodos , Enfermedades Transmisibles/diagnóstico , Enfermedades Transmitidas por los Alimentos/diagnóstico , Planificación en Salud , Administración en Salud Pública , Deportes , Aniversarios y Eventos Especiales , Enfermedades Transmisibles/microbiología , Diagnóstico Precoz , Gobierno Federal , Enfermedades Transmitidas por los Alimentos/microbiología , Enfermedades Transmitidas por los Alimentos/prevención & control , Organizaciones de Planificación en Salud , Humanos , Centros de Información/organización & administración , Londres , Infecciones del Sistema Respiratorio/diagnóstico , Infecciones del Sistema Respiratorio/prevención & control , Microbiología del AguaRESUMEN
We report a fatal case of community-acquired Legionnaires' disease in an infant aged under six months. Epidemiological and microbiological investigations suggested that a free-standing cold water humidifier using domestic tap water contaminated with Legionella pneumophila serogroup 1 served as a vehicle for infection. These findings were corroborated by sequence-based typing (SBT). Humidifier-associated Legionnaires' disease can be prevented by appropriate control measures. This case also illustrates the emerging role of SBT in the investigation of legionellosis.
Asunto(s)
Diseño de Equipo/efectos adversos , Humedad , Pediatría , Frío/efectos adversos , Contaminación de Equipos , Humanos , Lactante , Israel , Legionella pneumophila/genética , Legionella pneumophila/aislamiento & purificación , Enfermedad de los Legionarios/etiología , Enfermedad de los Legionarios/microbiología , Tipificación Molecular , Microbiología del AguaRESUMEN
During the winter period 2010/11 27 epidemiologically unlinked, confirmed cases of oseltamivir-resistant influenza A(H1N1)2009 virus infection have been detected in multiple, geographically dispersed settings. Three of these cases were in community settings, with no known exposure to oseltamivir. This suggests possible onward transmission of resistant strains and could be an indication of a possibility of changing epidemiology of oseltamivir-resistant influenza A(H1N1)2009 virus.
Asunto(s)
Antivirales/uso terapéutico , Farmacorresistencia Viral , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Gripe Humana/epidemiología , Oseltamivir/uso terapéutico , Adolescente , Adulto , Anciano , Niño , Preescolar , Femenino , Humanos , Lactante , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N1 del Virus de la Influenza A/aislamiento & purificación , Gripe Humana/tratamiento farmacológico , Gripe Humana/transmisión , Gripe Humana/virología , Masculino , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Neuraminidasa/genética , Pandemias , Polimorfismo de Nucleótido Simple , Vigilancia de la Población , Estaciones del Año , Reino Unido/epidemiología , Adulto JovenRESUMEN
OBJECTIVE: Legionella pneumophila serogroup 1 (Lp1) sequence type (ST) 1 is globally widespread in the environment and accounts for a significant proportion of Legionella infections, including nosocomial Legionnaires' disease (LD). This study aimed to design a sensitive and specific detection method for Lp ST1 that will underpin epidemiological investigations and risk assessment. METHODS: A total of 628 Lp genomes (126 ST1s) were analyzed by comparative genomics. Interrogation of more than 900 accessory genes revealed seven candidate targets for specific ST1 detection and specific primers and hydrolysis probes were designed and evaluated. The analytical sensitivity and specificity of the seven primer and probe sets were evaluated on serially diluted DNA extracted from the reference strain CIP107629 and via qPCR applied on 200 characterized isolates. The diagnostic performance of the assay was evaluated on 142 culture-proven clinical samples from LD cases and a real-life investigation of a case cluster. RESULTS: Of seven qPCR assays that underwent analytical validation, one PCR target (lpp1868) showed higher sensitivity and specificity for ST1 and ST1-like strains. The diagnostic performance of the assay using respiratory samples corresponded to a sensitivity of 95% (19/20) (95% CI (75.1-99.9)) and specificity of 100% (122/122) (95% CI (97-100)). The ST1 PCR assay could link two out of three culture-negative hospitalized LD cases to ST1 during a real-time investigation. CONCLUSION: Using whole genome sequencing (WGS) data, we developed and validated a sensitive and specific qPCR assay for the detection of Lp1 belonging to the ST1 clonal complex by amplification of the lpp1868 gene. The ST1 qPCR is expected to deliver an added value for Lp control and prevention, in conjunction with other recently developed molecular assays.
Asunto(s)
Legionella pneumophila/clasificación , Enfermedad de los Legionarios/diagnóstico , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Proteínas Bacterianas/genética , Cartilla de ADN/genética , Sondas de ADN , Genoma Bacteriano , Genómica , Genotipo , Humanos , Legionella pneumophila/aislamiento & purificación , Enfermedad de los Legionarios/microbiología , Tipificación Molecular/métodos , Sensibilidad y Especificidad , Análisis de Secuencia de ADN , Serogrupo , Secuenciación Completa del GenomaRESUMEN
BACKGROUND: Microbial whole genome sequencing (WGS) has many advantages over standard microbiological methods. However, it is not yet widely implemented in routine hospital diagnostics due to notable challenges. OBJECTIVES: The aim was to extract managerial, financial and clinical criteria supporting the decision to implement WGS in routine diagnostic microbiology, across different operational models of implementation in the hospital setting. METHODS: This was a systematic review of literature identified through PubMed and Web of Science. English literature studies discussing the applications of microbial WGS without limitation on publication date were eligible. A narrative approach for categorization and synthesis of the sources identified was adopted. RESULTS: A total of 98 sources were included. Four main alternative operational models for incorporating WGS in clinical microbiology laboratories were identified: full in-house sequencing and analysis, full outsourcing of sequencing and analysis and two hybrid models combining in-house/outsourcing of the sequencing and analysis components. Six main criteria (and multiple related sub-criteria) for WGS implementation emerged from our review and included cost (e.g. the availability of resources for capital and operational investment); manpower (e.g. the ability to provide training programmes or recruit trained personnel), laboratory infrastructure (e.g. the availability of supplies and consumables or sequencing platforms), bioinformatics requirements (e.g. the availability of valid analysis tools); computational infrastructure (e.g. the availability of storage space or data safety arrangements); and quality control (e.g. the existence of standardized procedures). CONCLUSIONS: The decision to incorporate WGS in routine diagnostics involves multiple, sometimes competing, criteria and sub-criteria. Mapping these criteria systematically is an essential stage in developing policies for adoption of this technology, e.g. using a multicriteria decision tool. Future research that will prioritize criteria and sub-criteria that were identified in our review in the context of operational models will inform decision-making at clinical and managerial levels with respect to effective implementation of WGS for routine use. Beyond WGS, similar decision-making challenges are expected with respect to future integration of clinical metagenomics.
Asunto(s)
Enfermedades Transmisibles/diagnóstico , Pruebas Diagnósticas de Rutina/normas , Técnicas Microbiológicas/normas , Secuenciación Completa del Genoma/normas , Técnicas de Apoyo para la Decisión , Pruebas Diagnósticas de Rutina/economía , Pruebas Diagnósticas de Rutina/instrumentación , Humanos , Metagenómica , Técnicas Microbiológicas/economía , Técnicas Microbiológicas/instrumentación , Control de Calidad , Secuenciación Completa del Genoma/economía , Secuenciación Completa del Genoma/instrumentaciónRESUMEN
OBJECTIVES: Shotgun sequencing is increasingly applied in clinical microbiology for unbiased culture-independent diagnosis. While software solutions for metagenomics proliferate, integration of metagenomics in clinical care requires method standardization and validation. Virtual metagenomics samples could underpin validation by substituting real samples and thus we sought to develop a novel solution for simulation of metagenomics samples based on user-defined clinical scenarios. METHODS: We designed the Microbial Metagenomics Mock Scenario-based Sample Simulation (M3S3) workflow, which allows users to generate virtual samples from raw reads or assemblies. The M3S3 output is a mock sample in FASTQ or FASTA format. M3S3 was tested by generating virtual samples for 10 challenging infectious disease scenarios, involving a background matrix 'spiked' in silico with pathogens including mixtures. Replicate samples (seven per scenario) were used to represent different compositional ratios. Virtual samples were analysed using Taxonomer and Kraken db. RESULTS: The 10 challenge scenarios were successfully applied, generating 80 samples. For all tested scenarios, the virtual samples showed sequence compositions as predicted from the user input. Spiked pathogen sequences were identified with the majority of the replicates and most exhibited acceptable abundance (deviation between expected and observed abundance of spiked pathogens), with slight differences observed between software tools. CONCLUSIONS: Despite demonstrated proof-of-concept, integration of clinical metagenomics in routine microbiology remains a substantial challenge. M3S3 is capable of producing virtual samples on-demand, simulating a spectrum of clinical diagnostic scenarios of varying complexity. The M3S3 tool can therefore support the development and validation of standardized metagenomics applications.
Asunto(s)
Enfermedades Transmisibles/diagnóstico , Metagenómica/normas , Técnicas de Diagnóstico Molecular/normas , Simulación por Computador , Humanos , Metagenómica/métodos , Técnicas de Diagnóstico Molecular/métodosRESUMEN
BACKGROUND: Next generation sequencing (NGS) is increasingly being used in clinical microbiology. Like every new technology adopted in microbiology, the integration of NGS into clinical and routine workflows must be carefully managed. AIM: To review the practical aspects of implementing bacterial whole genome sequencing (WGS) in routine diagnostic laboratories. SOURCES: Review of the literature and expert opinion. CONTENT: In this review, we discuss when and how to integrate whole genome sequencing (WGS) in the routine workflow of the clinical laboratory. In addition, as the microbiology laboratories have to adhere to various national and international regulations and criteria for their accreditation, we deliberate on quality control issues for using WGS in microbiology, including the importance of proficiency testing. Furthermore, the current and future place of this technology in the diagnostic hierarchy of microbiology is described as well as the necessity of maintaining backwards compatibility with already established methods. Finally, we speculate on the question of whether WGS can entirely replace routine microbiology in the future and the tension between the fact that most sequencers are designed to process multiple samples in parallel whereas for optimal diagnosis a one-by-one processing of the samples is preferred. Special reference is made to the cost and turnaround time of WGS in diagnostic laboratories. IMPLICATIONS: Further development is required to improve the workflow for WGS, in particular to shorten the turnaround time, reduce costs, and streamline downstream data analyses. Only when these processes reach maturity will reliance on WGS for routine patient management and infection control management become feasible, enabling the transformation of clinical microbiology into a genome-based and personalized diagnostic field.
Asunto(s)
Enfermedades Transmisibles/diagnóstico , Pruebas Diagnósticas de Rutina/métodos , Técnicas Microbiológicas/métodos , Secuenciación Completa del Genoma/métodos , Flujo de Trabajo , HumanosRESUMEN
BACKGROUND: Presently, the bottleneck in the deployment of high-throughput sequencing technology is the ability to analyse the increasing amount of data produced in a fit-for-purpose manner. The field of microbial bioinformatics is thriving and quickly adapting to technological changes, which creates difficulties for nonbioinformaticians in following the complexity and increasingly obscure jargon of this field. AIMS: This review is directed towards nonbioinformaticians who wish to gain understanding of the overall microbial bioinformatic processes, from raw data obtained from sequencers to final outputs. SOURCES: The software and analytical strategies reviewed are based on the personal experience of the authors. CONTENT: The bioinformatic processes of transforming raw reads to actionable information in a clinical and epidemiologic context is explained. We review the advantages and limitations of two major strategies currently applied: read mapping, which is the comparison with a predefined reference genome, and de novo assembly, which is the unguided assembly of the raw data. Finally, we discuss the main analytical methodologies and the most frequently used freely available software and its application in the context of bacterial infectious disease management. IMPLICATIONS: High-throughput sequencing technologies are overhauling outbreak investigation and epidemiologic surveillance while creating new challenges due to the amount and complexity of data generated. The continuously evolving field of microbial bioinformatics is required for stakeholders to fully harness the power of these new technologies.
Asunto(s)
Biología Computacional/métodos , Técnicas Microbiológicas/métodos , Epidemiología Molecular/métodos , Análisis de Secuencia de ADN/métodos , HumanosRESUMEN
OBJECTIVES: Whole genome sequencing (WGS) has revolutionized the subtyping of Legionella pneumophila but calling the traditional sequence-based type from genomic data is hampered by multiple copies of the mompS locus. We propose a novel bioinformatics solution for rectifying that limitation, ensuring the feasibility of WGS for cluster investigation. METHODS: We designed a novel approach based on the alignment of raw reads with a reference sequence. With WGS, reads originating from either of the two mompS copies cannot be differentiated. Therefore, when non-identical copies were present, we applied a read-filtering strategy based on read alignment to a reference sequence via unique 'anchors'. If minimal read coverage was achieved after filtration (≥3X), a consensus sequence was built based on mapped reads followed by calling the sequence-based typing allele. The entire procedure was implemented using a Perl script. RESULTS: The method was validated using a diverse sample of 265 L. pneumophila genomes, consisting of 59 different sequence types (STs) and 23 mompS variants; 57 of the 265 (22%) had non-identical mompS copies. In 237 of the 265 samples (89.4%), mompS calling was successful and no erroneous calling occurred. A 98.1% success was recorded among 109 samples meeting quality requirements. The method was superior to alternative approaches. CONCLUSIONS: As WGS becomes more accessible, technical difficulties in routine clinical and surveillance work will arise. The case of mompS in L. pneumophila serves as an example for such limitations that necessitate the development of novel computational solutions that meet end-user demands.
Asunto(s)
Técnicas de Tipificación Bacteriana , Genes Bacterianos , Genómica , Legionella pneumophila/clasificación , Biología Computacional , ADN Bacteriano/genética , Bases de Datos Genéticas , Estudios de Asociación Genética , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADNRESUMEN
During 2013/14, Israel witnessed the silent reintroduction and sustained transmission of wild poliovirus type 1 (WPV1) detected through routine environmental surveillance performed on sewage samples. The public health response to silent poliovirus transmission in a population with high inactivated polio vaccine (IPV) coverage poses an emerging challenge towards the 'End Game' of global poliovirus eradication. This paper reviews the risk assessment, risk management and risk communication aspects of this poliovirus incident. Special emphasis is placed on the use of scientific data generated in the risk assessment phase to inform the public health response. Reintroducing a live vaccine in supplemental immunization activities in response to transmission of WPV or vaccine-derived poliovirus should be considered close to the 'End Game' of polio eradication, especially if targeting the population at risk is feasible. Such circumstances require a comprehensive contingency plan that will support the generation of important public health evidence at the risk assessment stage, thereby allowing to tailor the risk management approaches and underpin appropriate risk communication.