RESUMEN
Shigella spp. are a leading cause of human diarrheal disease worldwide, with Shigella flexneri being the most frequently isolated species in developing countries. This serogroup is presently classified into 19 serotypes worldwide. We report here a multicenter validation of a multiplex-PCR-based strategy previously developed by Q. Sun, R. Lan, Y. Wang, A. Zhao, et al. (J Clin Microbiol 49:3766-3770, 2011) for molecular serotyping of S. flexneri This study was performed by seven international laboratories, with a panel of 71 strains (researchers were blind to their identity) as well as 279 strains collected from each laboratory's own local culture collections. This collaborative work found a high extent of agreement among laboratories, calculated through interrater reliability (IRR) measures for the PCR test that proved its robustness. Agreement with the traditional method (serology) was also observed in all laboratories for 14 serotypes studied, while specific genetic events could be responsible for the discrepancies among methodologies in the other 5 serotypes, as determined by PCR product sequencing in most of the cases. This work provided an empirical framework that allowed the use of this molecular method to serotype S. flexneri and showed several advantages over the traditional method of serological typing. These advantages included overcoming the problem of availability of suitable antisera in testing laboratories as well as facilitating the analysis of multiple samples at the same time. The method is also less time-consuming for completion and easier to implement in routine laboratories. We recommend that this PCR be adopted, as it is a reliable diagnostic and characterization methodology that can be used globally for laboratory-based shigella surveillance.
Asunto(s)
Reacción en Cadena de la Polimerasa Multiplex/métodos , Serotipificación/métodos , Shigella flexneri/clasificación , Técnicas de Tipificación Bacteriana/métodos , Técnicas de Tipificación Bacteriana/normas , ADN Bacteriano/genética , Humanos , Internacionalidad , Reacción en Cadena de la Polimerasa Multiplex/normas , Serogrupo , Shigella flexneri/inmunologíaAsunto(s)
Antibacterianos/administración & dosificación , Ceftazidima/administración & dosificación , Fosfomicina/administración & dosificación , Infecciones por Serratia/tratamiento farmacológico , Serratia marcescens/enzimología , beta-Lactamasas/metabolismo , Farmacorresistencia Bacteriana , Quimioterapia Combinada , Humanos , Pruebas de Sensibilidad Microbiana , Infecciones por Serratia/microbiología , Serratia marcescens/efectos de los fármacos , Serratia marcescens/aislamiento & purificación , Resultado del Tratamiento , beta-Lactamasas/genéticaAsunto(s)
Infección Hospitalaria/microbiología , Farmacorresistencia Bacteriana Múltiple/genética , Infecciones por Serratia/microbiología , Serratia marcescens/genética , beta-Lactamasas/genética , Argentina/epidemiología , Infección Hospitalaria/epidemiología , Brotes de Enfermedades , Farmacorresistencia Bacteriana Múltiple/efectos de los fármacos , Humanos , Infecciones por Serratia/epidemiología , Serratia marcescens/aislamiento & purificación , Serratia marcescens/patogenicidadRESUMEN
BACKGROUND: To implement effective control measures, timely outbreak detection is essential. Shigella is the most common cause of bacterial diarrhea in Argentina. Highly resistant clones of Shigella have emerged, and outbreaks have been recognized in closed settings and in whole communities. We hereby report our experience with an evolving, integrated, laboratory-based, near real-time surveillance system operating in six contiguous provinces of Argentina during April 2009 to March 2012. METHODOLOGY: To detect localized shigellosis outbreaks timely, we used the prospective space-time permutation scan statistic algorithm of SaTScan, embedded in WHONET software. Twenty three laboratories sent updated Shigella data on a weekly basis to the National Reference Laboratory. Cluster detection analysis was performed at several taxonomic levels: for all Shigella spp., for serotypes within species and for antimicrobial resistance phenotypes within species. Shigella isolates associated with statistically significant signals (clusters in time/space with recurrence interval ≥365 days) were subtyped by pulsed field gel electrophoresis (PFGE) using PulseNet protocols. PRINCIPAL FINDINGS: In three years of active surveillance, our system detected 32 statistically significant events, 26 of them identified before hospital staff was aware of any unexpected increase in the number of Shigella isolates. Twenty-six signals were investigated by PFGE, which confirmed a close relationship among the isolates for 22 events (84.6%). Seven events were investigated epidemiologically, which revealed links among the patients. Seventeen events were found at the resistance profile level. The system detected events of public health importance: infrequent resistance profiles, long-lasting and/or re-emergent clusters and events important for their duration or size, which were reported to local public health authorities. CONCLUSIONS/SIGNIFICANCE: The WHONET-SaTScan system may serve as a model for surveillance and can be applied to other pathogens, implemented by other networks, and scaled up to national and international levels for early detection and control of outbreaks.
Asunto(s)
Brotes de Enfermedades , Disentería Bacilar/epidemiología , Monitoreo Epidemiológico , Shigella/aislamiento & purificación , Argentina/epidemiología , Análisis por Conglomerados , Electroforesis en Gel de Campo Pulsado , Humanos , Pruebas de Sensibilidad Microbiana , Tipificación Molecular , Serotipificación , Shigella/clasificación , Shigella/genéticaRESUMEN
Cronobacter species are opportunistic pathogens associated with severe infections in neonates and immunocompromised infants. From January 2009 through September 2010, two cases of neonatal infections associated with Cronobacter malonaticus and one case associated with Cronobacter sakazakii, two of them fatal, were reported in the same hospital. These are the first clinical isolates of Cronobacter spp. in Argentina. The objective of this work was to characterize and subtype clinical isolates of Cronobacter spp. in neonate patients, as well as to establish the genetic relationship between these isolates and the foodborne isolates previously identified in the country. Pulsed-field gel electrophoresis analysis showed a genetic relationship between the C. malonaticus isolates from two patients. Different results were found when the pulsed-field gel electrophoresis patterns of clinical isolates were compared with those deposited in the National Database of Cronobacter spp.
Asunto(s)
Cronobacter sakazakii/clasificación , Cronobacter sakazakii/aislamiento & purificación , Argentina , Técnicas de Tipificación Bacteriana , Electroforesis en Gel de Campo Pulsado , Femenino , Humanos , Recién Nacido , MasculinoRESUMEN
Cronobacter species are opportunistic pathogens associated with severe infections in neonates and immunocompromised infants. From January 2009 through September 2010, two cases of neonatal infections associated with Cronobacter malonaticus and one case associated with Cronobacter sakazakii, two of them fatal, were reported in the same hospital. These are the first clinical isolates of Cronobacter spp. in Argentina. The objective of this work was to characterize and subtype clinical isolates of Cronobacter spp. in neonate patients, as well as to establish the genetic relationship between these isolates and the foodborne isolates previously identified in the country. Pulsed-field gel electrophoresis analysis showed a genetic relationship between the C. malonaticus isolates from two patients. Different results were found when the pulsed-field gel electrophoresis patterns of clinical isolates were compared with those deposited in the National Database of Cronobacter spp.
Asunto(s)
Cronobacter sakazakii/clasificación , Cronobacter sakazakii/aislamiento & purificación , Argentina , Electroforesis en Gel de Campo Pulsado , Femenino , Humanos , Masculino , Recién Nacido , Técnicas de Tipificación BacterianaRESUMEN
Serratia marcescens causes health care-associated infections with important morbidity and mortality. Particularly, outbreaks produced by multidrug-resistant isolates of this species, which is already naturally resistant to several antibiotics, including colistin, are usually described with high rates of fatal outcomes throughout the world. Thus, it is important to survey factors associated with increasing frequency and/or emergence of multidrug-resistant S. marcescens nosocomial infections. We report the investigation and control of an outbreak with 40% mortality due to multidrug-resistant S. marcescens infections that happened from November 2007 to April 2008 after treatment with colistin for Acinetobacter baumannii meningitis was started at hospital H1 in 2005. Since that year, the epidemiological pattern of frequently recovered species has changed, with an increase of S. marcescens and Proteus mirabilis infections in 2006 in concordance with a significant decrease of the numbers of P. aeruginosa and A. baumannii isolates. A single pulsed-field gel electrophoresis (PFGE) cluster of S. marcescens isolates was identified during the outbreak. When this cluster was compared with S. marcescens strains (n = 21) from 10 other hospitals (1997 to 2010), it was also identified in both sporadic and outbreak isolates circulating in 4 hospitals in Argentina. In132::ISCR1::blaCTX-M-2 was associated with the multidrug-resistant cluster with epidemic behavior when isolated from outbreaks. Standard infection control interventions interrupted transmission of this cluster even when treatment with colistin continued in several wards of hospital H1 until now. Optimizing use of colistin should be achieved simultaneously with improved infection control to prevent the emergence of species naturally resistant to colistin, such as S. marcescens and P. mirabilis.
Asunto(s)
Infecciones por Acinetobacter/tratamiento farmacológico , Antibacterianos/uso terapéutico , Colistina/uso terapéutico , Infección Hospitalaria/epidemiología , Brotes de Enfermedades , Meningitis Bacterianas/tratamiento farmacológico , Infecciones por Serratia/epidemiología , Acinetobacter baumannii/aislamiento & purificación , Adulto , Anciano , Anciano de 80 o más Años , Argentina/epidemiología , Infección Hospitalaria/mortalidad , Farmacorresistencia Bacteriana Múltiple , Electroforesis en Gel de Campo Pulsado , Femenino , Genotipo , Hospitales , Humanos , Masculino , Persona de Mediana Edad , Epidemiología Molecular , Tipificación Molecular , Estudios Retrospectivos , Infecciones por Serratia/mortalidad , Serratia marcescens/clasificación , Serratia marcescens/efectos de los fármacos , Serratia marcescens/genética , Serratia marcescens/aislamiento & purificación , Adulto JovenRESUMEN
Cronobacter species are opportunistic pathogens associated with severe infections in neonates and immunocompromised infants. From January 2009 through September 2010, two cases of neonatal infections associated with Cronobacter malonaticus and one case associated with Cronobacter sakazakii, two of them fatal, were reported in the same hospital. These are the first clinical isolates of Cronobacter spp. in Argentina. The objective of this work was to characterize and subtype clinical isolates of Cronobacter spp. in neonate patients, as well as to establish the genetic relationship between these isolates and the foodborne isolates previously identified in the country. Pulsed-field gel electrophoresis analysis showed a genetic relationship between the C. malonaticus isolates from two patients. Different results were found when the pulsed-field gel electrophoresis patterns of clinical isolates were compared with those deposited in the National Database of Cronobacter spp.
Asunto(s)
Cronobacter sakazakii/clasificación , Cronobacter sakazakii/aislamiento & purificación , Argentina , Técnicas de Tipificación Bacteriana , Electroforesis en Gel de Campo Pulsado , Femenino , Humanos , Recién Nacido , MasculinoRESUMEN
Cronobacter (formerly known as Enterobacter sakazakii) is a genus comprising seven species regarded as opportunistic pathogens that can be found in a wide variety of environments and foods, including powdered infant formula (PIF). Cronobacter sakazakii, the major species of this genus, has been epidemiologically linked to cases of bacteremia, meningitis in neonates, and necrotizing enterocolitis, and contaminated PIF has been identified as an important source of infection. Robust and reproducible subtyping methods are required to aid in the detection and investigation, of foodborne outbreaks. In this study, a pulsed-field gel electrophoresis (PFGE) protocol was developed and validated for subtyping Cronobacter species. It was derived from an existing modified PulseNet protocol, wherein XbaI and SpeI were the primary and secondary restriction enzymes used, generating an average of 14.7 and 20.3 bands, respectively. The PFGE method developed was both reproducible and discriminatory for subtyping Cronobacter species.
Asunto(s)
Cronobacter/clasificación , Tipificación Molecular/métodos , Análisis del Polimorfismo de Longitud de Fragmentos Amplificados , Animales , Cronobacter/genética , Cronobacter/aislamiento & purificación , Cronobacter/metabolismo , Cronobacter sakazakii/clasificación , Cronobacter sakazakii/genética , Cronobacter sakazakii/aislamiento & purificación , Cronobacter sakazakii/metabolismo , Enzimas de Restricción del ADN , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Bacteriano/metabolismo , Electroforesis en Gel de Campo Pulsado , Infecciones por Enterobacteriaceae/diagnóstico , Infecciones por Enterobacteriaceae/microbiología , Alimentos en Conserva/microbiología , Enfermedades Transmitidas por los Alimentos/diagnóstico , Enfermedades Transmitidas por los Alimentos/microbiología , Humanos , Leche/microbiología , Reproducibilidad de los Resultados , Vibrio cholerae/clasificación , Vibrio cholerae/genética , Vibrio cholerae/aislamiento & purificación , Vibrio cholerae/metabolismo , Yersinia pestis/clasificación , Yersinia pestis/genética , Yersinia pestis/aislamiento & purificación , Yersinia pestis/metabolismoRESUMEN
Shigella flexneri is one of the agents most frequently linked to diarrheal illness in developing countries and often causes outbreaks in settings with poor hygiene or sanitary conditions. Travel is one of the means by which S. flexneri can be imported into developed countries, where this pathogen is not commonly seen. A robust and discriminatory subtyping method is needed for the surveillance of S. flexneri locally and regionally, and to aid in the detection and investigation of outbreaks. The PulseNet International network utilizes standardized pulsed-field gel electrophoresis (PFGE) protocols to carry out laboratory-based surveillance of foodborne pathogens in combination with epidemiologic data. A multicenter validation was carried out in nine PulseNet laboratories located in North and South America, Europe, and Asia, and it demonstrated that a new protocol is highly robust and reproducible for subtyping of S. flexneri. This protocol, already approved for PulseNet laboratories, applies NotI and XbaI as primary and secondary restriction enzymes, respectively, under electrophoresis conditions of initial switch time of 5 s to final switch time of 35 s, at 6 volts/cm.