Noroviruses - SAVALnet PY
Transcripción
Noroviruses - SAVALnet PY
FAMILY CALICIVIRIDAE: Noroviruses Dra. Sonia Montenegro Departamento de Pediatría Hospital Regional GGB 5 de octubre, 2008 Temas a considerar: Taxonomía, estructure y genoma Enfermedad y sintomas clínicos Diagnóstico Transmisión Epidemiología e Incidencia Resistencia Ambiental Prevención y Control (Infección Nosocomial) Taxonomía de Norovirus Previamente- Familia Caliciviridae Genero:1. Norwalk-Like Viruses (NLVs) 2. Sapporo-Like Viruses (SLVs) Recientemente- Género Noroviruses (antes Norwalk-Like Viruses) Morfología Viral Sin envoltura,forma icosahedrica 27-40 nm Estruct. única proteína capsid, 60kD - 180 moléculas, formada de 90 dímeros Características Especiales : 32 depresiones como conos en los ejes del icosahedro Estructura de Norovirus por M.E. www.pubmed.gov PMID: 13679618 Genoma (+) ss RNA, 7900 nt de longitud 3 Open Reading Frames: ORF1= proteins no-estructurales - RdRp+ helicase ORF2= proteina de capsid estructural ORF3= proteína peq., función desc. ORF1 5’ RdRp hel ORF2 cap ORF3 ? 3’ RdRp Structure Estructura cristal. de www.pubmed.gov PMID: 12706072 Estrategia de Replicacion Replicación Típica de virus RNA positive sense ss Wagner et al. Basic Virology. www.netlibrary.com Manifestaciones Enfermedad Infecta intestino delgado y causa gastroenteritis Expansión de los vellosidades del ID proximal y acortamiento microvellosidad. - celulas epiteliales permanecen intactas Periodo Incubación : 24-48 hours Conocido como- “stomach flu” Via de Infección Wagner et al. Basic Virology. www.netlibrary.com Sintomatología Clínica Diarrea No-sanguinea. Nausea Vómitos Cólicos Abdominal. Malestar Myalgias Dolor de cabeza Fiebre baja Síntomas duran 12-60 horas. Niños tienden a sufrir de vómitos, mientras que los adultos tienden a sufrir de diarrea. Transmisión Fecal-Oral Típicamente por agua de beber contaminada Muchas fuentes encontradas : suministro de agua municipal de mala calidad pozos lagos recreacionales piscinas agua de cruceros alimento manipulado por personas infectadas o lavada con agua contaminada Prevalencia de Fuentes de Transmission Characteristics of Noroviruses Feature Observation Consequences Low infectious dose <102 virus particles Droplet/person-to-person, foodhandler spread, second. spread Prolonged asymptomatic shedding Environmental stability Up to 2 weeks Increases risk of secondary spread, problems with control of food handlers Survives 10 ppm chlorine, freezing, 67°C Hard to eliminate from contaminated water; survives in ice, partially cooked oysters Great strain diversity Many genetic and Composite diagnostics; repeated infections with many different antigenic types, underestimate prevalence antigenic types Lack of longtime immunity Disease with reinfection Childhood exposure doesn’t protect in adulthood; difficult to develop vaccine with lifelong protection Reservoir Humans Only in human host— identificat. of related animal strains Clinical Characteristics • Excretion of virus in stool begins a few hours before the onset of symptoms and a maximum of 24-72 hours after exposure • Symptoms usually last 24-60 h; 30% of infections may be asymptomatic • Virus can be present in stool of infected persons for a week or more after recovery or even if never sick Norovirus Susceptibility and Resistance Susceptibility to norovirus is related to histo-blood group antigens (HBGAs) status: • NV attach to carbohydrates of the histo-blood group family on human gastroduodenal epithelial cells (and saliva) if present • Affinity may vary by viral genotype • Human milk contains decoy receptors • Oysters contain similar receptors Factors Associated with Secondary AGE Among Household Contacts • Age of Contact; transmission inversely proportional to age of contact (<2yrs 4 times more likely to transmit infection) • Bed sharing • Exposure to primary vomiting episode • Member of index family Perry et al. EID Vol. 11, No. 7, July 2005 Epidemiología e Incidencia Norovirus hay en todo el mundo. Carga: - aprox. 267,000,000 casos anuales - 612,000 hospitalizaciones - 3,000 muertes Altamnete contagioso: menos de 100 partículas virales pueden causar infeccion. El 2002- un barco cricero para Alaska, brote con 13% de los 1266 a bordo afectados; agua de beber contaminated Outbreaks of Gastroenteritis Associated with Noroviruses on Cruise Ships --- United States, 2002 During January 1--December 2, 2002, CDC's Vessel Sanitation Program (VSP), which conducts surveillance for acute gastroenteritis (AGE) on cruise ships with foreign itineraries sailing into U.S. ports (1), received reports of 21 outbreaks of AGE* on 17 cruise ships. Of the 21 outbreaks, nine were confirmed by laboratory analysis of stool specimens from affected persons to be associated with noroviruses, three were attributable to bacterial agents, and nine were of unknown etiology. Seven outbreaks were reported in 2001, and of these, four were confirmed to be associated with norovirus (CDC, unpublished data, 2002). This report describes five of the norovirus outbreaks that occurred during July 1--December 2, 2002, on cruise ships. EH Cramer, D Forney, Vessel Sanitation Program; AL Dannenberg, MD, Div of Emergency and Environ.Health Svcs, National Center for Environmental Health; MA Widdowson, et al Div of Viral and Rickettsial Diseases; E Mintz, C Stover,, Div of Bacterial Diseases, Nat Cent. of Infectious Diseas; E Isakbaeva, et al MD, CDC. Outbreaks of Gastroenteritis Associated with Noroviruses on Cruise Ships --- United States, 2002 • • • • • • • • Cruise Ship C. On September 28, cruise ship C, owned by cruise line B, embarked 1,984 passengers and 941 crew members for a 7-day round-trip cruise from Florida to the Caribbean. Several passengers had AGE within 24 hours of embarkation, and by October 1, a total of 70 (4%) passengers and two (0.2%) crew members reported illness. On October 3, CDC investigators boarded the ship to conduct an epidemiologic and environmental investigation. Questionnaires completed by 1,879 (95%) passengers and 860 (91%) crew members identified 356 (19%) passengers and 13 (1.5%) crew members who met the AGE case definition. The epidemiologic investigation suggested a point source of infection, followed by cases associated with person-to-person transmission. The investigation identified an association between illness among passengers and lunch served at embarkation (odds ratio=2.4; 95% confidence interval=1.1--5.2; p value=0.02). Four of 11 stool specimens from patients were positive for norovirus by RT-PCR. Characterization of the strain by sequence analysis of RT-PCR products matched those from an outbreak on the same ship that occurred 3 weeks previously but was not identical to the outbreak strain on cruise ships A and B. CDC recommended reinforcing sanitation practices and excluding ill foodhandlers from the work place. Cruise ship C continued service, and no new cases were reported on subsequent cruises. Outbreaks of Gastroenteritis Associated with Noroviruses on Cruise Ships --- United States, 2002 Cruise Ship A. On July 18,2002 cruise ship A, owned by cruise line A, embarked 1,318 passengers and 564 crew members for a 7-day cruise from Vancouver to Alaska. On July 19, five passengers reported to the ship's infirmary with symptoms of Acute Gastro Eenteritis AGE. By July 25, a total of 167 (13%) passengers and nine (2%) crew members had reported illness. Among the 176 patients, the predominant symptoms were vomiting (76%) and diarrhea (73%). Five of 10 stool specimens from ill passengers were positive for norovirus by reverse transcriptase polymerase chain reaction (RT-PCR). On July 25, when passengers disembarked, the ship was disinfected in accordance with CDC recommendations, and the same day, a new group of passengers embarked for another 7-day cruise. During the cruise, 189 (14%) of 1,336 passengers and 30 (5.3%) of 571 crew members had AGE with diarrhea (91%) and vomiting (85%) . An environmental health inspection conducted by CDC revealed no sanitary deficiencies. Cruise line A cancelled a subsequent cruise and voluntarily took the ship out of service for 1 week for aggressive cleaning and sanitizing. No outbreaks were reported on subsequent cruises. Outbreaks of Gastroenteritis Associated with Noroviruses on Cruise Ships --- United States, 2002 Cruise Ship B. On October 1, cruise ship B, also owned by cruise line A, embarked 1,281 passengers and 598 crew members for a 21-day cruise from Washington to Florida. By October 16, a total of 101 (8%) passengers and 14 (2%) crew members reported to the infirmary with AGE symptoms. On October 18, CDC investigators boarded the ship to conduct an epidemiologic and environmental investigation. Of 972 surveyed passengers, 399 (41%) met the case definition for AGE. Investigators found no association between illness and water, specific meals served on the ship, or with offshore excursions. Stool specimens from 12 of 13 patients tested positive for norovirus. Characterization of the strain by sequence analysis of RT-PCR products matched those from cruise ship A. Despite implementation of control measures that included disinfection of the vessel and quarantine of ill passengers and crew members, a total of 264 passengers and 41 crew members reported illness on three subsequent 10-day cruises. Cruise line A voluntarily withdrew cruise ship B from service for 10 days for aggressive cleaning and sanitizing. No outbreaks were reported on subsequent voyages. Prevención y Control Altamente estable en el ambienteresistente a: CONGELAMIENTO CALENTAMIENTO A 60ºC DESINFECCIÓN C/ CLORO CONDICIONES DE ACIDEZ VINAGRE ALCOHOL ALTA CONCENTRACIÓN DE AZÚCAR Prevencion y Control Infección produce IgG, IgA and IgM, pero los anticuerpos no son protectores. No produce inmunidad duradera ni protección a la reinfección. Inmunidad sólo transitoria que dura 3-4 meses. No hay vacuna y no es probable su desarrollo. No hay drogas antivirales,sin embargo recuperacion completa es común. Prevenir con lavado de manos,buena higiene, manejo adecuado del agua y de la preparación de alimentos. DEFINITION “Nosocomial infection is a localized or systemic condition that results from adverse reaction to the presence of an infectious agent(s) or its toxin(s) and that was not present or incubating at the time of admission to the hospital” CDC definitionns of nosocomial infections. APIC Infection Control and Applied Epidemiology: Priciples and Practice. 1996:A-1-20 Norovirus en Chile Vidal R. Et al 2006.Novel Recombinant Norovirus Causing Outbreaks of Gastroenteritis in Santiago, Chile. -The purpose of this study was to characterize genetically the capsid of the NV strains detected in gastroenteritis outbreaks -Outbreaks of gastroenteritis occurring in Santiago, between 2001 and 2003 were caused by genetically diverse NV strains that grouped mainly into GII. Norovirus en Chile Vidal, R., et al 2005 V. 2005. Caliciviruses and foodborne gastroenteritis, Chile. Emerg. Infect. Dis. 11:1134–37. The purpose of this study was to determine the t role of HuCVs as a cause of gastroenteritis outbreaks from June 1, 2000, to January 30, 2003, in Santiago, Chile, by using recently improved antigen and genome detection assays, and to characterize genetically the circulating strains. PREVALENCIA Y DIVERSIDAD GENETICA DE NOROVIRUS AISLADOS DE NIÑOS CON GASTROENTERITIS AGUDA EN CONCEPCION PROYECTO DIUC N0 207.86207.86-0606-1 Metodología Trujillo et al. 2006. J Clin Microbiol 44:1405-12. Use of TaqMan real-time reverse transcription-PCR for rapid detection, quantification, and typing of norovirus. OBJETIVOS 1. Determinar prevalencia de Norovirus por diagnsótico molecular en deposiciones de niños con gastroenteritis aguda. 2. Clasificación genética de aislados de Norovirus por secuenciamiento. 3. Determinar prevalencia de Rotavirus, Adenovirus bacterias y C jejuni spp diagnóstico convenc. 4. Caracterización clínica y epidemiológica de los casos por Norovirus en niños SDA MÉTODOS Estudio caso-control Dic 2007 a Sept. 2008 126 niños de 0-14 años con sospecha de SDA. 58 niños asintomáticos de 0-3 año (guardería) Análisis de deposiciones por RT-PCR “onestep” a tiempo real, con sondas Taqman especificas para detección y tipificación de Norovirus Genogrupos I y II. Presencia de Rotavirus y Adenovirus en heces por inmunocromatografía dual kit VIKIA™ “Rota-Adeno” de Biomerieux , realizado en el lab. de Microbiología del Hospital Regional GGB Datos del Paciente Datos del Paciente Ficha inicial Datos del Paciente Ficha inicial Datos del Paciente Ficha inicial Datos del Paciente Ficha inicial METODOS SECUENCIAS DE PRIMERS Y SONDAS Primers Norovirus Genotipo I: COG 1R : COG 1F : RING 1A : RING 1B : Tamaño de banda: 84 bp 5' - CTT AGA CGC CAT CAT TYA - 3 5' - CGY TGG ATG CGI TTY CAT GA - 3’ 5' - AGA TYG CGA TCY CCT GTC CA - 3’ 5' - AGA TCG CCG TCT CCT GTC CA - 3 Primers Norovirus Genotipo II: Tamaño de banda: 97 bp COG 2R : 5’ - TCT ACG CCA TCT TCA TTC ACA - 3’ COG 2F : 5’ - CAR GAR BCN ATG TTY AGR TGG ATG AG - 3’ NOV RING 2 : 5’ - TGG GAG GGC GAT CGC AAT CT - 3’ METODOS PROGRAMA TERMOCICLADOR Norovirus RT-PCR RT: Denaturacion : 30 min, 50˚C 10 min, 95 ˚C PCR : (#45) 15 sec, 94˚C 25 sec, 56˚C 25 sec, 72˚C RESULTADOS Norovirus Real-time Taqman PCR C+ 1074 GII C+ 1074 GII C+ KS200 C+ KS200 C+ 1293 GI C+ 1293 GI C+ 1057 GI C+ 1057 GI RESULTADOS Amplification Plots Samples RESULTADOS Norovirus Real-time Taqman PCR 1. AGUA C(T)* 2. M. 1293 G1 38 3. M. 1074 GII 30 RESULTADOS EN 126 NIÑOS: 113 hospitalizados, 13 ambulatorios 23.8 % Norovirus: 19 % G II ; 4.8 % G I Rotavirus 14.4 5.6% de Adenovirus. En los niños controles se encontró 5.2% Norovirus G1 y 1.7% Norovirus GII. RESULTADOS EN 90 NIÑOS DE 0-2 AÑOS 31% Norovirus: 25.6% GII y 5.6% GI. 16.85% Rotavirus y 7.86% Adenovirus Infección Mixta en 3 niños (norovirus GI y GII) NIÑOS HOSPITALIZADOS POR SDA 46% ( de 90) 31.25% Norovirus GII; 21.9% Rota 9.1 %Adeno. NOSOCOMIAL? 54% HOSPITAL. OTRAS CAUSAS 26.3% Norovirus GII, 13.2% Rotavirus y 2.6% Adenovirus. DISTRIBUCION ESTACIONAL VERANO 70 niños 3/70 4.3% OTOÑO 38 niños 0 INVIERNO 17niños 1/17 5.9% Norovirus genotipo II 6/70 8.6% 11/38 28.9% 5/17 29.4% Rotavirus 13/70 18.6% 3/70 4.3% 2/17 11.76% Norovirus genotipo I Adenovirus 3/70 4.3% 4/38 10.5% 0 CONCLUSIONES La mayor prevalencia de Norovirus se encontró en niños de 0-2 años con mayor frecuencia de Norovirus genotipo II. La prevalencia de Norovirus en niños 0-2 años fue superior a la de Rotavirus y es posible que un porcentaje importante sea adquirido como infección intra hospitalaria. Resultados Liu Z. et al 2007 Jul; Infect &Digest Tract Dis . Clinical manifestations of norovirus gastroenteritis in infants and children. RESULTS: 24.8% positive for norovirus antigen . 60.8% October to December, seasonal preference of the virus. 91.2% positives found under 2 years of age. Rotavirus 20.3%, co-infected by these two viruses. Principal diferencia clinica severidad de fiebre en Rotavirus. 18% of Noro positives admitted to hospitals with diagnosis other than gastroenteritis symptoms of diarrhea between 1 to 11 days after hospitalization. Nosocomial infection (NI) Exogenous: microorganisms originated – Hands – Devices – Infusions, injections It can be prevented by high hygiene!!! Endogenous: microorganisms originated – Bowel system – Skin surface – Mounth – Genital system