Diagnostics, epidemiology of diarrhoeal diseases of viral origin; vaccination against rotavirus infection Krisztián Bányai ANTSZ Baranya County Institute of State Public Health Service, Regional Laboratory of Virology Historical overview Early 20th century • „pseudocholera infantum”; „winter vomiting disease” 1940s and 1950s • transmissible agent 1950s and1970s • identification of viruses in diarrheic feces 1970s • identification of the first human enteric viruses 1980s and 1990s • identification of new, potential enteric viruses Viruses found in the gut Found in the gut but not associated with gastroenteritis • Polio Enteroviruses 68-71 • Coxsackie A Coxsackie B • Echo • Hepatitis A Hepatitis E • Adenoviruses 1-39, 42-51 Reoviruses Found in the gut as opportunistic infection • CMV VZV • HSV HIV Human enteric viruses Virus Genome Virus morphology Serotypes Rotavirus A,B,C, RNA ~70-75 nm, double-shelled particle, 11 G (Reoviridae, Double-stranded, ‘wheel’-like appearance 12 P Rotavirus) 11 segments Calicivirus (Caliciviridae, RNA ~27-32 nm, featureless surface several Norovirus Positive sense Sapovirus) RNA ~33 nm, David-star surface several Positive sense Astrovirus RNA ~27-30 nm, 5 or 6 pointed star surface 8 (Astroviridae, Positive sense Astrovirus) Adenovirus DNA ~74 nm, classic icosahedral structure 2 (Adenoviridae, Double-stranded Mastadenovirus) Possible human enteric viruses Virus Genome Virus morphology Serotypes Aichi virus RNA ~ 28-35 nm, structured 1 ? (Picornaviridae, Positive sense surface (kobu = knob) Kobuvirus) Parechovirus RNA ~27-32 nm, featureless 3 ? (Picornaviridae, Positive sense surface Parechovirus) Picobirnavirus RNA ~ 28-35 nm, featureless ? Picotrirnavirus Double- surface (Non-classified) stranded, 2 or 3 segments Torovirus RNA ~100-140 nm, torus-like 1 ? (Coronaviridae) Positive sense shape Laboratory diagnostics •Virus detection in the faeces Cultivation Particle detection Antigen detection Nucleic acid detection •Virus-specific antibody in the serum Cultivation Category Viruses Culturable Rotaviruses yes Confirmed Human caliciviruses (noro,sapo) NO Astroviruses yes Enteric adenoviruses (Group F) yes Coronaviruses yes Possible Parechoviruses yes Picobirna-, picotrirnaviruses NO Pestiviruses NO Toroviruses NO Aichi virus yes Not confirmed (Non Group F) Adenoviruses yes Coxsackie A and B viruses yes Echoviruses yes Atmar and Estes, Clin Microbiol Rev 14:15-37, 2001 Electron microscopy Rotavirus Astrovirus Norovirus Sapovirus Parecho Aichi Picobirna- virus Adenovirus Torovirus http://www.epa.gov/nerlcwww/images.htm Antigen-based methods Commercial kits •EIA (RV, AdV, AstV, NV) •IF (RV) •LA (RV, AdV) •ICh (RV, AdV) Nucleic acid based methods RFLP PAGE 4 4 2 3 2 3 2 2 Mixed Identification infections of new viruses No kits (PBV) (RT-)PCR /multiplex/ Sensitivity EM < LA (PAGE) < EIA < (RT-)PCR < Cultivation 106-7 105-6 104-6 101-4 100-1 /ml Limitations: quantity of shed virus particles/antigens (e.g., RV, 1010-1012; NV, < 106-107 ml) epidemiologic background are the detected viruses infectious? Virus Routinely used methods Alternative methods Rotavirus EM PAGE, RT-PCR EIA, LA, ICh Cultivation Serology Cailiciviruses EM Serology (Sapovirus EIA (noro) Norovirus) RT-PCR Astrovirus EM Cultivation EIA Serology RT-PCR Enteric adenovirus EM PCR EIA, LA, ICh RFLP Cultivation Aichi virus Cultivation EIA RT-PCR Parechoviruses Cultivation RT-PCR Picobirnavirus PAGE, RT-PCR Torovirus EM RT-PCR Epidemiology US Bangladesh Episodes/child/year 1-2 4-7 Total episodes by age 5 5-7 20-35 Agents Viruses RV 1 1 AstV 1 1 CVs 1-2 2-5 EAdV 1 1 Bacteria, parasites 1 >20 Risk of death 1 in 12,000 1 in 40 Glass et al., 2001 Endemic childhood disease Epidemic disease Viruses Rotavirus A Norovirus Astrovirus Rotavirus B Enteric adenovirus Norovirus Sapovirus Rotavirus C Astrovirus Torovirus Rotavirus A és C Coronavirus Glass et al., 2001 Kapikian et al., 2001 Differences in the epidemiology of RV Industrial World Developing Countries Seasonality Winter Year round Age (% < 1 yr) 40% 80% Serotypes 5 common Mostly common Some additional Mixed infections Rare Common Case fatality Low High 1 in 100 000 1 in 200 400,000 - 500,000 deaths annually ! Glass et al., 1998 Endemic childhood disease Epidemic disease Viruses Rotavirus A Noroviruses Astrovirus Rotavirus B Enteric adenovirus Norovirus Sapovirus Rotavirus C Astrovirus Torovirus Rotavirus A és C Coronavirus Glass et al., 2001 Etiology of gastroenteritis outbreaks (1998-2002; N=829) Salmonella Shigella Campylobacter Other/More bacteria Non-bacterial Krisztalovics et al., 2004 Etiology of non-bacterial gastroenteritis outbreaks (1998-2002; N=490) Adenovirus Rotavirus Calicivirus Non-bacterial, no further tested Krisztalovics et al., 2004 Total no. of calicivirus (CV) and non-bacterial gastroenteritis outbreaks - laboratory surveillance (1998-2002) CV Non-bacterial Krisztalovics et al., 2004 Settings 9% 11% Kórházak,Hospitals, nursing szoc. homesotthonok 43% Éttermek,Restaurants, fogadások catered meals 11% Iskolák,Schools, dayóvodák care centers VakációVacation settings EgyébOther 26% CDC, 1996-1997 Glass et al., 2000 Source of infection 21% Élelmiszer-közvetítettFood Személyes kontaktus 43% Infected person KagylófogyasztásShellfish 11% VízWater közvetített IsmeretlenUnkown 6% NincsNo data adat 3% 16% CDC, 1996-1997 Glass et al., 2000 Endemic childhood disease Epidemic disease Mode of transmission direct contact food aerosol water contaminated surfaces personal contact aerosol Reservoir humans humans animals animals Immunity high seroprevalence seroconversion by age of 5 y during the outbreak good short-lived Glass et al., 2001 Transmission through ‘aerosol’ Moreno-Espinosa et al., 2004 Endemic childhood disease Epidemic disease Mode of transmission direct contact food aerosol water contaminated surfaces personal contact aerosol Reservoir humans humans animals animals Immunity high seroprevalence seroconversion by age of 5 y during the outbreak good short-lived Glass et al., 2001 Animals as reservoirs of rotavirus strains B4106 (P[14]G3) Belgium (11 LA) AU-1, Ro-5829 (P3[9]G3) 116E (P8[11]G9) Japan, Israel (11 FE) India (10 HU / 1 BO) I321 (P8[11]G10 India (2 HU / 9 BO) Ro1845, HCR3 (P5[3]G3 ) Israel, USA (11 FE / CA) PA151 (P3[9]G6) PCP5 (P3[9]G3) (7 HU / 4 BO) Br1054 (P[8]G5) US1205 (P[8]G9, P[6]G9) S-Am (8 HU / 3 PO) Global (10 HU / 1 PO) Endemic childhood disease Epidemic disease Mode of transmission direct contact food aerosol water contaminated surfaces personal contact aerosol Reservoir humans humans animals animals Immunity high seroprevalence seroconversion by age of 5 y during the outbreak good short-lived Glass et al., 2001 Endemic childhood disease Epidemic disease Virus variants limited in a given area several genotypes various in different areas Clinical might be mild, self-limiting manifestation severe (life threatening) Prevention vaccination (Rotavirus A) outbreak control safer food handling water surveillance Glass et al., 2001 100 Baranya County, 1984-2000 90 80 70 G1 60 G2 50 G3 G4 40 G6 G9 30 20 10 0 1984-19851985-19861986-19871987-19881988-19891989-19901990-19911991-19921992-19931993-19941994-19951995-19961996-19971997-19981998-19991999-2000 100 Budapest, 1988-2003 90 80 70 G1 60 G2 G3 50 G4 40 G6 G9 30 20 10 0 1988-19891989-19901990-19911991-19921992-19931993-19941994-19951995-19961996-19971997-19981998-19991999-20002000-20012001-20022002-2003 Endemic childhood disease Epidemic disease Virus variants limited in a given area several genotypes various in different areas Clinical might be mild, self-limiting manifestation severe (life threatening) Prevention vaccination (Rotavirus A) outbreak control safer food handling water surveillance Glass et al., 2001 Endemic childhood disease Epidemic disease Virus variants limited in a given area several genotypes various in different areas Clinical might be mild, self-limiting manifestation severe (life threatening) Prevention vaccination (Rotavirus A) outbreak control safer food handling water surveillance Glass et al., 2001 Kapikian et al., 2001 Estimated no. of rotavirus infections accordig to the disease severity 400 000 - 500 000 deaths 2 000 000 hospital admission 25 000 000 medical visit 111 000 000 home treatment Parashar et al., 2003 Why is vaccination What benefits are needed? expected? •high mortality •prevention of severe outcome infections •high morbidity decreasing the no. •high costs of treatment of hospitalizations and •limited influence of public deaths health interventions and •reduction of costs improvemments in water assoc. with the supply treatment •regional inequalities in health care systems •lack of antiviral therapy Animal strains Animal-human reassortant strains Attenuated human strains Alternative approaches Santos and Hoshino, 2004 Parental strains Reassortant progenies Animal strains Animal-human reassortant strains Attenuated human strains Alternative approaches Santos and Hoshino, 2004.