title: Molecular Characterization of Polio from Environmental Samples: ISSP, The Israeli Sewage Surveillance Protocol.
abstract: Polioviruses are enteric viruses that cause paralytic poliomyelitis in less than 0.5 % of infections and are asymptomatic in >90 % infections of naïve hosts. Environmental surveillance monitors polio in populations rather than in individuals. When this very low morbidity to infection ratio, drops drastically in highly vaccinated populations, environmental surveillance employing manual or automatic sampling coupled with molecular analysis carried out in well-equipped central laboratories becomes the surveillance method of choice since polioviruses are excreted by infected individuals regardless of whether or not the infection is symptomatic. This chapter describes a high throughput rapid turn-around time method for molecular characterization of polioviruses from sewage. It is presented in five modules: (1) Sewage collection and concentration of the viruses in the sewage; (2) Cell cultures for identification of virus in the concentrated sewage; (3) Nucleic acid extractions directly from sewage and from tissue cultures infected with aliquots of concentrated sewage; (4) Nucleic Acid Amplification for poliovirus serotype identification and intratypic differentiation (discriminating wild and vaccine derived polioviruses form vaccine strains); and (5) Molecular characterization of viral RNA by qRT-PCR, TR-PCR, and Sequence analysis. Monitoring silent or symptomatic transmission of vaccine-derived polioviruses or wild polioviruses is critical for the endgame of poliovirus eradication. We present methods for adapting standard kits and validating the changes for this purpose based on experience gained during the recent introduction and sustained transmission of a wild type 1 poliovirus in Israel in 2013 in a population with an initial IPV vaccine coverage >90 %.
authors.affiliation: Environmental Virology Laboratory, Central Virology Laboratory, Public Health Services, Chaim Sheba Medical Center, Tel Hashomer, 52621, Israel. Lester.shulman@sheba.gov.health.il.|Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel. Lester.shulman@sheba.gov.health.il.
authors.forename: Lester M
authors.initials: LM
authors.lastname: Shulman
authors.affiliation: National Center for Viruses in the Environment, Central Virology Laboratory, Public Health Services, Chaim Sheba Medical Center, Tel Hashomer, 52621, Israel.
authors.forename: Yossi
authors.initials: Y
authors.lastname: Manor
authors.affiliation: Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel.|Molecular Assay Section, Central Virology Laboratory, Public Health Services, Chaim Sheba Medical Center, Tel Hashomer, 52621, Israel.
authors.forename: Musa
authors.initials: M
authors.lastname: Hindiyeh
authors.affiliation: Central Virology Laboratory, Public Health Services, Chaim Sheba Medical Center, Tel Hashomer, 52621, Israel.
authors.forename: Danit
authors.initials: D
authors.lastname: Sofer
authors.affiliation: Department of Epidemiology and Preventive Medicine, School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel.|Central Virology Laboratory, Public Health Services, Chaim Sheba Medical Center, Tel Hashomer, 52621, Israel.
authors.forename: Ella
authors.initials: E
authors.lastname: Mendelson
mesh_terms: D004784:Environmental Monitoring / Q000379:methods*; D006801:Humans; D007557:Israel; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D011051:Poliomyelitis / Q000821:virology*; D017955:Poliovirus / Q000235:genetics* / Q000302:isolation & purification*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D017423:Sequence Analysis, RNA / Q000379:methods; D012722:Sewage / Q000821:virology*; D046509:Tissue Culture Techniques / Q000379:methods
chemical_list: D012367:RNA, Viral; D012722:Sewage
keywords: Environmental surveillance; Intratypic differentiation; Oral polio vaccine (OPV); Plaque assay; Poliovirus; RNA purification; RT-PCR; Sequence analysis; Tissue culture; Vaccine-derived poliovirus (VDPV); Wild poliovirus; qRT-PCR