Biomaterial Database

Simultaneous detection of enteric viruses by multiplex real-time RT-PCR. 2004

Christian Beuret

Spiez Laboratory, Christian Beuret, 3700 Spiez, Switzerland. Christian.beuret@gr.admin.ch

A multiplex real-time RT-PCR protocol for the simultaneous detection of noroviruses ("Norwalk-like viruses") of genogroups I and II, human astroviruses and enteroviruses is described. The protocol was developed and evaluated using the LightCycler and corresponding SYBR Green reagents. New primers were designed within conserved genome regions to optimize the detection range of virus subtypes of each genus. To enable the development of a multiplex PCR assay within one tube (capillary), similar mastermix- and cycling-conditions were respected for each individual primer system. Subsequent melting curve analysis allowed the determination of possible dual-contaminations of entero- and noro- or astroviruses by the formation of dual peaks. Special care was taken to minimize the loss of sensitivity, since the detection of small viral contaminations is a crucial parameter especially for food analysis. The multiplex assay was compared successfully to the single SYBR Green assay, and revealed to be at least 10 times more sensitive than the one obtained with an endpoint PCR thermocycler protocol published previously.

UI	MeSH Term	Description	Entries
D009663	Norwalk virus	The type species in the genus NOROVIRUS, first isolated in 1968 from the stools of school children in Norwalk, Ohio, who were suffering from GASTROENTERITIS. The virions are non-enveloped spherical particles containing a single protein. Multiple strains are named after the places where outbreaks have occurred.	Norwalk Agent
D009930	Organic Chemicals	A broad class of substances containing carbon and its derivatives. Many of these chemicals will frequently contain hydrogen with or without oxygen, nitrogen, sulfur, phosphorus, and other elements. They exist in either carbon chain or carbon ring form.	Organic Chemical,Chemical, Organic,Chemicals, Organic
D011804	Quinolines
D003959	Diamines	Organic chemicals which have two amino groups in an aliphatic chain.	Diamine
D004770	Enterovirus	A genus of the family PICORNAVIRIDAE whose members preferentially inhabit the intestinal tract of a variety of hosts. The genus contains many species. Newly described members of human enteroviruses are assigned continuous numbers with the species designated "human enterovirus".	Coxsackie Viruses,Coxsackieviruses
D005759	Gastroenteritis	INFLAMMATION of any segment of the GASTROINTESTINAL TRACT from ESOPHAGUS to RECTUM. Causes of gastroenteritis are many including genetic, infection, HYPERSENSITIVITY, drug effects, and CANCER.	Gastroenteritides
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001257	Mamastrovirus	A genus of small, circular RNA viruses in the family ASTROVIRIDAE. They cause GASTROENTERITIS and are found in the stools of several vertebrates including humans. Transmission is by the fecal-oral route and there are at least eight human serotypes. The type species is Human astrovirus.	Human astrovirus,Human astroviruses,Mamastroviruses,astroviruses, Human
D012367	RNA, Viral	Ribonucleic acid that makes up the genetic material of viruses.	Viral RNA
D052160	Benzothiazoles	Compounds with a benzene ring fused to a thiazole ring.