U.S. flagAn official website of the United States government
                    Here's how you know **Official websites use .gov**
                            
                            A **.gov** website belongs to an official government organization in the United States.
 **Secure .gov websites use HTTPS**
                            
                            A **lock** (                            
                              A locked padlock                                                      
                            
                            ) or **https://** means you’ve safely connected to the .gov website. Share sensitive information only on official, secure websites.


Menu Agricultural Research Service
						
                    U.S. DEPARTMENT OF AGRICULTURE
 close



Research
                Research Home
                            National Programs
                            Research Projects
                            Scientific Manuscripts
                            International Engagement
                            Scientific Software/Models
                            Databases and Datasets
                            Office of Scientific Quality Review
            Media
                News & Features
                            Multimedia
                            Briefing Room
                            Down on the Farm
                            Press Room
                            Science in Your Shopping Cart
                            Scientific Discoveries
                            Tellus
                            Under the Microscope
            About ARS
                About ARS Home
                            Staff Directory
                            Labs and Research Centers (Map)
                            Headquarter Offices
                            Organizational Chart
                            Employee Services (REE Employees Only)
                            Office of Outreach, Diversity, and Equal Opportunity
            Work With Us
                Work With Us Home
                            ARS Vacancies at USAJOBS
                            Careers at ARS Information
                            Scientific Collaborations
                            Extramural Agreements
                            1890 Faculty Research Sabbatical ProgramSearch small
                    Search
            
            ARS Home
                                About ARS
                                Contact Us




ARS Home Research Publications at this Location





- ARS National Programs
- Search for a scientific manuscript
- Search for a research project
- ARS Software
- Programs and Projects
- Collaborations
- Subjects of Investigation
- Publications

Title:   Author  item
                                            Richards, Garyitem
                                            Watson, Michaelitem
                                            Kingsley, David
  Submitted to: Journal of Virological Methods
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 10/28/2003
                                    Publication  Date: 1/1/2004
                                    Citation: RICHARDS, G.P., WATSON, M.A., KINGSLEY, D.H. A SYBR GREEN REAL-TIME RT-PCR METHOD TO DETECT AND QUANTITATE NORWALK VIRUS IN STOOLS. JOURNAL OF VIROLOGICAL METHODS. 2004. vol. 116. pg. 67-70.Interpretive Summary: Norwalk virus is a common cause of illness in the United States and tests to detect this virus in human stools have been complicated.  We developed a new test, known as real-time reverse transcription-polymerase chain reaction, to detect Norwalk virus in stools.  We also evaluated a simple dilution and heating process in place of more complicated procedures to extract the viruses from stools.  These rapid and simple procedures were applied to 68  human stool isolates from a volunteer study and were successful in detecting Norwalk viruses in stools suspected to contain viruses.  The levels of virus present were determined using a new procedure, referred to as the dilution end-point standard curve.  One stool contained 60 billion Norwalk viruses per gram, which is far more than previously detected in human stools.  Such high counts provide a clue to why Norwalk-like viruses are the leading cause of non-bacterial, gastrointestinal illness in the United States.  Two of the volunteers shed Norwalk virus in their stools, even though they did not show any overt signs of illness.  This suggests the possibility that health care workers and food handlers could unknowingly spread Norwalk virus in the workplace.
                                Technical Abstract: This study was performed with three objectives: a) to develop a real-time reverse transcription-PCR (rt RT-PCR) procedure for a commonly used laboratory strain of Norwalk virus (NV), b) to evaluate the potential of sample dilution and heat release of viral RNA as an alternative to more complex procedures to extract viruses from stool specimens, and c) to identify the presence and relative quantity of NV in stools obtained from a highly controlled volunteer study.  We describe the development of a simple, single tube, hot start, rt RT-PCR technique using SYBR green fluorescence for the detection of the 8FIIa strain of NV in diluted stools.  Real-time RT-PCR was applied to 68 stool isolates from patients participating in a NV volunteer study.  Ten of the 20 volunteers (50%) exhibited clinical signs of NV illness (diarrhea or vomiting) and seroconverted.  The equivalent of 1 μl of a 1:1000 dilution of their stools was amplified by rt RT-PCR and consistently tested positive for NV amplicon without the need for virus extraction.  Based on the development of a dilution end-point standard curve to semi-quantitate minimum virus levels, the number of RT-PCR amplifiable viruses in stool specimens is as high as 61.6 billion NV per gram of stool.  Such high counts provide a clue to why Norwalk-like viruses are the number one cause of non-bacterial gastroenteritis in the United States. Two patients without clinical symptoms, but who seroconverted within 4 weeks of the challenge, shed NV in their stools, demonstrating a carrier state among asymptomatic individuals.  This has major public health implications, particularly with health care workers and food handlers who could be at risk of unknowingly spreading NV contamination.


Return to top


Connect with ARS Sign up


ARS Home
                            USDA.gov
                            Plain Writing
                            Policies & Links
                            Civil Rights Statements
                            FOIA
                            Accessibility Statement
                            Privacy Policy
                            Non-Discrimination Statement
                            Quality of Information
                            USA.gov
                            WhiteHouse.gov