I have searched hard to find a reliable and best research paper writing service and finally i got a good option for my needs as ⇒ www.WritePaper.info ⇐, Hello! Briefly, cells inoculated with clinical material were incubated at 35°C for 7 days. Representative results for the protocol of RT-PCR plus hybridization. Thirteen specimens were collected from an outbreak in 2001 and 2002 in Henan province, and nine were collected from a single outbreak in 2000 and 2001 in Anhui province. Rubella virus is the sole member of the genus Rubivirus in the family Togaviridae. One word – EXCELLENT! Detection of rubella virus is often necessary to confirm rubella cases and to identify specimens to be used to characterize wild-type rubella viruses.
Rub the swab firmly over the back of the throat, both tonsils and any areas of inflammation, exudation or ulceration.
Viral isolates were obtained from both provinces by using the culture methods described above; the isolates were characterized further for molecular epidemiologic purposes (data not shown).
Blood for culture should be collected instead. The methods used were reverse transcription (RT)-PCR followed by Southern hybridization using RNA extracted directly from clinical specimens, virus growth in tissue culture followed by virus detection by RT-PCR, low-background immunofluorescence in infected tissue culture cells using monoclonal antibodies to the structural proteins of rubella virus, and a replicon-based method of detecting infectious virus. Routine rubella vaccination is not included in the national immunization program in China, although rubella vaccine is available in certain urban areas. The technique used here relies on a stable cell line expressing a replicon and requires only standard virus passage and fluorescent microscopic techniques for the detection of infections virus (i.e., no RNA synthesis, no transfection, etc.). Nevertheless, approximately 3 to 23% of adults remain susceptible in various countries and areas (4, 8, 15, 17). Since the high sensitivity of RT-PCR was increased by the addition of the hybridization step rather than a nested PCR, contamination by PCR products, which often occurs with the nested RT-PCR technique, was minimized. It contains a 5′-proximal open reading frame (ORF) that encodes nonstructural proteins, which are responsible for viral genome replication, and a 3′-proximal ORF, which encodes three structural proteins, C, E2, and E1 (10). Many rubella cases were IgM negative and infectious virus positive at 1 day after the onset of rash (2). It was estimated in 2003 that more than 100,000 infants were born with CRS each year worldwide (16). A puromycin resistance gene was inserted after the first subgenomic promoter, and the gene encoding GFP was inserted after the second subgenomic promoter. and mouth specimens However, virus is usually present in the throat or nasopharynx from a few days before until about 5 days after the onset of rash, making virus detection a possible accessory test for diagnosis (2).
Among the techniques requiring tissue culture, the replicon-based method is particularly simple to do, requiring only tissue culture and microscopic techniques. RT-PCR.Viral RNA from throat swabs was extracted using Tri-Reagent LS for liquid samples (Molecular Research Center, Cincinnati, OH) according to the manufacturer's protocol. They can swab the back of the throat or nose and test it for the bacteria that cause diphtheria.
My professor asked me to write a research paper based on a field I have no idea about. Thus, compared with RT-PCR plus hybridization, these culture-based methods each detected about 69% (9/13) of the positive specimens from Henan.
The techniques described here require only equipment that is commonly available, unlike some other techniques (e.g., real-time RT-PCR).
By RT-PCR alone, 68% of the specimens (15/22) tested positive.
The use of this low-background IFA protocol was necessary since rubella virus replicates to relatively low levels compared to levels of other viruses (e.g., measles virus). However, the other three methods also performed well, detecting most of the specimens positive by RT-PCR plus hybridization (81%). The method using culture plus RT-PCR and the IFA method are rather complex procedures, require good molecular reagents, and are time-consuming, requiring 3 weeks to complete all passages. If you continue browsing the site, you agree to the use of cookies on this website. The BRDII attenuated vaccine strain of rubella virus (Tian Tan Biological Products Corporation, Beijing, China) was introduced in 1994 in some parts of China including the city of Shanghai and Shandong province.
A throat swab culture, or throat culture, is a test commonly used to diagnose bacterial infections in the throat. Thus, this technique is considered to be the least difficult method to perform and could be automated. In China, 331 subnational laboratories were established and a laboratory surveillance network was started in 2000. Among these four methods, direct RT-PCR followed by hybridization was the most sensitive method; the replicon-based method was the least difficult to perform. American Society for Microbiology These primers were part of a nested set reported previously (3). Mock-infected Vero cells were always carried through this protocol with each set of clinical specimens as a negative control. See our Privacy Policy and User Agreement for details.
Comparison of sensitivities of techniques for detecting infectious rubella virus or viral RNA using specimens from Anhui and Henan.
Clinical specimens.Twenty-two throat swabs specimens were collected from patients in China with clinically suspected rubella virus infection (Table 1).
GFP-expressing replicons used for the detection of rubella virus as described previously are cumbersome for a number of reasons, including the need to transfect previously infected Vero cells with replicon RNA (19). By RT-PCR alone, 68% of the specimens (15/22) tested positive. False-negative results may be obtained if only a single serum sample taken around the day of rash onset is collected, the most likely day for serum collection. We do not retain these email addresses. In addition, an imported measles-mumps-rubella vaccine containing the rubella virus vaccine strain RA27/3 (Merck and Co., Inc., Whitehouse Station, NJ) has been available since 1996 in large cities (20, 24). The 143-bp DNA probe was made by RT-PCR using a forward primer (RV13 [5′-CTC GAG GTC CAG GTC CYG CC-3′]) and a reverse primer (RV14 [5′-GAA TGG CGT TGG CAA ACC GG-3′]), DIG-labeled dUTPs (Roche Molecular Biochemicals, Mannheim, Germany), and Therien RNA.
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