Real-time PCR for mumps diagnosis on clinical specimens—Comparison with results of conventional methods of virus detection and nested PCR

Krause, C.; Eastick, Kirstine; Ogilvie, Marie M.

doi:10.1016/j.jcv.2006.07.009

Cited by 52 publications

(32 citation statements)

References 14 publications

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“…Follicle-stimulating and several similar hormones are excreted in urine at great enough concentration to be extracted commercially. Alternatively, TSE infectivity may be excreted by processes analogous to those responsible for the low-level virurias that occur during infections of the nervous system by mumps, measles, and West Nile virus (28)(29)(30).…”

Section: Discussionmentioning

confidence: 99%

Excretion of Transmissible Spongiform Encephalopathy Infectivity in Urine

Gregori¹,

Kovács²,

Alexeeva³

et al. 2008

Emerg. Infect. Dis.

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The route of transmission of most naturally acquired transmissible spongiform encephalopathy (TSE) infections remains speculative. To investigate urine as a potential source of TSE exposure, we used a sensitive method for detection and quantitation of TSE infectivity. Pooled urine collected from 22 hamsters showing clinical signs of 263K scrapie contained 3.8 ± 0.9 infectious doses/mL of infectivity. Titration of homogenates of kidneys and urinary bladders from the same animals gave concentrations 20,000-fold greater. Histologic and immunohistochemical examination of these same tissues showed no indications of infl ammatory or other pathologic changes except for occasional deposits of disease-associated prion protein in kidneys. Although the source of TSE infectivity in urine remains unresolved, these results establish that TSE infectivity is excreted in urine and may thereby play a role in the horizontal transmission of natural TSEs. The results also indicate potential risk for TSE transmission from human urine-derived hormones and other medicines.T ransmissible spongiform encephalopathies (TSEs) are fatal neurologic diseases. In humans, a long asymptomatic incubation period is followed by a progressive clinical course that typically lasts a few months to a year. TSE infectivity and pathologic changes are concentrated in the nervous system; however, much of the transmission risk results from parenteral exposure to the much lower concentrations of infectivity found in tissues outside the nervous system. Thus, despite the very low concentration of TSE infectivity in blood (1,2), 4 human cases of transmission of variant Creutzfeldt-Jakob disease through blood transfusions have been documented (3,4). If TSE infectivity were excreted, human urine, which is a source of injectible fertility hormones and other drugs (5,6), could also pose a risk for transmission. Infected urine might also account for the horizontal transmission of sheep scrapie and might contribute to the natural spread of chronic wasting disease in deer and elk.Early attempts to transmit Creutzfeldt-Jakob disease by cross-species inoculation of rodents and primates with urine from diseased patients failed (7,8). More recent attempts in which urine from infected hamsters was injected back into hamsters have produced variable results (9,10). Two other studies have reported infectivity in urine (11) and infectivity with disease-specifi c prion protein (PrP d ) in kidneys of mice with simultaneous scrapie and nephritis but not in those with scrapie alone (12). To resolve these discrepancies, we used a highly sensitive and precise method of measuring low concentrations of TSE infectivity, which we have successfully used for quantitation of TSE infectivity in blood (1,2), to measure the concentration of TSE infectivity in urine of scrapie-infected hamsters. Materials and Methods Urine Collection and ProcessingUrine was collected from a cohort of 22 Syrian hamsters (Harlan Sprague-Dawley, Haslet, MI, USA) that had been infected by intracranial injection with...

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Section: Discussionmentioning

confidence: 99%

Excretion of Transmissible Spongiform Encephalopathy Infectivity in Urine

Gregori¹,

Kovács²,

Alexeeva³

et al. 2008

Emerg. Infect. Dis.

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“…Other PCR diagnostic tests for mumps virus detection have been developed previously, including nested and two-step RT-PCR assays (11,17,19,22). A recent study describes a successful modification of the real-time RT-PCR, used by Uchida and colleagues, into a one-step assay (12). The authors reported a high degree of sensitivity and specificity for oral specimens but not for urine specimens (12).…”

Section: Discussionmentioning

confidence: 99%

Real-Time Reverse Transcription-PCR Assay for Detection of Mumps Virus RNA in Clinical Specimens

et al. 2007

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The mumps virus is a negative-strand RNA virus in the family Paramyxoviridae. Mumps infection results in an acute illness with symptoms including fever, headache, and myalgia, followed by swelling of the salivary glands. Complications of mumps can include meningitis, deafness, pancreatitis, orchitis, and first-trimester abortion. Laboratory confirmation of mumps infection can be made by the detection of immunoglobulin M-specific antibodies to mumps virus in acute-phase serum samples, the isolation of mumps virus in cell culture, or by detection of the RNA of the mumps virus by reverse transcription (RT)-PCR. We developed and validated a multiplex real-time RT-PCR assay for rapid mumps diagnosis in a clinical setting. This assay used oligonucleotide primers and a TaqMan probe targeting the mumps SH gene, as well as primers and a probe that targeted the human RNase P gene to assess the presence of PCR inhibitors and as a measure of specimen quality. The test was specific, since it did not amplify a product from near-neighbor viruses, as well as sensitive and accurate. Real-time RT-PCR results showed 100% correlation with results from viral culture, the gold standard for mumps diagnostic testing. Assay efficiency was over 90% and displayed good precision after performing inter-and intraassay replicates. Thus, we have developed and validated a molecular method for rapidly diagnosing mumps infection that may be used to complement existing techniques.

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“…For this reason, the elution volume was decreased for the High Pure PCR Template Preparation kit, the input volume was doubled for the QIAamp DNA Mini kit and water was not added to the swab when the RTP DNA/RNA Virus Mini kit was employed. Increasing sample volume has also been performed by Krause et al (2006), who used oral samples for the detection of mumps and obtained good recovery.…”

Section: Discussionmentioning

confidence: 99%

A comparison of molecular methods for hepatitis B virus (HBV) DNA detection from oral fluid samples

Portilho

Martins

Lampe

et al. 2012

Journal of Medical Microbiology

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The objective of the present study was to evaluate four commercial DNA extraction methods and three PCR protocols for hepatitis B virus (HBV) detection in artificially contaminated oral fluid samples. The extraction protocols were selected based on ease of use and cost, and were also compared with respect to sensitivity and cost. Prior PCR optimization was conducted, in which the sample volume for DNA extraction and the concentrations of DNA and Taq DNA polymerase in the PCR were adjusted. One-round PCR, used to amplify the core region of the HBV genome, achieved high levels of sensitivity in comparison with nested and semi-nested PCR experiments that were designed for the amplification of HBV surface protein genes. Of the four extraction protocols evaluated, the RTP DNA/RNA Virus Mini kit and the QIAamp DNA Mini kit gave the highest recovery rates, presenting 20 copies of HBV DNA ml "1 as the limit of detection. These results suggest that HBV DNA can be detected from oral fluid samples but that the optimization of the PCR assays and the choice of extraction methods must be determined by laboratories before the implementation of this method in routine diagnostics.

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Real-time PCR for mumps diagnosis on clinical specimens—Comparison with results of conventional methods of virus detection and nested PCR

Cited by 52 publications

References 14 publications

Excretion of Transmissible Spongiform Encephalopathy Infectivity in Urine

Excretion of Transmissible Spongiform Encephalopathy Infectivity in Urine

Real-Time Reverse Transcription-PCR Assay for Detection of Mumps Virus RNA in Clinical Specimens

A comparison of molecular methods for hepatitis B virus (HBV) DNA detection from oral fluid samples

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