Application of a Receptor-Binding Capture Quantitative Reverse Transcription-PCR Assay To Concentrate Human Norovirus from Sewage and To Study the Distribution and Stability of the Virus

Tian, Peng; Yang, David; Pan, Liangwen; Mandrell, Robert E.

doi:10.1128/aem.06875-11

Cited by 30 publications

(13 citation statements)

References 40 publications

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“…In addition, the partially purified PGM powder used in this study contained 0.5 to 1.5% sialic acid (Sigma-Aldrich), which was reported to be the receptor for murine norovirus (MNV) (11). Utilizing the ability of PGM to bind to HuNoV, Tian et al (9,12,13) and Pan et al (14) used PGM-conjugated magnetic beads (PGM-MBs) to extract and concentrate HuNoV from food matrices, such as fresh produce, salad, and sewage. After binding, only bound virus particles can be quantified using qRT-PCR.…”

mentioning

confidence: 99%

Evaluation of the Porcine Gastric Mucin Binding Assay for High-Pressure-Inactivation Studies Using Murine Norovirus and Tulane Virus

Chen

2015

Appl Environ Microbiol

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We compared the results of high-hydrostatic-pressure (HHP) inactivation of murine norovirus type 1 (MNV-1) and Tulane virus (TV) obtained by a porcine gastric mucin binding assay followed by quantitative reverse transcription-PCR (referred to here as the PGM-MB/PCR assay) and a plaque assay and evaluated HHP inactivation of a human norovirus (HuNoV) genogroup I genotype 1 (GI.1) strain and a HuNoV GII.4 strain by using the PGM-MB/PCR assay. Viruses were treated at different pressure levels for 2 min at 4 or 21°C in culture medium of neutral pH and in culture medium of pH 4 at 21°C. The log reductions of infectious MNV-1 and TV particles caused by HHP were assessed using the PGM-MB/PCR and plaque assays, while the log reductions of HuNoVs were assessed by the PGM-MB/PCR assay only. For TV and MNV-1, the two pressure inactivation curves obtained using the plaque and PGM-MB/PCR assays were almost identical at <2-log-reduction levels regardless of the treatment temperature and pH. Further increasing the pressure over the 2-log-reduction level resulted in higher log reductions of TV and MNV-1, as assessed by the plaque assay, but did not increase the log reductions, as assessed by the PGM-MB/PCR assay. HHP treatments could achieve maximum reductions of ϳ3 and 3.5 log units for GI.1 and GII.4, respectively, as assessed by the PGM-MB/PCR assay. On the basis of these results, it can reasonably be concluded that the PGM-MB/PCR assay would very likely be able to estimate HHP inactivation of HuNoV at <2-log-reduction levels. It would also likely conservatively quantify HHP inactivation of the GI.1 strain at 2-to 3-log-reduction levels and the GII.4 strain at 2-to 3.5-log-reduction levels.H uman norovirus (HuNoV) is the leading cause of foodborne illnesses in the United States (1). It has frequently been associated with a variety of ready-to-eat foods, including berries, salsa, and guacamole, and raw shellfish, such as oysters (2-4). Currently, the lack of suitable cell culture systems and practical small-animal models has been hindering research for developing and/or identifying effective processing methods to inactivate HuNoV (1, 5). Therefore, evaluation of the efficacy of processing treatments must rely on HuNoV surrogates. The accuracy of methods that use these surrogates is questionable since direct comparison of methods that use surrogates with methods that use HuNoV is not possible. Molecular biology methods, mostly quantitative reverse transcription-PCR (qRT-PCR), can be used for HuNoV quantification. However, these methods can detect only the presence of HuNoV RNA and cannot distinguish between infectious and noninfectious virus particles.HuNoVs are reported to bind to histo-blood group antigens (HBGAs) in the human intestinal tract, and HBGAs are considered the attachment factors necessary for infection (6, 7). To initiate infection, HuNoVs need to be able to bind to HBGAs to enter the host cells. Porcine gastric mucin (PGM) contains HBGAs and has been shown to be able to bind to genogroup I (GI) and genogroup II...

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confidence: 99%

Evaluation of the Porcine Gastric Mucin Binding Assay for High-Pressure-Inactivation Studies Using Murine Norovirus and Tulane Virus

Chen

2015

Appl Environ Microbiol

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“…The slope was −1.496 cycles/log 10 for GI HuNoVs with an R 2 of 0.9974 and was −1.4648 cycles/log 10 for GII HuNoVs with an R 2 of 0.9991 (Tian et al, 2012). …”

Section: Methodsmentioning

confidence: 99%

In Situ Capture RT-qPCR: A New Simple and Sensitive Method to Detect Human Norovirus in Oysters

Zhou

Tian

et al. 2017

Front. Microbiol.

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Human noroviruses (HuNoVs) are the major cause worldwide for non-bacterial acute gastroenteritis. In this study, we applied a novel viral receptor mediated in situ capture RT-qPCR (ISC-RT-qPCR) to detect HuNoVs in oysters and compared with the traditional RT-qPCR method. Ten HuNoVs RT-PCR positive and 5 negative clinical samples from gastroenteritis patients were used to compare specificity and sensitivity of ISC-RT-qPCR against that of the RT-qPCR assay. ISC-RT-qPCR had at a one-log and a two-log increase in sensitivity over that of the RT-qPCR assay for genotype I (GI) and GII, respectively. Distributions of HuNoVs in oyster tissues were investigated in artificially inoculated oysters. GI HuNoVs could be detected in all tissues in inoculated oysters by both ISC-RT-qPCR and RT-qPCR. GII HuNoVs could only be detected in gills and digestive glands by both methods. The number of viral genomic copies (vgc) measured by ISC-RT-qPCR was comparable with RT-qPCR in the detection of GI and GII HuNoVs in inoculated oysters. Thirty-six oyster samples from local market were assayed for HuNoVs by both assays. More HuNoVs could be detected by ISC-RT-qPCR in retail oysters. The detection rates of GI HuNoVs in gills, digestive glands, and residual tissues were 33.3, 25.0, and 19.4% by ISC-RT-qPCR; and 5.6, 11.1, and 11.1% by RT-qPCR, respectively. The detection rates of GII HuNoVs in gills were 2.8% by ISC-RT-qPCR; no GII HuNoV was detected in these oysters by RT-qPCR. Overall, all results demonstrated that ISC-RT-qPCR is a promising method for detecting HuNoVs in oyster samples.

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“…However, the use of filters limits the speed of the procedure, is ineffective for processing samples with high amounts of particulate matter, and demands relatively large volumes of eluent for primary and secondary virus recovery. Alternatively, bioaffinity approaches that utilize magnetic separation have been developed in which magnetic beads are coated with capsid-binding ligands like HBGAs or porcine gastric mucin (PGM) that can be used to concentrate HuNoV from food or environmental water samples (33,175,229,(237)(238)(239)(240). Positively charged magnetic beads, which combine charge-based separation with the advantages of magnetic separation, have been used.…”

Section: Sampling and Presampling Considerationsmentioning

confidence: 99%

Response to the Questions Posed by the Food Safety and Inspection Service, the Centers for Disease Control and Prevention, the National Marine Fisheries Service, and the Defense Health Agency, Veterinary Services Activity Regarding Control Strategies for Reducing Foodborne Norovirus Infections†

2016

Journal of Food Protection

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Application of a Receptor-Binding Capture Quantitative Reverse Transcription-PCR Assay To Concentrate Human Norovirus from Sewage and To Study the Distribution and Stability of the Virus

Cited by 30 publications

References 40 publications

Evaluation of the Porcine Gastric Mucin Binding Assay for High-Pressure-Inactivation Studies Using Murine Norovirus and Tulane Virus

Evaluation of the Porcine Gastric Mucin Binding Assay for High-Pressure-Inactivation Studies Using Murine Norovirus and Tulane Virus

In Situ Capture RT-qPCR: A New Simple and Sensitive Method to Detect Human Norovirus in Oysters

Response to the Questions Posed by the Food Safety and Inspection Service, the Centers for Disease Control and Prevention, the National Marine Fisheries Service, and the Defense Health Agency, Veterinary Services Activity Regarding Control Strategies for Reducing Foodborne Norovirus Infections†

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