• Title/Summary/Keyword: Human calicivirus

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Application of Buoyant Density Centrifugation Method for the Rapid Detection of Feline Calicivirus in Oyster and Lettuce as Norovirus Surrogate

  • Cho, Yun-Sik;Lee, Kang-Whie;Jang, Keum-Il;Ahn, Jun-Bae;Kim, Kwang-Yup
    • Food Science and Biotechnology
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    • v.17 no.5
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    • pp.925-930
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    • 2008
  • Norovirus has become the most common cause of human gastroenteritis in developed countries. Detection procedures of foodborne viruses from foods require several steps. The concentration step using polyethylene glycol (PEG) is time-consuming and the detection efficiency of reverse transcription-polymerase chain reaction (RT-PCR) is affected by inhibitors from food components. In this study, a rapid detection method based on buoyant density centrifugation was developed to replace the time-consuming chloroform-polyethylene glycol-Tris Tween method. Feline calicivirus that belongs to the family Caliciviridae was used as a surrogate model for norovirus. After artificial inoculation of feline calcivirus (FCV) to oyster and lettuce, 830 ${\mu}L$ of homogenized sample suspension was layered on the top of 670 ${\mu}L$ 20% percoll and centrifuged. Then RNA extraction step was proceeded with the supernatant. By varying several physical conditions, the detection limits were lowered to $2.4{\times}10^2$ PFU per 1 g in oyster and $2.4{\times}10^0$ PFU per 1 g in lettuce. The protocol obtained in this study could be used to develop new detection method for norovirus in foods.

Antiviral Effect of Korean Red Ginseng Extract and Ginsenosides on Murine Norovirus and Feline Calicivirus as Surrogates for Human Norovirus

  • Lee, Min-Hwa;Lee, Bog-Hieu;Jung, Ji-Youn;Cheon, Doo-Sung;Kim, Kyung-Tack;Choi, Chang-Sun
    • Journal of Ginseng Research
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    • v.35 no.4
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    • pp.429-435
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    • 2011
  • Korean red ginseng has been studied various biological activities such as immune, anti-oxidative, anti-microbial, and anticancer activities but antiviral mechanism needs further studies. In this study, we aimed to examine the antiviral effects of Korea red ginseng extract and ginsenosides on norovirus surrogate, including murine norovirus (MNV) and feline calicivirus (FCV). We evaluated the pre-, co-, and post-treatment effects of Korean red ginseng (KRG), ginsenosides $Rb_1$ and $Rg_1$. To measure the antiviral effect and cytotoxicity of KRG extract, and ginsenosides $Rb_1$ and $Rg_1$, we treated Crandell-Reese Feline Kidney for FCV or RAW264.7 cells for MNV with concentrations of 0, 5, 6.7, 10, 20 ug/mL total saponin. There was cytotoxic effect in the highest concentration 20 ug/mL of KRG extract so this concentration was excluded in this study. The FCV titer was significantly reduced to 0.23-0.83 $log_{10}$ 50% tissue culture infectious dose ($TCID_{50}$)/mL in groups pre-treated with red ginseng extract or ginsenosides. The titer of MNV was significantly reduced to 0.37-1.48 $log_{10}$ $TCID_{50}$/mL in groups pre-treated with red ginseng extract or ginsenosides. However, there was no observed antiviral effect in groups co-treated or post-treated with KRG and its constituents. Our data suggest that KRG extract has an antiviral effect against norovirus surrogates. The antiviral mechanisms of KRG and ginsenosides should be addressed in future studies.

Expression and Antigenicity of Replicase Protein from Snow Mountain-Like Caliciviruses, Korean Isolates (한국형 사람 Calicivirus Replicase 단백의 발현 및 항원성 평가)

  • Chang, Mi-Yoon;Yang, Jai-Myung;Kim, Kyung-Hee
    • The Journal of Korean Society of Virology
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    • v.27 no.2
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    • pp.151-160
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    • 1997
  • In view of the potential of replicase protein as a diagnostic reagent for human caliciviruses (HuCVs), we have cloned and over-expressed this gene from the Snow Mountain-like Korean strains in Escherichia coli as a fusion protein with glutathione S-transferase (GST), and described the preliminary antigenic characterization of the recombinant products. Each 470bp fragment corresponding to highly conserved region of RNA-dependent RNA polymerase was generated by RT-PCR from stools of two diarrheal children, cloned in pMOSBlue T-vector, and subcloned between the EcoRI and SalI restriction sites of pGEX-4T-3, a GST gene fusion vector, yielding $pGCV_{pol}$. This construct expressed a Snow Mountain-like HuCV replicase under the control of the IPTG-inducible tac promoter. An extract prepared by sonication of the E. coli cell inclusion bodies bearing $pGCV_{pol}$ products was purified and analyzed by SDS-PAGE. After Coomassie blue staining, it was shown that the recombinant replicase migrated on the gels with an approximate molecular mass of 46.5 kDa, that was subsequently cleaved into a 26 kDa GST fragment and a 20.5 kDa replicase protein upon digestion with thrombin protease. The replicase was recognized on immunoblotting with the sera from symptomatic children with the HuCV-associated diarrhea but not by asymptomatic sera from adults. The results presented the first biological activity of individually expressed HuCV replicase subunit and provided important reagents for diagnosis of HuCV infection.

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Seroprevalence of Norwalk-Like Viruses (NLVs) in Seoul: Detection Using Baculovirus-Expressed Norwalk Virus Capsid Antigen (Baculovirus에서 발현된 Norwalk Virus 캡시드 항원에 대한 국내인의 혈청학적 반응도)

  • Kang, Shin-Jai;Nam, Ki-Bum;Cho, Yang-Ja;Kim, Kyung-Hee
    • The Journal of Korean Society of Virology
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    • v.27 no.2
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    • pp.161-168
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    • 1997
  • The present study was designed to estimate the seroprevalence of NLVs among diarrheagenic children and in healthy adults in Seoul and its vicinity with the use of an EIA and an Western blot (WB) based on recombinant Norwalk virus capsid protein (rNV) and crude virus preparations as antigen. Seroconversion was observed in 34 (83%) of 41 tested using the EIA and in 21 (54%) of 39 using the WB, suggesting that the NLVs with epitopes common to rNV are prevalent in Seoul area. Diarrheal children who were known to have been infected with several other strains of the NLVs showed no significant antibody response to the rNV. Infection with rNV occurred earlier in life: primary infections with rNV were common before the age of 6 months and over 91 % of children had evidence of infection by that age by the EIA. Since the amount of the NLV antigens available for seroepidemiologic surveys is limited, we tried to detect NLV antibody by using crude virus preparations as antigen. One crude virus preparation of a child whose stool yielded genetically distinct NLV revealed the presence of the plural number of bands upon SDS-PAGE, but precipitated only one band (62 kDa) after the WB with a serum (collected 10 days after the onset of symptoms) of another diarrheal child. The WB assay we present in this report revealed that the NLVs are prevalent among Korean population and that the sera contained antibody to a single major structural protein, with molecular sizes of 58 to 62 kDa, compatible with the sizes reported for the Norwalk virus and Snow Mountain agent proteins, respectively. When the results of the WB were compared with those obtained by the EIA, the EIA antibody assay was sensitive enough to detect an antibody rise of as much as 4096-fold but not as specific as the WB. The WB assay presented in this paper will provide a powerful tool to elucidate not only antigenic structures of the NL Vs but also seroepidemiology of the NLV infection. The availability of an unlimited source of antigen will enable a large scale serologic studies that will greatly increase our understanding of the role of NLVs in human enteric illness.

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Recent (2010-2019) foodborne outbreaks caused by viruses in the Republic of Korea along with their detection and inactivation methods (바이러스에 의한 최근(2010-2019) 국내 식중독 사고와 검출법 및 제어법에 대한 동향 조사)

  • Kwon, Seung-Wook;Kim, Sang-Soon
    • Korean Journal of Food Science and Technology
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    • v.53 no.1
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    • pp.1-11
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    • 2021
  • In this review, recent foodborne outbreaks caused by viruses in the Republic of Korea (2010-2019) were analyzed. The human norovirus was found to be the major foodborne virus causing an average of 94.9% of the viral outbreaks. Reverse-transcription polymerase chain reaction (PCR) with electrophoresis has been widely used to detect viruses, but several rapid detection methods, including real-time PCR, multiplex PCR, and quantum dot assay, have also been suggested. For norovirus inactivation studies, surrogates such as murine norovirus and feline calicivirus have been widely used to identify the reduction rate owing to the limitations in laboratory cultivation. Conversely, direct cell infection studies have been conducted for other foodborne viruses such as adenovirus, astrovirus, rotavirus, and hepatitis A or E virus. Moreover, virucidal mechanisms using various physical and chemical treatments have been revealed. These recent studies suggest that rapid in situ detection and effective control are valuable for ensuring food safety against viral infections.

Clinical Manifestations of Norovirus Infection in Korean Pediatric Cancer Patients (한국 소아 암환자에서 노로바이러스 감염증의 임상 양상)

  • Choi, Hyunshin;Choi, Young Bae;Hwang, Ji-Young;Cheon, Doo-Sung;Jeong, Hye Sook;Choe, Yon Ho;Yoo, Keon Hee;Sung, Ki Woong;Koo, Hong Hoe;Kim, Yae-Jean
    • Pediatric Infection and Vaccine
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    • v.18 no.1
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    • pp.40-47
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    • 2011
  • Purpose : Norovirus infection, a common cause of community-acquired gastroenteritis, can also lead to severe illness in immunocompromised patients. We investigated clinical manifestations of norovirus infection in pediatric cancer patients. Methods : Stool specimens were collected from pediatric patients with gastrointestinal symptoms between November 2008 and September 2009 at Samsung Medical Center, Seoul, Korea. Norovirus infection was identified by reverse-transcription polymerase chain reaction (RT-PCR). A retrospective chart review was performed in pediatric cancer patients who were diagnosed with norovirus infection. Results : Ten patients were diagnosed with norovirus infection by RT-PCR in stool samples. The median age was 0.83 years (range 0.25-5.5 years) and the male to female ratio was 1.5:1 (6 males and 4 females). Underlying diseases were hematologic malignancies (4/10, 40%), neuroblastoma (4/10, 40%), and brain tumors (2/10, 20%). Three patients were infected before hematopoietic cell transplantation (HCT) and four patients after HCT. All patients had diarrhea (10/10, 100%), with a median frequency of diarrhea of 8.5 times/day (range 4-22 times/day). Median virus shedding duration was 72.5 days (range 19-299 days). Four patients with pneumatosis intestinalis were conservatively treated with bowel rest and total parenteral nutrition. One patient with severe diarrhea and bloody stool had concomitant chronic gut graft-versus-host disease (GVHD). Norovirus infection-related mortality was not observed. Conclusion : Norovirus infection can cause significant clinical manifestations with prolonged viral shedding in immunocompromised patients. Norovirus should be considered in pediatric cancer patients with severe gastrointestinal symptoms.