Detection of hepatitis A virus in environmental samples by antigen-capture PCR

The efficacy of the antigen-capture PCR (AC-PCR) method for the detection of hepatitis A virus (HAV) in environmental samples was demonstrated. HAV was captured from a seeded liquid waste or a shellfish sample with homologous antibody and then heat denatured and subjected to reverse transcription an...

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Veröffentlicht in:Applied and Environmental Microbiology 1994-06, Vol.60 (6), p.1927-1933
Hauptverfasser: Deng, M Y, Day, S P, Cliver, D O
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container_title Applied and Environmental Microbiology
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creator Deng, M Y
Day, S P
Cliver, D O
description The efficacy of the antigen-capture PCR (AC-PCR) method for the detection of hepatitis A virus (HAV) in environmental samples was demonstrated. HAV was captured from a seeded liquid waste or a shellfish sample with homologous antibody and then heat denatured and subjected to reverse transcription and the PCR, all in the same tube. Subsequently, the AC-PCR products were analyzed by oligonucleotide probe hybridization in solution, agarose gel electrophoresis, and autoradiography. The AC-PCR detected as little as 0.053 PFU of cell culture-adapted HAV strain HM175/18f. The results of cDNA-RNA hybridization indicated that the particle/ PFU ratio of this virus strain is approximately 79:1. Therefore, the detection limit of the AC-PCR was estimated to be four virus particles. No amplified products were observed when poliovirus 1, coxsackievirus A9, coxsackievirus B3, echovirus 6, reovirus 1, adenovirus type 40, human rotavirus type 1, and bovine enterovirus type 2 were tested, confirming the specificity of the assay. There were no differences between the nucleotide sequences of AC-PCR products of HAV strain HM175/18f and the sequences of wild-type HAV strain HM175 derived from molecularly cloned cDNA. Of 121 waste and shellfish samples tested by both plaque assays (PA) in cell cultures and the AC-PCR, 81 (67%) were positive and 31 (26%) were negative as determined by both methods, whereas 9 (7%) were positive as determined by the AC-PCR and negative as determined by the PA, and none were positive as determined by the PA and negative as determined by the AC-PCR.
doi_str_mv 10.1128/aem.60.6.1927-1933.1994
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HAV was captured from a seeded liquid waste or a shellfish sample with homologous antibody and then heat denatured and subjected to reverse transcription and the PCR, all in the same tube. Subsequently, the AC-PCR products were analyzed by oligonucleotide probe hybridization in solution, agarose gel electrophoresis, and autoradiography. The AC-PCR detected as little as 0.053 PFU of cell culture-adapted HAV strain HM175/18f. The results of cDNA-RNA hybridization indicated that the particle/ PFU ratio of this virus strain is approximately 79:1. Therefore, the detection limit of the AC-PCR was estimated to be four virus particles. No amplified products were observed when poliovirus 1, coxsackievirus A9, coxsackievirus B3, echovirus 6, reovirus 1, adenovirus type 40, human rotavirus type 1, and bovine enterovirus type 2 were tested, confirming the specificity of the assay. There were no differences between the nucleotide sequences of AC-PCR products of HAV strain HM175/18f and the sequences of wild-type HAV strain HM175 derived from molecularly cloned cDNA. Of 121 waste and shellfish samples tested by both plaque assays (PA) in cell cultures and the AC-PCR, 81 (67%) were positive and 31 (26%) were negative as determined by both methods, whereas 9 (7%) were positive as determined by the AC-PCR and negative as determined by the PA, and none were positive as determined by the PA and negative as determined by the AC-PCR.</description><identifier>ISSN: 0099-2240</identifier><identifier>EISSN: 1098-5336</identifier><identifier>DOI: 10.1128/aem.60.6.1927-1933.1994</identifier><identifier>PMID: 8031088</identifier><identifier>CODEN: AEMIDF</identifier><language>eng</language><publisher>United States: American Society for Microbiology</publisher><subject>alimentos ; almejas ; amplification chaine polymerase ; Base Sequence ; biodegradacion ; biodegradation ; bivalvia ; bovin ; cattle ; Cellular biology ; cerdo ; clam ; clams ; contaminacion ; contamination ; Effluents ; enterovirus ; estiercol ; farmyard manure ; Food science ; foods ; fumier ; ganado bovino ; Hepatitis ; hepatitis A virus ; Hepatovirus - isolation &amp; purification ; huitre ; immunological techniques ; infeccion ; infection ; Molecular Sequence Data ; ostra ; oysters ; polymerase chain reaction ; Polymerase Chain Reaction - methods ; porcin ; produit alimentaire ; reaccion de cadenas de polimerasa ; Sensitivity and Specificity ; Sewage ; Shellfish ; Shellfish - microbiology ; swine ; technique immunologique ; tecnicas inmunologicas ; Viral Plaque Assay</subject><ispartof>Applied and Environmental Microbiology, 1994-06, Vol.60 (6), p.1927-1933</ispartof><rights>Copyright American Society for Microbiology Jun 1994</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c705t-7d3017a4a087dd01c439aaeacfb61a41b6df30a2914f0d0872f9b98432b7911c3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC201582/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC201582/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,727,780,784,885,3188,3189,27924,27925,53791,53793</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8031088$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Deng, M Y</creatorcontrib><creatorcontrib>Day, S P</creatorcontrib><creatorcontrib>Cliver, D O</creatorcontrib><title>Detection of hepatitis A virus in environmental samples by antigen-capture PCR</title><title>Applied and Environmental Microbiology</title><addtitle>Appl Environ Microbiol</addtitle><description>The efficacy of the antigen-capture PCR (AC-PCR) method for the detection of hepatitis A virus (HAV) in environmental samples was demonstrated. HAV was captured from a seeded liquid waste or a shellfish sample with homologous antibody and then heat denatured and subjected to reverse transcription and the PCR, all in the same tube. Subsequently, the AC-PCR products were analyzed by oligonucleotide probe hybridization in solution, agarose gel electrophoresis, and autoradiography. The AC-PCR detected as little as 0.053 PFU of cell culture-adapted HAV strain HM175/18f. The results of cDNA-RNA hybridization indicated that the particle/ PFU ratio of this virus strain is approximately 79:1. Therefore, the detection limit of the AC-PCR was estimated to be four virus particles. No amplified products were observed when poliovirus 1, coxsackievirus A9, coxsackievirus B3, echovirus 6, reovirus 1, adenovirus type 40, human rotavirus type 1, and bovine enterovirus type 2 were tested, confirming the specificity of the assay. There were no differences between the nucleotide sequences of AC-PCR products of HAV strain HM175/18f and the sequences of wild-type HAV strain HM175 derived from molecularly cloned cDNA. Of 121 waste and shellfish samples tested by both plaque assays (PA) in cell cultures and the AC-PCR, 81 (67%) were positive and 31 (26%) were negative as determined by both methods, whereas 9 (7%) were positive as determined by the AC-PCR and negative as determined by the PA, and none were positive as determined by the PA and negative as determined by the AC-PCR.</description><subject>alimentos</subject><subject>almejas</subject><subject>amplification chaine polymerase</subject><subject>Base Sequence</subject><subject>biodegradacion</subject><subject>biodegradation</subject><subject>bivalvia</subject><subject>bovin</subject><subject>cattle</subject><subject>Cellular biology</subject><subject>cerdo</subject><subject>clam</subject><subject>clams</subject><subject>contaminacion</subject><subject>contamination</subject><subject>Effluents</subject><subject>enterovirus</subject><subject>estiercol</subject><subject>farmyard manure</subject><subject>Food science</subject><subject>foods</subject><subject>fumier</subject><subject>ganado bovino</subject><subject>Hepatitis</subject><subject>hepatitis A virus</subject><subject>Hepatovirus - isolation &amp; purification</subject><subject>huitre</subject><subject>immunological techniques</subject><subject>infeccion</subject><subject>infection</subject><subject>Molecular Sequence Data</subject><subject>ostra</subject><subject>oysters</subject><subject>polymerase chain reaction</subject><subject>Polymerase Chain Reaction - methods</subject><subject>porcin</subject><subject>produit alimentaire</subject><subject>reaccion de cadenas de polimerasa</subject><subject>Sensitivity and Specificity</subject><subject>Sewage</subject><subject>Shellfish</subject><subject>Shellfish - microbiology</subject><subject>swine</subject><subject>technique immunologique</subject><subject>tecnicas inmunologicas</subject><subject>Viral Plaque Assay</subject><issn>0099-2240</issn><issn>1098-5336</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1994</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU9v1DAQxS0EKtvCRwAMB25Zxnbi2AcO1VL-SBUgoGdrkji7rhI7tZOifnu82lVFueCLR3q_Nx7PI-QVgzVjXL1DO64lrOWaaV4XTAuRK10-IisGWhWVEPIxWQFoXXBewlNymtI1AJQg1Qk5USAYKLUiXz_Y2bazC56Gnu7shLObXaLn9NbFJVHnqfW5DH60fsaBJhynwSba3FH0s9taX7Q4zUu09PvmxzPypMch2efH-4xcfbz4tflcXH779GVzflm0NVRzUXcCWI0lgqq7DlhbCo1ose0bybBkjex6Acg1K3voMsR73WhVCt7UmrFWnJH3h77T0oy2a_NsEQczRTdivDMBnXmoeLcz23BrOLBK8ex_e_THcLPYNJvRpdYOA3oblmSYVEryvMX_gkLmU1cZfPMPeB2W6PMS8ptVDonXLEP1AWpjSCna_n5iBmafq8m5GglGmn2uZp-r2eeanS_-_vC97xhk1l8f9J3b7n67aA2m8WG3zLw8MD0Gg9vokrn6mbtXkAdUXIk_rdCz0g</recordid><startdate>19940601</startdate><enddate>19940601</enddate><creator>Deng, M Y</creator><creator>Day, S P</creator><creator>Cliver, D O</creator><general>American Society for Microbiology</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QO</scope><scope>7SN</scope><scope>7SS</scope><scope>7ST</scope><scope>7T7</scope><scope>7TM</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>SOI</scope><scope>7QH</scope><scope>7TV</scope><scope>5PM</scope></search><sort><creationdate>19940601</creationdate><title>Detection of hepatitis A virus in environmental samples by antigen-capture PCR</title><author>Deng, M Y ; Day, S P ; Cliver, D O</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c705t-7d3017a4a087dd01c439aaeacfb61a41b6df30a2914f0d0872f9b98432b7911c3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1994</creationdate><topic>alimentos</topic><topic>almejas</topic><topic>amplification chaine polymerase</topic><topic>Base Sequence</topic><topic>biodegradacion</topic><topic>biodegradation</topic><topic>bivalvia</topic><topic>bovin</topic><topic>cattle</topic><topic>Cellular biology</topic><topic>cerdo</topic><topic>clam</topic><topic>clams</topic><topic>contaminacion</topic><topic>contamination</topic><topic>Effluents</topic><topic>enterovirus</topic><topic>estiercol</topic><topic>farmyard manure</topic><topic>Food science</topic><topic>foods</topic><topic>fumier</topic><topic>ganado bovino</topic><topic>Hepatitis</topic><topic>hepatitis A virus</topic><topic>Hepatovirus - isolation &amp; purification</topic><topic>huitre</topic><topic>immunological techniques</topic><topic>infeccion</topic><topic>infection</topic><topic>Molecular Sequence Data</topic><topic>ostra</topic><topic>oysters</topic><topic>polymerase chain reaction</topic><topic>Polymerase Chain Reaction - methods</topic><topic>porcin</topic><topic>produit alimentaire</topic><topic>reaccion de cadenas de polimerasa</topic><topic>Sensitivity and Specificity</topic><topic>Sewage</topic><topic>Shellfish</topic><topic>Shellfish - microbiology</topic><topic>swine</topic><topic>technique immunologique</topic><topic>tecnicas inmunologicas</topic><topic>Viral Plaque Assay</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Deng, M Y</creatorcontrib><creatorcontrib>Day, S P</creatorcontrib><creatorcontrib>Cliver, D O</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Ecology Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Environment Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>Environment Abstracts</collection><collection>Aqualine</collection><collection>Pollution Abstracts</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Applied and Environmental Microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Deng, M Y</au><au>Day, S P</au><au>Cliver, D O</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Detection of hepatitis A virus in environmental samples by antigen-capture PCR</atitle><jtitle>Applied and Environmental Microbiology</jtitle><addtitle>Appl Environ Microbiol</addtitle><date>1994-06-01</date><risdate>1994</risdate><volume>60</volume><issue>6</issue><spage>1927</spage><epage>1933</epage><pages>1927-1933</pages><issn>0099-2240</issn><eissn>1098-5336</eissn><coden>AEMIDF</coden><abstract>The efficacy of the antigen-capture PCR (AC-PCR) method for the detection of hepatitis A virus (HAV) in environmental samples was demonstrated. HAV was captured from a seeded liquid waste or a shellfish sample with homologous antibody and then heat denatured and subjected to reverse transcription and the PCR, all in the same tube. Subsequently, the AC-PCR products were analyzed by oligonucleotide probe hybridization in solution, agarose gel electrophoresis, and autoradiography. The AC-PCR detected as little as 0.053 PFU of cell culture-adapted HAV strain HM175/18f. The results of cDNA-RNA hybridization indicated that the particle/ PFU ratio of this virus strain is approximately 79:1. Therefore, the detection limit of the AC-PCR was estimated to be four virus particles. No amplified products were observed when poliovirus 1, coxsackievirus A9, coxsackievirus B3, echovirus 6, reovirus 1, adenovirus type 40, human rotavirus type 1, and bovine enterovirus type 2 were tested, confirming the specificity of the assay. There were no differences between the nucleotide sequences of AC-PCR products of HAV strain HM175/18f and the sequences of wild-type HAV strain HM175 derived from molecularly cloned cDNA. Of 121 waste and shellfish samples tested by both plaque assays (PA) in cell cultures and the AC-PCR, 81 (67%) were positive and 31 (26%) were negative as determined by both methods, whereas 9 (7%) were positive as determined by the AC-PCR and negative as determined by the PA, and none were positive as determined by the PA and negative as determined by the AC-PCR.</abstract><cop>United States</cop><pub>American Society for Microbiology</pub><pmid>8031088</pmid><doi>10.1128/aem.60.6.1927-1933.1994</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record>
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ispartof Applied and Environmental Microbiology, 1994-06, Vol.60 (6), p.1927-1933
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source American Society for Microbiology; MEDLINE; PubMed Central (Training); Alma/SFX Local Collection
subjects alimentos
almejas
amplification chaine polymerase
Base Sequence
biodegradacion
biodegradation
bivalvia
bovin
cattle
Cellular biology
cerdo
clam
clams
contaminacion
contamination
Effluents
enterovirus
estiercol
farmyard manure
Food science
foods
fumier
ganado bovino
Hepatitis
hepatitis A virus
Hepatovirus - isolation & purification
huitre
immunological techniques
infeccion
infection
Molecular Sequence Data
ostra
oysters
polymerase chain reaction
Polymerase Chain Reaction - methods
porcin
produit alimentaire
reaccion de cadenas de polimerasa
Sensitivity and Specificity
Sewage
Shellfish
Shellfish - microbiology
swine
technique immunologique
tecnicas inmunologicas
Viral Plaque Assay
title Detection of hepatitis A virus in environmental samples by antigen-capture PCR
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