Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform
Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative pol...
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| Veröffentlicht in: | Food and environmental virology 2022-12, Vol.14 (4), p.364-373 |
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| creator | Donia, Ahmed Furqan Shahid, Muhammad Hassan, Sammer-ul Shahid, Ramla Ahmad, Aftab Javed, Aneela Nawaz, Muhammad Yaqub, Tahir Bokhari, Habib |
| description | Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and RT-LAMP assays were performed on extracted RNA of seven wastewater samples from COVID-19 hotspots. RT‑LAMP assay was also performed on wastewater samples without RNA extraction. Current detection of SARS-CoV-2 is mainly by RT-qPCR of ORF (ORF1ab) and N genes so we targeted both to find the best target gene for SARS-CoV-2 detection. We also performed RT-LAMP with/without RNA extraction inside microfluidic device to target both genes. Positivity rates of RT-qPCR and RT-LAMP performed on extracted RNA were 100.0% (7/7) and 85.7% (6/7), respectively. RT-qPCR results revealed that all 7 wastewater samples were positive for N gene (Ct range 37–39), and negative for ORF1ab, suggesting that N gene could be the best target gene for SARS-CoV-2 detection. RT-LAMP of N and ORF (ORF1a) genes performed on wastewater samples without RNA extraction indicated that all 7 samples remains pink (negative). The color remains pink in all microchannels except microchannels which subjected to RT-LAMP for targeting N region after RNA extraction (yellow color) in 6 out of 7 samples. This study shows that SARS-CoV-2 was successfully detected from wastewater samples using RT-LAMP in microfluidic chips. This study brings the novelty involving the use of wastewater samples for detection of SARS-CoV-2 without previous virus concentration and with/without RNA extraction.
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| doi_str_mv | 10.1007/s12560-022-09522-3 |
| format | Article |
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Graphical abstract</description><identifier>ISSN: 1867-0334</identifier><identifier>ISSN: 1867-0342</identifier><identifier>EISSN: 1867-0342</identifier><identifier>DOI: 10.1007/s12560-022-09522-3</identifier><identifier>PMID: 35508752</identifier><language>eng</language><publisher>New York: Springer US</publisher><subject>Biomedical and Life Sciences ; Biomedicine ; Chemistry/Food Science ; Color ; COVID-19 ; COVID-19 - diagnosis ; COVID-19 Testing ; Disease hot spots ; Food Science ; Gene amplification ; Genes ; Humans ; Lab-on-a-chip ; Microchannels ; Microfluidic devices ; Microfluidics ; N gene ; Original Paper ; Point-of-Care Systems ; Polymerase chain reaction ; Reverse transcription ; Ribonucleic acid ; RNA ; RNA-directed DNA polymerase ; SARS-CoV-2 - genetics ; Sensitivity and Specificity ; Severe acute respiratory syndrome coronavirus 2 ; Target detection ; Virology ; Wastewater ; Water sampling</subject><ispartof>Food and environmental virology, 2022-12, Vol.14 (4), p.364-373</ispartof><rights>The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature 2022</rights><rights>2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.</rights><rights>The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature 2022.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c474t-addbbe99ef4f38ad8414eeca7435a37bcb8b4d08f94dcfc51e0e2a87ddedab113</citedby><cites>FETCH-LOGICAL-c474t-addbbe99ef4f38ad8414eeca7435a37bcb8b4d08f94dcfc51e0e2a87ddedab113</cites><orcidid>0000-0002-2130-3700</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s12560-022-09522-3$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s12560-022-09522-3$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>230,314,780,784,885,27924,27925,41488,42557,51319</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/35508752$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Donia, Ahmed</creatorcontrib><creatorcontrib>Furqan Shahid, Muhammad</creatorcontrib><creatorcontrib>Hassan, Sammer-ul</creatorcontrib><creatorcontrib>Shahid, Ramla</creatorcontrib><creatorcontrib>Ahmad, Aftab</creatorcontrib><creatorcontrib>Javed, Aneela</creatorcontrib><creatorcontrib>Nawaz, Muhammad</creatorcontrib><creatorcontrib>Yaqub, Tahir</creatorcontrib><creatorcontrib>Bokhari, Habib</creatorcontrib><title>Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform</title><title>Food and environmental virology</title><addtitle>Food Environ Virol</addtitle><addtitle>Food Environ Virol</addtitle><description>Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and RT-LAMP assays were performed on extracted RNA of seven wastewater samples from COVID-19 hotspots. RT‑LAMP assay was also performed on wastewater samples without RNA extraction. Current detection of SARS-CoV-2 is mainly by RT-qPCR of ORF (ORF1ab) and N genes so we targeted both to find the best target gene for SARS-CoV-2 detection. We also performed RT-LAMP with/without RNA extraction inside microfluidic device to target both genes. Positivity rates of RT-qPCR and RT-LAMP performed on extracted RNA were 100.0% (7/7) and 85.7% (6/7), respectively. RT-qPCR results revealed that all 7 wastewater samples were positive for N gene (Ct range 37–39), and negative for ORF1ab, suggesting that N gene could be the best target gene for SARS-CoV-2 detection. RT-LAMP of N and ORF (ORF1a) genes performed on wastewater samples without RNA extraction indicated that all 7 samples remains pink (negative). The color remains pink in all microchannels except microchannels which subjected to RT-LAMP for targeting N region after RNA extraction (yellow color) in 6 out of 7 samples. This study shows that SARS-CoV-2 was successfully detected from wastewater samples using RT-LAMP in microfluidic chips. This study brings the novelty involving the use of wastewater samples for detection of SARS-CoV-2 without previous virus concentration and with/without RNA extraction.
Graphical abstract</description><subject>Biomedical and Life Sciences</subject><subject>Biomedicine</subject><subject>Chemistry/Food Science</subject><subject>Color</subject><subject>COVID-19</subject><subject>COVID-19 - diagnosis</subject><subject>COVID-19 Testing</subject><subject>Disease hot spots</subject><subject>Food Science</subject><subject>Gene amplification</subject><subject>Genes</subject><subject>Humans</subject><subject>Lab-on-a-chip</subject><subject>Microchannels</subject><subject>Microfluidic devices</subject><subject>Microfluidics</subject><subject>N gene</subject><subject>Original Paper</subject><subject>Point-of-Care Systems</subject><subject>Polymerase chain reaction</subject><subject>Reverse transcription</subject><subject>Ribonucleic acid</subject><subject>RNA</subject><subject>RNA-directed DNA polymerase</subject><subject>SARS-CoV-2 - genetics</subject><subject>Sensitivity and Specificity</subject><subject>Severe acute respiratory syndrome coronavirus 2</subject><subject>Target detection</subject><subject>Virology</subject><subject>Wastewater</subject><subject>Water sampling</subject><issn>1867-0334</issn><issn>1867-0342</issn><issn>1867-0342</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2022</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kU1vVCEUhm-MxtbqH3BhSNy4Qfm693I3JpPRapNpnLSjLgkXDlOaO1CBadOtv1x02vFj4QZIznMeOLxN85yS15SQ_k2mrO0IJoxhMrR15Q-aQyq7HhMu2MP9mYuD5knOl4R0nLX8cXPA25bIvmWHzfeTUGCddPExoOjQ2QovZqdLpINFp96k6Katt96gFZiLEKe4vkUuJvQOCpj7pvPZ2Tmexy-YIR_QV50L3OgCCelcRWhmr3UwYNEy-lBwdHiuE6DlpEtVbZ42j5yeMjy724-az8fvV_OPePHpw8l8tsBG9KJgbe04wjCAE45LbaWgAsDoXvBW8340oxyFJdINwhpnWgoEmJa9tWD1SCk_at7uvFfbcQPWQChJT-oq-Y1Otypqr_6uBH-h1vFaDaTr5dBVwas7QYrftpCL2vhsYJp0gLjNinUdoUSQQVb05T_oZdymUMdTrOeDaLnshkqxHVX_OecEbv8YStTPiNUuYlUjVr8iVrw2vfhzjH3LfaYV4Dsg11JYQ_p993-0PwD1XrN-</recordid><startdate>20221201</startdate><enddate>20221201</enddate><creator>Donia, Ahmed</creator><creator>Furqan Shahid, Muhammad</creator><creator>Hassan, Sammer-ul</creator><creator>Shahid, Ramla</creator><creator>Ahmad, Aftab</creator><creator>Javed, Aneela</creator><creator>Nawaz, Muhammad</creator><creator>Yaqub, Tahir</creator><creator>Bokhari, Habib</creator><general>Springer US</general><general>Springer Nature B.V</general><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>7X8</scope><scope>5PM</scope><orcidid>https://orcid.org/0000-0002-2130-3700</orcidid></search><sort><creationdate>20221201</creationdate><title>Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform</title><author>Donia, Ahmed ; Furqan Shahid, Muhammad ; Hassan, Sammer-ul ; Shahid, Ramla ; Ahmad, Aftab ; Javed, Aneela ; Nawaz, Muhammad ; Yaqub, Tahir ; Bokhari, Habib</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c474t-addbbe99ef4f38ad8414eeca7435a37bcb8b4d08f94dcfc51e0e2a87ddedab113</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2022</creationdate><topic>Biomedical and Life Sciences</topic><topic>Biomedicine</topic><topic>Chemistry/Food Science</topic><topic>Color</topic><topic>COVID-19</topic><topic>COVID-19 - diagnosis</topic><topic>COVID-19 Testing</topic><topic>Disease hot spots</topic><topic>Food Science</topic><topic>Gene amplification</topic><topic>Genes</topic><topic>Humans</topic><topic>Lab-on-a-chip</topic><topic>Microchannels</topic><topic>Microfluidic devices</topic><topic>Microfluidics</topic><topic>N gene</topic><topic>Original Paper</topic><topic>Point-of-Care Systems</topic><topic>Polymerase chain reaction</topic><topic>Reverse transcription</topic><topic>Ribonucleic acid</topic><topic>RNA</topic><topic>RNA-directed DNA polymerase</topic><topic>SARS-CoV-2 - genetics</topic><topic>Sensitivity and Specificity</topic><topic>Severe acute respiratory syndrome coronavirus 2</topic><topic>Target detection</topic><topic>Virology</topic><topic>Wastewater</topic><topic>Water sampling</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Donia, Ahmed</creatorcontrib><creatorcontrib>Furqan Shahid, Muhammad</creatorcontrib><creatorcontrib>Hassan, Sammer-ul</creatorcontrib><creatorcontrib>Shahid, Ramla</creatorcontrib><creatorcontrib>Ahmad, Aftab</creatorcontrib><creatorcontrib>Javed, Aneela</creatorcontrib><creatorcontrib>Nawaz, Muhammad</creatorcontrib><creatorcontrib>Yaqub, Tahir</creatorcontrib><creatorcontrib>Bokhari, Habib</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Food and environmental virology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Donia, Ahmed</au><au>Furqan Shahid, Muhammad</au><au>Hassan, Sammer-ul</au><au>Shahid, Ramla</au><au>Ahmad, Aftab</au><au>Javed, Aneela</au><au>Nawaz, Muhammad</au><au>Yaqub, Tahir</au><au>Bokhari, Habib</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform</atitle><jtitle>Food and environmental virology</jtitle><stitle>Food Environ Virol</stitle><addtitle>Food Environ Virol</addtitle><date>2022-12-01</date><risdate>2022</risdate><volume>14</volume><issue>4</issue><spage>364</spage><epage>373</epage><pages>364-373</pages><issn>1867-0334</issn><issn>1867-0342</issn><eissn>1867-0342</eissn><abstract>Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and RT-LAMP assays were performed on extracted RNA of seven wastewater samples from COVID-19 hotspots. RT‑LAMP assay was also performed on wastewater samples without RNA extraction. Current detection of SARS-CoV-2 is mainly by RT-qPCR of ORF (ORF1ab) and N genes so we targeted both to find the best target gene for SARS-CoV-2 detection. We also performed RT-LAMP with/without RNA extraction inside microfluidic device to target both genes. Positivity rates of RT-qPCR and RT-LAMP performed on extracted RNA were 100.0% (7/7) and 85.7% (6/7), respectively. RT-qPCR results revealed that all 7 wastewater samples were positive for N gene (Ct range 37–39), and negative for ORF1ab, suggesting that N gene could be the best target gene for SARS-CoV-2 detection. RT-LAMP of N and ORF (ORF1a) genes performed on wastewater samples without RNA extraction indicated that all 7 samples remains pink (negative). The color remains pink in all microchannels except microchannels which subjected to RT-LAMP for targeting N region after RNA extraction (yellow color) in 6 out of 7 samples. This study shows that SARS-CoV-2 was successfully detected from wastewater samples using RT-LAMP in microfluidic chips. This study brings the novelty involving the use of wastewater samples for detection of SARS-CoV-2 without previous virus concentration and with/without RNA extraction.
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| subjects | Biomedical and Life Sciences Biomedicine Chemistry/Food Science Color COVID-19 COVID-19 - diagnosis COVID-19 Testing Disease hot spots Food Science Gene amplification Genes Humans Lab-on-a-chip Microchannels Microfluidic devices Microfluidics N gene Original Paper Point-of-Care Systems Polymerase chain reaction Reverse transcription Ribonucleic acid RNA RNA-directed DNA polymerase SARS-CoV-2 - genetics Sensitivity and Specificity Severe acute respiratory syndrome coronavirus 2 Target detection Virology Wastewater Water sampling |
| title | Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform |
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