Development and Evaluation of Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Rapid and Real-Time Detection of the Swine-Origin Influenza A H1N1 Virus
Identifieur interne : 000C57 ( Main/Exploration ); précédent : 000C56; suivant : 000C58Development and Evaluation of Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Rapid and Real-Time Detection of the Swine-Origin Influenza A H1N1 Virus
Auteurs : Manmohan Parida [Inde] ; Jyoti Shukla [Inde] ; Shashi Sharma [Inde] ; Sanna Ranghia Santhosh [Inde] ; Vasanthapuram Ravi [Inde] ; Reeta Mani [Inde] ; Maria Thomas [Inde] ; Shashi Khare [Inde] ; Arvind Rai [Inde] ; Radha Kant Ratho [Inde] ; Sujit Pujari [Inde] ; Bijayanti Mishra [Inde] ; Putcha Venkata Lakshmana Rao [Inde] ; Rajagopalan Vijayaraghavan [Inde]Source :
- The Journal of Molecular Diagnostics : JMD [ 1525-1578 ] ; 2011.
Abstract
The recent emergence of the swine-origin influenza A H1N1 virus (S-OIV) poses a serious global health threat. Rapid detection and differentiation of S-OIV from seasonal influenza is crucial for patient management and control of the epidemics. A one-step, single-tube accelerated and quantitative S-OIV-specific H1 reverse transcription loop-mediated isothermal amplification (RTLAMP) assay for clinical diagnosis of S-OIV by targeting the H1 gene is reported in this article. A comparative evaluation of the H1-specific RTLAMP assay vis-à-vis the World Health Organization-approved real-time polymerase chain reaction (RTPCR), involving 239 acute-phase throat swab samples, demonstrated exceptionally higher sensitivity by picking up all of the 116 H1N1-positive cases and 36 additional positive cases among the negatives that were sequence-confirmed as S-OIV H1N1. None of the real-time RTPCR-positive samples were missed by the RTLAMP system. The comparative analysis revealed that S-OIV RTLAMP was up to tenfold more sensitive than the World Health Organization real-time RTPCR; it had a detection limit of 0.1 tissue culture infectious dosage of 50/ml. One of the most attractive features of this isothermal gene amplification assay is that it seems to have an advantage in monitoring gene amplification by means of SYBR Green I dye-mediated naked-eye visualization within 30 minutes compared to 2 to 3 hours for a real-time reverse transcription polymerase chain reaction. This suggests that the RTLAMP assay is a valuable tool for rapid, real-time detection and quantification of S-OIV in acute-phase throat swab samples without requiring sophisticated equipment.
Url:
DOI: 10.1016/j.jmoldx.2010.11.003
PubMed: 21227400
PubMed Central: 3069812
Affiliations:
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Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author><author><name sortKey="Shukla, Jyoti" sort="Shukla, Jyoti" uniqKey="Shukla J" first="Jyoti" last="Shukla">Jyoti Shukla</name><affiliation wicri:level="1"><nlm:aff id="aff1">Division of Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author><author><name sortKey="Sharma, Shashi" sort="Sharma, Shashi" uniqKey="Sharma S" first="Shashi" last="Sharma">Shashi Sharma</name><affiliation wicri:level="1"><nlm:aff id="aff1">Division of Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author><author><name sortKey="Ranghia Santhosh, Sanna" sort="Ranghia Santhosh, Sanna" uniqKey="Ranghia Santhosh S" first="Sanna" last="Ranghia Santhosh">Sanna Ranghia Santhosh</name><affiliation wicri:level="1"><nlm:aff id="aff1">Division of Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author><author><name sortKey="Ravi, Vasanthapuram" sort="Ravi, Vasanthapuram" uniqKey="Ravi V" first="Vasanthapuram" last="Ravi">Vasanthapuram Ravi</name><affiliation wicri:level="1"><nlm:aff id="aff2">Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore</wicri:regionArea><wicri:noRegion>Bangalore</wicri:noRegion></affiliation></author><author><name sortKey="Mani, Reeta" sort="Mani, Reeta" uniqKey="Mani R" first="Reeta" last="Mani">Reeta Mani</name><affiliation wicri:level="1"><nlm:aff id="aff2">Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore</wicri:regionArea><wicri:noRegion>Bangalore</wicri:noRegion></affiliation></author><author><name sortKey="Thomas, Maria" sort="Thomas, Maria" uniqKey="Thomas M" first="Maria" last="Thomas">Maria Thomas</name><affiliation wicri:level="1"><nlm:aff id="aff2">Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, National Institute of Mental Health and Neuro Sciences, Bangalore</wicri:regionArea><wicri:noRegion>Bangalore</wicri:noRegion></affiliation></author><author><name sortKey="Khare, Shashi" sort="Khare, Shashi" uniqKey="Khare S" first="Shashi" last="Khare">Shashi Khare</name><affiliation wicri:level="1"><nlm:aff id="aff3">Department of Virology, National Centre for Disease Control, Delhi, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, National Centre for Disease Control, Delhi</wicri:regionArea><wicri:noRegion>Delhi</wicri:noRegion></affiliation></author><author><name sortKey="Rai, Arvind" sort="Rai, Arvind" uniqKey="Rai A" first="Arvind" last="Rai">Arvind Rai</name><affiliation wicri:level="1"><nlm:aff id="aff3">Department of Virology, National Centre for Disease Control, Delhi, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, National Centre for Disease Control, Delhi</wicri:regionArea><wicri:noRegion>Delhi</wicri:noRegion></affiliation></author><author><name sortKey="Kant Ratho, Radha" sort="Kant Ratho, Radha" uniqKey="Kant Ratho R" first="Radha" last="Kant Ratho">Radha Kant Ratho</name><affiliation wicri:level="1"><nlm:aff id="aff4">Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh</wicri:regionArea><wicri:noRegion>Chandigarh</wicri:noRegion></affiliation></author><author><name sortKey="Pujari, Sujit" sort="Pujari, Sujit" uniqKey="Pujari S" first="Sujit" last="Pujari">Sujit Pujari</name><affiliation wicri:level="1"><nlm:aff id="aff4">Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh</wicri:regionArea><wicri:noRegion>Chandigarh</wicri:noRegion></affiliation></author><author><name sortKey="Mishra, Bijayanti" sort="Mishra, Bijayanti" uniqKey="Mishra B" first="Bijayanti" last="Mishra">Bijayanti Mishra</name><affiliation wicri:level="1"><nlm:aff id="aff4">Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Department of Virology, Post Graduate Institute of Medical Education and Research, Chandigarh</wicri:regionArea><wicri:noRegion>Chandigarh</wicri:noRegion></affiliation></author><author><name sortKey="Lakshmana Rao, Putcha Venkata" sort="Lakshmana Rao, Putcha Venkata" uniqKey="Lakshmana Rao P" first="Putcha Venkata" last="Lakshmana Rao">Putcha Venkata Lakshmana Rao</name><affiliation wicri:level="1"><nlm:aff id="aff1">Division of Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author><author><name sortKey="Vijayaraghavan, Rajagopalan" sort="Vijayaraghavan, Rajagopalan" uniqKey="Vijayaraghavan R" first="Rajagopalan" last="Vijayaraghavan">Rajagopalan Vijayaraghavan</name><affiliation wicri:level="1"><nlm:aff id="aff1">Division of Virology, Defence Research & Development Establishment, Gwalior, India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, Defence Research & Development Establishment, Gwalior</wicri:regionArea><wicri:noRegion>Gwalior</wicri:noRegion></affiliation></author></analytic><series><title level="j">The Journal of Molecular Diagnostics : JMD</title><idno type="ISSN">1525-1578</idno><idno type="eISSN">1943-7811</idno><imprint><date when="2011">2011</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>The recent emergence of the swine-origin influenza A H1N1 virus (S-OIV) poses a serious global health threat. Rapid detection and differentiation of S-OIV from seasonal influenza is crucial for patient management and control of the epidemics. A one-step, single-tube accelerated and quantitative S-OIV-specific H1 reverse transcription loop-mediated isothermal amplification (RTLAMP) assay for clinical diagnosis of S-OIV by targeting the H1 gene is reported in this article. A comparative evaluation of the H1-specific RTLAMP assay vis-à-vis the World Health Organization-approved real-time polymerase chain reaction (RTPCR), involving 239 acute-phase throat swab samples, demonstrated exceptionally higher sensitivity by picking up all of the 116 H1N1-positive cases and 36 additional positive cases among the negatives that were sequence-confirmed as S-OIV H1N1. None of the real-time RTPCR-positive samples were missed by the RTLAMP system. The comparative analysis revealed that S-OIV RTLAMP was up to tenfold more sensitive than the World Health Organization real-time RTPCR; it had a detection limit of 0.1 tissue culture infectious dosage of <sub>50</sub>/ml. One of the most attractive features of this isothermal gene amplification assay is that it seems to have an advantage in monitoring gene amplification by means of SYBR Green I dye-mediated naked-eye visualization within 30 minutes compared to 2 to 3 hours for a real-time reverse transcription polymerase chain reaction. This suggests that the RTLAMP assay is a valuable tool for rapid, real-time detection and quantification of S-OIV in acute-phase throat swab samples without requiring sophisticated equipment.</p></div></front></TEI><affiliations><list><country><li>Inde</li></country></list><tree><country name="Inde"><noRegion><name sortKey="Parida, Manmohan" sort="Parida, Manmohan" uniqKey="Parida M" first="Manmohan" last="Parida">Manmohan Parida</name></noRegion><name sortKey="Kant Ratho, Radha" sort="Kant Ratho, Radha" uniqKey="Kant Ratho R" first="Radha" last="Kant Ratho">Radha Kant Ratho</name><name sortKey="Khare, Shashi" sort="Khare, Shashi" uniqKey="Khare S" first="Shashi" last="Khare">Shashi Khare</name><name sortKey="Lakshmana Rao, Putcha Venkata" sort="Lakshmana Rao, Putcha Venkata" uniqKey="Lakshmana Rao P" first="Putcha Venkata" last="Lakshmana Rao">Putcha Venkata Lakshmana Rao</name><name sortKey="Mani, Reeta" sort="Mani, Reeta" uniqKey="Mani R" first="Reeta" last="Mani">Reeta Mani</name><name sortKey="Mishra, Bijayanti" sort="Mishra, Bijayanti" uniqKey="Mishra B" first="Bijayanti" last="Mishra">Bijayanti Mishra</name><name sortKey="Pujari, Sujit" sort="Pujari, Sujit" uniqKey="Pujari S" first="Sujit" last="Pujari">Sujit Pujari</name><name sortKey="Rai, Arvind" sort="Rai, Arvind" uniqKey="Rai A" first="Arvind" last="Rai">Arvind Rai</name><name sortKey="Ranghia Santhosh, Sanna" sort="Ranghia Santhosh, Sanna" uniqKey="Ranghia Santhosh S" first="Sanna" last="Ranghia Santhosh">Sanna Ranghia Santhosh</name><name sortKey="Ravi, Vasanthapuram" sort="Ravi, Vasanthapuram" uniqKey="Ravi V" first="Vasanthapuram" last="Ravi">Vasanthapuram Ravi</name><name sortKey="Sharma, Shashi" sort="Sharma, Shashi" uniqKey="Sharma S" first="Shashi" last="Sharma">Shashi Sharma</name><name sortKey="Shukla, Jyoti" sort="Shukla, Jyoti" uniqKey="Shukla J" first="Jyoti" last="Shukla">Jyoti Shukla</name><name sortKey="Thomas, Maria" sort="Thomas, Maria" uniqKey="Thomas M" first="Maria" last="Thomas">Maria Thomas</name><name sortKey="Vijayaraghavan, Rajagopalan" sort="Vijayaraghavan, Rajagopalan" uniqKey="Vijayaraghavan R" first="Rajagopalan" last="Vijayaraghavan">Rajagopalan Vijayaraghavan</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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