Development of a new method for diagnosis of Group B Coxsackie genome by reverse transcription loop-mediated isothermal amplification

Jaianand, K.; Saravanan, Nirmala; Gunasekaran, Palani; Sheriff, A Khaleefathullah

doi:10.4103/0255-0857.81780

Cited by 6 publications

(7 citation statements)

References 17 publications

(18 reference statements)

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“…The sensitivity of the conventional RT-PCR has been established at approximately 1 to100 PFU of virus per gram of tissue (Shen et al, 2009), the sensitivity of NASBA similar to RT-PCR (Saeedinia et al, 2008), and the sensitivity of the real-time PCR assay have been reported 100 copies per reaction for CVB3 (Dierssen et al, 2008). Most researchers that have developed RT-LAMP method for detection of Enteroviruses have shown that sensitivity of the RT-LAMP was approximately 10 copies per reaction that was 10-fold higher sensitive than RT-PCR (Jaianand et al, 2010;Yaqing et al, 2012;Zhao et al, 2014).The RT-LAMP assay developed in this work and comparative analysis of RT-LAMP and RT-PCR sensitivity suggested that RT-LAMP was 100-fold higher sensitive than RT-PCR. Also due to the designing four primers that recognize six specific areas in target sequence, RT-LAMP assay has a high specificity, therefore cross-reactivity with other Enterovirus were not observed.…”

Section: Discussionmentioning

confidence: 72%

“…Also, reverse transcription LAMP (RT-LAMP) assay which is carried out in a single tube is a simple, high specific, rapid, and cost-effectiveness method . Less time-consuming of RT-LAMP than conventional PCR-based methods has been used successfully for rapid detection of pathogenic RNA viruses such as Enteroviruses (Arita et al, 2009;Jaianand et al, 2010Jaianand et al, , 2011Shi et al, 2011;Nie et al, 2012;Yaqing et al, 2012;Zhang et al, 2012;Zhao et al, 2013;Chen et al, 2014;Ding et al, 2014;Jiang et al, 2014;Zhao et al, 2014;Wang et al, 2014) and less prone to inhibit from impurity of template sample (Caipang et al, 2004;Kaneko et al, 2007;Tomita et al, 2008;Mori and Notomi, 2009;Ghosh et al, 2015). The RT-LAMP assay is very specific in compared to other molecular detection methods, because in this method four primers are used to recognize six specific regions of the target gene for amplification .…”

Section: Introductionmentioning

confidence: 99%

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Evaluation of a rapid detection for Coxsackievirus B3 using one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP)

Monazah

Zeinoddini

Saeeidinia

2017

Journal of Virological Methods

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Coxsackievirus B3 (CVB3) is a member of the genus Enterovirus within the family Picornaviridae and is an important pathogen of viral myocarditis, which accounts for more than 50% viral myocarditis cases. VP1 is major capsid protein that this region has a low homology in both amino acid and nucleotide sequences among Enteroviruses. Therefore we have chosen this region for designed a set of RT-LAMP primers for CVB3 detection. For this the total RNA was extracted from 24-h post infected-HeLa cells with complete cytopathic effect (CPE), and applied to a one-step reverse transcription loop-mediated isothermal amplification reaction (RT-LAMP) using CVB3-specific primers. The optimization of RT-LAMP reaction was carried out with three variables factors including MgSO concentration, temperature and time of incubation. Amplification was analyzed by using 2% agarose gel electrophoresis and ethidium bromide and SYBR Green staining. Our results were shown the ladder-like pattern of the VP1 gene amplification. The LAMP reaction mix was optimized and the best result observed at 4mM MgSO and 60°C for 90min incubation. RT-LAMP had high sensitivity and specificity for detection of CVB3 infection. This method can be used as a rapid and easy diagnostic test for detection of CVB3 in clinical laboratories.

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Section: Discussionmentioning

confidence: 72%

Section: Introductionmentioning

confidence: 99%

Evaluation of a rapid detection for Coxsackievirus B3 using one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP)

Monazah

Zeinoddini

Saeeidinia

2017

Journal of Virological Methods

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“…), one detecting group B CV (Jaianand et al . ). Only one RT‐LAMP assay was established for detection of CVA16 which used hydroxynaphthol blue (HNB) dye mediated visualization (Nie et al .…”

Section: Resultsmentioning

confidence: 97%

Rapid and visual detection of human enterovirus coxsackievirus A16 by reverse transcription loop-mediated isothermal amplification combined with lateral flow device

Guo

Liu

Zhao

et al. 2015

Lett Appl Microbiol

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Coxsackievirus A16 (CVA16) is one of the major causative agents of hand, foot and mouth disease (HFMD). Rapid and reliable detection and typing of it can limit the spread. We developed a reverse transcription loop-mediated isothermal amplification (RT-LAMP) combined with lateral flow device (LFD) technology to rapidly detect CVA16. The high sensitivity and specificity and its ease of use make this assay ideal for use in resource-limited settings such as primary care facilities and clinical laboratories in developing countries.

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“…Notomi et al (2000) first described an autocycling strand displacement DNA synthesis using the Bst DNA polymerase, the LAMP method is carried out widely in diagnostic clinical laboratories for detection of bacteria and viruses (12). The LAMP assay as a rapid and cost-effective method does not require thermocycler and gel electrophoresis in some cases (using turbidity) and the inhibitory effects of substances in samples are largely eliminated (4,13). A reliable LAMP mostly depends on the specificity of the primer sets (4).…”

Section: Loop-mediated Isothermal Amplification (Lamp) For …mentioning

confidence: 99%

Loop-Mediated Isothermal Amplification (LAMP) for the Rapid Diagnosis of Herpes Simplex Virus Type 1 (HSV-1)

Pourhossein

Soleimanjahi

Behzadian³

et al. 2011

Iran J Virol

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Background and Aims: considering difficulties in usual laboratory methods in detection of viral infections, improved DNA-based diagnostic techniques are more reliable. Loop mediated isothermal amplification method (LAMP) is a nucleic acid amplification method that amplifies DNA using six primers which has been developed to diagnose viruses as a rapid and high efficiency test. In this study, the LAMP was used for detection of Herpes simplex virus. Methods: The set of primers were designed, for detection of HSV-based on gG fragment. The genome of the virus was extracted from the culture supernatant of infected cells. LAMP technique was optimized in term of temperature, time and the ingredients used for test. For detection of HSV-1 infection, sensitivity and specificity of this technique were determined by various dilutions of virus and infected samples with HSV-2 that was taken from Day Hospital of Tehran. Results: The sensitivity and specificity of HSV-1 specific LAMP method, reached about 500 copies/tube and 99.9% respectively. Furthermore, both the agarose gel electrophoresis containing Ethidium Bromide (EtBr) and the turbidity assay directly detected HSV-1 virus LAMP products in reactions. Conclusion: In this study, the reliability of LAMP for detection of HSV-was approved; therefore this rapid, accurate, and cost-effective detection and quantification method may perhaps be an investigative tool which can be valid for detection of target viral genome in clinical specimens in the absence of the necessary facilities for performing PCR.

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Development of a new method for diagnosis of Group B Coxsackie genome by reverse transcription loop-mediated isothermal amplification

Cited by 6 publications

References 17 publications

Evaluation of a rapid detection for Coxsackievirus B3 using one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP)

Evaluation of a rapid detection for Coxsackievirus B3 using one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP)

Rapid and visual detection of human enterovirus coxsackievirus A16 by reverse transcription loop-mediated isothermal amplification combined with lateral flow device

Loop-Mediated Isothermal Amplification (LAMP) for the Rapid Diagnosis of Herpes Simplex Virus Type 1 (HSV-1)

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