Loop-mediated isothermal amplification for rapid and sensitive detection of nervous necrosis virus (NNV)

Abstract

A loop-mediated isothermal amplification (LAMP) of DNA is currently one of the most commonly used molecular diagnostic tools due to its simple, fast and easy to amplify target DNA under isothermal conditions. In the present study, a LAMP test was designed from the coat protein gene of Nervous necrosis virus (NNV) which has been responsible for some of the most explosive epidemics of emerging viral diseases in many Asian countries, resulting in huge economical losses on a fish farm. Using a set of the synthesized primers matching the specific region of the NNV gene sequences provided by NCBI database, not originated from NNV-infected fish, we observed that the efficiency and specificity of LAMP were mainly dependent on the concentration of DNA polymerase and reaction temperature, suggesting that they act as main determinants in LAMP assay. In particular, we demonstrated that they were even applied to detect NNV infection in red sea bream. These data suggest that the development of LAMP primers based on genetic information provided by public database, not virus-infected samples, may provide very simple and convenient method to determine viral infection in aquatic organism.st was designed from the coat protein gene of Nervous necrosis virus (NNV) which has been responsible for some of the most explosive epidemics of emerging viral diseases in many Asian countries, resulting in huge economical losses on a fish farm. Using a set of the synthesized primers matching the specific region of the NNV gene sequences provided by NCBI database, not originated from NNV-infected fish, we observed that the efficiency and specificity of LAMP were mainly dependent on the concentration of DNA polymerase and reaction temperature, suggesting that they act as main determinants in LAMP assay. In particular, we demonstrated that they were even applied to detect NNV infection in red sea bream. These data suggest that the development of LAMP primers based on genetic information provided by public database, not virus-infected samples, may provide very simple and convenient method to determine viral infection in aquatic organism.