Development and application of quantitative detection method for nervous necrosis virus (NNV) isolated from sevenband grouper Hyporthodus septemfasciatus

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• 作者：Jong-Oh Kim ^{jongoh.kim77@gmail.com" class="auth_mail" title="E-mail the corresponding author} ; Jae-Ok Kim ; Wi-Sik Kim ; Myung-Joo Oh ; ^{ohmj@jnu.ac.kr" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Nervous necrosis virus (NNV) ; Quantitative detection ; Diagnostic ; Sevenband grouper
• 刊名：Asian Pacific Journal of Tropical Medicine
• 出版年：2016
• 出版时间：August 2016
• 年：2016
• 卷：9
• 期：8
• 页码：742-748
• 全文大小：1199 K

文摘

To develop the rapid and efficient quantitative detection tool for nervous necrosis virus isolated from sevenband grouper Hyporhodus septemfasciatus.

Methods

The viral genes of the NNV (SGYeosu08) isolated from sevenband grouper were phylogenetically analyzed. In addition, novel quantitative PCR primers based on the genomic sequence of SGYeosu08 isolate were designed and compared it with the conventional bio-assay method (TCID₅₀) using in vitro and in vivo samples.

Results

The phylogenetic analysis of viral genes demonstrated the relationship of SGYeosu08 with members of red-spotted grouper nervous necrosis virus (RGNNV). The qNNV_R1 primer set (R1_F and R1_R) and the qNNV_R2 primer set (R2_F and R2_R) revealed 93% primer efficiency (regression: y = −0.2861x + 9.9401, R² = 0.9976) and the revealed 108% primer efficiency (regression: y = −0.3172x + 10.0611, R² = 0.9982), respectively. Its comparison with viral infectivity calculated by TCID₅₀ method showed similar kinetic pattern at in vitro and NNV challenged fish (in vivo) samples.

Conclusions

Result show that this method is rapid and efficient to diagnose NNV infection compare to traditional bioassay method (TCID₅₀).