The development and application of a Dot-ELISA assay for diagnosis of southern rice black-streaked dwarf disease in the field.

Zhenchao Wang,Jiaju Liu,Mengjiao Zeng,Baoan Song,Song Yang,Dandan Yu,Linhong Jin,Zhuo Chen,Deyu Hu,Xiangyang Li,Liang Bi

doi:10.3390/v4010167

Zhenchao Wang, Jiaju Liu + Show 9 more

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https://doi.org/10.3390/v4010167

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Journal: Viruses	Publication Date: Jan 23, 2012
Citations: 50	License type: CC BY 4.0

Affiliation: Guizhou University, South China University of Technology

Abstract

Outbreaks of the southern rice black-streaked dwarf virus (SRBSDV) have caused significant crop losses in southern China in recent years, especially in 2010. There are no effective, quick and practicable methods for the diagnosis of rice dwarf disease that can be used in the field. Traditional reverse transcription-polymerase chain reaction (RT-PCR) methodology is accurate but requires expensive reagents and instruments, as well as complex procedures that limit its applicability for field tests. To develop a sensitive and reliable assay for routine laboratory diagnosis, a rapid dot enzyme-linked immunosorbent assay (dot-ELISA) method was developed for testing rice plants infected by SRBSDV. Based on anti-SRBSDV rabbit antiserum, this new dot-ELISA was highly reliable, sensitive and specific toward SRBSDV. The accuracy of two blotting media, polyvinylidene fluoride membrane (PVDF membrane) and nitrocellulose filter membrane (NC membrane), was compared. In order to facilitate the on-site diagnosis, three county laboratories were established in Shidian (Yunnan province), Jianghua (Hunan Province) and Libo (Guizhou province). Suspected rice cases from Shidian, Yuanjiang and Malipo in Yunnan province were tested and some determined to be positive for SRBSDV by the dot-ELISA and confirmed by the One Step RT-PCR method. To date, hundreds of suspected rice samples collected from 61 districts in southwestern China have been tested, among which 55 districts were found to have rice crops infected by SRBSDV. Furthermore, the test results in the county laboratories showed that Libo, Dehong (suspected samples were sent to Shidian) and Jianghua were experiencing a current SRBSDV outbreak.

Highlights

Southern rice black-streaked dwarf virus (SRBSDV) is currently one of the most damaging rice crop diseases in China
The southern rice black-streaked dwarf virus (SRBSDV) was identified in 2008, detection of infection has depended on reverse transcription-polymerase chain reaction (RT-PCR) methods
Economic losses from SRBSDV outbreak were so serious that the development of a new rapid SRBSDV diagnostic method to detect the virus at the early stages of infection was urgently needed to allow sufficient time for preventative measures to avoid the spread of the virus

Summary

Introduction

Southern rice black-streaked dwarf virus (SRBSDV) is currently one of the most damaging rice crop diseases in China. The SRBSDV is a novel member of Fijivirus (family Reoviridae) [1,2,3], first observed in Guangdong province in 2001 and identified in 2008. Wang reported the complete SRBSDV nucleotide sequences of isolates from Hainan and Guangdong provinces of China, and confirmed that it is a novel Fijivirus species [3]. SRBSDV has caused extensive crop damage in Jiangxi, Hunan, Guangdong, Fujian, Hainan, Guizhou, and Yunnan since 2009 in China. 5 million acres of rice had been infected by this virus in China and 100,000 acres of rice farm reported crop failure in 2009 [11]. In 2010, nearly 3 million acres of rice were infected by SRBSDV, suggesting a rapid spread and major losses in the coming years [12]

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