Nucleic-Acid Based Techniques for the Fine Diagnosis of Plant Viruses in India

Abstract

Field and horticultural crops are infected by large number of viruses and virus-like pathogens in India, which can be managed only by preventive measures and for that sensitive diagnosis is a prerequisite. Bioassay was initially utilized in detection of plant viruses. Following this, serological assay enzyme-linked immunosorbent assay (ELISA) was widely employed. In the last two decades, nucleic acid based detection assays were developed for the correct and fine diagnosis of plant viruses in India. Nucleic acid based detection assays in addition to providing sufficient sensitivity, specificity and accuracy are also robust and easily applicable. Correct identity of a large number of viruses which were misdiagnosed earlier was now diagnosed to their correct taxonomic status using nucleic acid based assays. Most commonly used nucleic acid based detection assays employed in India are nucleic acid spot hybridization (NASH), polymerase chain reaction (PCR) and reverse transcription-polymerase chain reaction (RT-PCR). These methods have been utilized for detection of large number of viruses and virus-like pathogens. Multiplex detection assays (multiplex PCR and multiplex–RT-PCR) were also developed for simultaneous detection of more than one virus in same reaction tube. Isothermal detection assay where amplification reaction can be carried out at constant temperature is developed and employed for simple and robust detection as well as characterization of plant viruses in India. The most commonly employed isothermal detection assays includes the rolling circle amplification (RCA) which has been employed to detect DNA viruses of the genera Begomovirus, Babuvirus and Badnavirus. Loop-mediated isothermal amplification (LAMP) are latest in the application of isothermal detection methods, which provides simple and easy detection of plant viruses and such detection assays have been developed for some viruses in India. The next generation sequencing (NGS) or deep sequencing, which involves massive parallel sequencing approach followed by bioinformatics analysis, has revolutionized the field of novel virus discovery in India. This chapter aims to highlight the application of various nucleic acid based detection assays in diagnosis and characterization of viruses infecting different crops in India. In this chapter, a crop-wise review of the application of different nucleic acid based detection methods in India has been presented.