Abstract
Plants are frequently infected by a wide range of viruses that cause important agronomic, economic and social impact. Detection of viruses at early stages of infection is crucial to reduce economic losses. Biological indexing and serological enzyme-linked immunosorbent assay (ELISA) methods are widely used for diagnosis. Nevertheless, molecular techniques have revolutionized plant virus detection and identification. Early molecular hybridization technologies were rapidly supplanted by more powerful nucleic acids amplification methods based on the polymerase chain reaction (PCR). Although molecular methods are highly discriminatory, allowing strain typing, routine testing has been hampered by problems in reproducibility. Continuous efforts have been made to overcome these barriers. Improved systems to prepare plant or insect samples have been developed. Efforts have also been directed at increasing the sensitivity and specificity of detection, which can be limited by the high content of enzyme inhibitors in plant materials. Nested and multiplex PCR offer high sensitivity and the possibility to detect several targets in one assay, respectively. Other technologies allow the amplification of nucleic acids in an isothermal reaction (nucleic acid sequence-based amplification [NASBA] or reverse transcription loop-mediated isothermal amplification [RT-LAMP] procedures). High-throughput testing has been achieved by real-time PCR, which allows the automation of PCR combined with fluorimetry. Real-time PCR simultaneously permits detection and quantitation of targets. The use of integrated protocols combining the specificity of serological techniques with the sensitivity of molecular methods will increase the accuracy and reliability of virus diagnostic. In the future, nucleic acid arrays and biosensors assisted by nanotechnology could revolutionize the detection of plant viruses.