The viral pathogens that infect crop plants constrain food production and economic development throughout the world's agricultural regions. Viral diseases are difficult to prevent, and once established few means are available to counter their impact on yield. As a result, development and deployment of resistance crop varieties remains the most effective manner in which to combat the evolving threats presented by plant viral diseases. Underpinning such efforts is the need for robust diagnostic capacities to identify the species and strains of viral pathogens infecting crop plants and their related wild species, and to understand their distribution within a given geographical region.
Access to simple, low cost tools for the molecular study of plant viral pathogens is central to generating the knowledge and improved germplasm required by scientists, breeders and farmers to combat these diseases and maximize crop yields. Effective methods for sampling, storage and retrieval of viral pathogens from infected plant tissues allows not only identification of the viral pathogens but also detailed molecular study of their genomes, generating increased understanding of their epidemiology, etiology and evolution. Diagnostic technologies are also required for virus indexing to facilitate certification of pathogen-free materials for the collection, maintenance and international exchange of the elite germplasm on which required plant improvement programs are based.
Molecular characterization of the viruses that infect plant material is currently achieved by direct electrophoretical isolation from total nucleic acid, followed by cloning and subsequent analysis, or amplification of full or partial genomic sequences by polymerase chain reaction (PCR). PCR is the more powerful technique due to its ability to recover viral sequences and whole genome components from very low viral titres, and is now the preferred approach for most applications. Currently, total viral and genomic nucleic acids are isolated from infected tissues by methods such as Dellaporta et al.  which involve multi-step protocols for DNA or RNA extraction, precipitation and purification. A frequent limitation for studying viruses at the molecular level is the ability to reliably obtain high quality nucleic acids from putatively infected plant material. Plant tissues to be analyzed must be collected and preserved in order to maintain integrity of the nucleic acids until they can be processed. This poses challenges when sample numbers are large and when working in the field, most especially in the tropical and sub-tropical regions where plant viral pathogens are abundant. Field studies are thus constrained by the resources required for sample preservation and transportation, placing restrictions on the number of samples that can be collected in a given time and the size and remoteness of the regions that can be effectively surveyed. Timely processing and/or storage of the samples before they spoil can also be problematic in locations where access to well equipped laboratory facilities is limited.
We report here the use of FTA technology for efficient sampling and recovery of viral pathogens from infected leaf tissues and their subsequent molecular analysis. Utilising the geminiviruses that infect maize (Zea mays), cassava (Manihot esculenta) and tomato (Lycopersicum esuclentum), in addition to Tobacco mosaic virus (TMV), Potato virus Y (PVY) and Tobacco etch virus (TEV), we provide evidence that diagnostic techniques can be applied to both DNA and RNA viruses eluted from FTA cards in a manner equivalent to conventional isolation methods, and that this cost-effective technology significantly simplifies the sampling and analysis of diseased plants in both the laboratory and field environments.
FTA is a paper-based technology designed for the collection and archiving of nucleic acids, either in their purified form or within pressed samples of fresh tissue. Proprietary chemicals impregnated into the paper act to lyze cellular material and fix and preserve DNA and RNA within the fibre matrix . After a short drying period, pressed samples can be stored at room temperature for extended periods and processed when required. Nucleic acids are recovered by removing small punches from the pressed area and washing with simple reagents. RNA and smaller DNA molecules, such as plasmids and viral genomic components, are eluted by a simple extraction buffer and used as template for amplification by PCR. Genomic DNA remains attached to the paper matrix but is available for amplification by PCR when the paper punch is included in the PCR reaction mix. Advantages of FTA technology have been realized for human DNA processing and forensic applications , for wildlife DNA samples  and applied to PCR-based genotyping [5, 6] but have not been well documented for use with plant pathogens. Recognizing the potential benefits this technology could bring to sampling and molecular study of viral crop diseases, we tested the efficacy of FTA for retrieval of viral pathogens from infected leaf tissues and for the detection of viral-derived transgene sequences in transgenic plants.