ARS | Publication request: Linear-motion tattoo machine and prefabricated needle sets for the delivery of plant viruses by vascular puncture inoculation

Submitted to: European Journal of Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: June 18, 2011
Publication Date: July 3, 2011
Citation: Weiland, J.J., Edwards, M.C. 2011. Linear-motion tattoo machine and prefabricated needle sets for the delivery of plant viruses by vascular puncture inoculation. European Journal of Plant Pathology. 131:553-558.

Interpretive Summary: The natural spread of plant viruses between host plants is enabled by numerous vector organisms, as well as by mechanical wounding. Transmission by rub inoculation on the leaves of host plants has facilitated experimental research on plant viruses for decades, yet many plant viruses, especially some of those transmitted by insects, cannot be propagated in this manner. A technique known as vascular puncture inoculation (or VPI) was developed years ago to address this problem. In this technique, the seed of a plant such as maize is inoculated rather than the plant. The virus is delivered by puncturing the seed with fine insect pins with the goal of delivering the virus to the vascular tissue present near the embryo in the seed. These pin assemblies are assembled by hand and driven by an engraving tool. In an effort to improve this technique, a linear-motion tattoo machine driving industry-standard needle arrays was tested as a means of delivering plant viruses into maize and small grain seed embryos. The new method was applied in the successful transmission of maize rayado fino virus (MRFV) and oat blue dwarf virus (OBDV), although no infection was obtained in repeated tests with barley yellow dwarf virus (BYDV-PAV) or cereal yellow dwarf virus (CYDV-RPV). Replacement of the original engraving-tool with a linear-motion tattoo machine in VPI provides greater flexibility and convenience in a quiet, readily-available instrument, while improving reproducibility through the use of prefabricated needle arrays.

Technical Abstract: Vascular puncture inoculation (VPI) of plant viruses previously has been conducted either manually or by use of a commercial engraving tool and laboratory-fabricated needle arrays. In an effort to improve this technique, a linear-motion tattoo machine driving industry-standard needle arrays was tested as a means of delivering plant viruses into maize and small grain seed embryos. The new method was applied in the successful transmission of maize rayado fino virus (MRFV), the type member of the genus Marafivirus, from an archived sample to maize. Subsequent transfer of MRFV from the sap of an infected plant using the method produced an average infection rate in maize of 70% (range 39% - 93%). Maize, oat, and triticale were successfully infected with oat blue dwarf virus (OBDV) using the method; similar infection rates were observed between maize seeds inoculated with the tattoo machine and those inoculated with the engraving machine when using prefabricated needle arrays. No infection was obtained in repeated tests with barley yellow dwarf virus (BYDV-PAV) or cereal yellow dwarf virus (CYDV-RPV) using either sap or RNA from infectious cloned cDNA. Replacement of the original engraving-tool with a linear-motion tattoo machine in VPI provides greater flexibility and convenience in a quiet, readily-available instrument, while improving reproducibility through the use of prefabricated needle arrays.