Syrah decline disease is a well documented problem in California and France. It is characterized by swelling and cracking of the graft union, stem pitting and grooving, and premature leaf reddening. French scientist have been studying this problem since 1999 and have failed to find any correlation with genetic incompatibility, crown gall infection, known pathogens (viruses, fungi, viroids), or environmental conditions. They concluded that the problem had no simple explanation and currently, examining diseased grapevines for a new virus. The potential of a virus causing Syrah decline is reasonable in that trunk stem markings support the hypothesis. In our earlier investigation working with diseased specimens, a new molecularly distinct strain of Rupestris stem pitting associated virus (RSPaV), designated as the Syrah strain, was characterized. However, in follow up assays, we were unable to establish correlations between Syrah decline and RSPaV-SY.
A new genome sequencer, the Genome Sequencer FLX, which is accessible through 454 Life Science Corporation (brandford, CT) offers an opportunity for mass sequencing of cDNA fragments produced from RNA viruses, other microbial organisms or plant related nucleic acids. This system circumvents the bacterial-cloning steps, which is inefficient in retaining clones of viruses in low titers and could sequence thousands of cDNA molecules of 100-300bp in size at a time. In 2007, cDNA samples were prepared from two different Syrah clones, Syrah 6 (showing pitting, wood necrosis and declining symptoms) and Syrah 8 (looks healthy with no visual symptoms), and sent to the 454 sequencing facilities. Total number of fragments (about average of 100-300 nt in size) sequenced by the Genome Sequencer were 169,184 and 182,406 for Syrah 6 and Syrah 8, respectively. Among them, 59,398 and 276 from Syrah 6 and Syrah 8, respectively, belonged to plant viruses and viroids. The viruses that were identified included: RSPaV, Grapevine rupestris vein feathering virus (GRVFV), Grapevine redglobe associated virus (GRGaV), Grapevine leafroll associated virus 9 (GLRaV-9) and a new uncharacterized virus. The viroids included: Hop stunt viroid (HSVd), Australian grapevine viroid AGVd), and Grapevine yellow speckle viroid (GYSVd). Only two viruses that previously have been found by RT-PCR were also found by the 454 sequencing facility in Syrah 8.
Work is in progress to assemble the sequences of all the segments for each virus/viroid and design specific PCR primers for their detection and use the primers to establish a possible correlation between one or multiple of these viruses/viroids with Syrah decline syndrome.