3.3.21
SUGARCANE STRIATE MOSAIC DISEASE: DEVELOPMENT OF A DIAGNOSTIC TEST

N THOMPSON1, Y CHOI2 and JW RANDLES1

1Department of Crop Protection, The University of Adelaide, Waite Campus, Glen Osmond, South Australia, 5064, Australia; 2Department of Plant Pathology, University of California, Riverside, California, 92521, USA

Background and objectives
Sugarcane striate mosaic disease (ScSMD) was first reported in 1961 in the Burdekin district of Queensland, Australia. The disease appears to be unique to this area and causes a loss of approximately $A 1 M per annum to the Australian sugar industry.

The symptoms of ScSMD include characteristic short chlorotic striations present on most leaves of the sugarcane plant. Other symptoms include a general yellowing, reduced growth rate, shorter internode length and less thickening of the cane. Symptoms of ScSMD become more severe for subsequent ratoons, with older ratoons showing dramatically reduced growth rate and severe symptoms. Diseased cane is usually ploughed out after the first ratoon because the reduced growth rate and other ScSMD symptoms make it uneconomical.

Diagnosis of disease currently depends on recognition of the symptoms described above, but as the severity of symptoms can vary between different varieties, time of year, and other environmental factors, disease diagnosis is unreliable.

ScSMD was recently suggested to have a viral etiology and a slightly flexuous rod-shaped virus-like particle with single stranded RNA was isolated from diseased cane and named sugarcane striate mosaic virus (ScSMV) [1]. The RNA was partially sequenced [1]. This paper describes the use of sequence data to develop molecular diagnostic methods for ScSMV so that etiology and epidemiology can be fully described.

Materials and methods
dsRNA was extracted from ScSMD affected sugarcane by the method of microgranular cellulose extraction [2], gel purified and clones constructed by random priming of the dsRNA. cDNA was primed with random hexanucleotide linked to the 3' end of a universal oligonucleotide with an EcoR I site for cloning and an additional 5' site for priming by PCR. Cloning of the cDNA was done using the EcoR I restriction site in the PCR primer to insert random fragments into the pGEM-7zf (+) plasmid [2]. Automated sequencing of clones generated by this method was done using the termination method utilising the T7 and SP6 priming sites on the pGEM-7zf (+) vector. Sequences were edited using the SeqEd program and genbank database comparison was done using the FastA program of the webANGIS interface.

Results and conclusions
Sequence analysis has revealed that ScSMV appears to be closely related to apple stem pitting virus (ASPV) [3] and members of the carlavirus group. ASPV has recently been designated as the type species of the proposed genus Foveavirus [4], and it is suggested that ScSMV may be a member of this genus.

References
1. Choi, Y 1997. PhD thesis, University of Adelaide.
2. Choi, Y and Randles, JW, 1997. BioTechniques 23, 610-611.
3. Jelkmann, W 1994. Journal of General Virology 75, 1535-1542.
4. ICTV, 1998. Unpublished.