NOVEL DIAGNOSTIC METHOD FOR RATOON STUNTING DISEASE: DEVELOPMENT AND IMPLICATIONS FOR RSD MANAGEMENT

A NEW MOLECULAR RSD diagnostic platform was developed using polymerase chain reaction (PCR) on pooled leaf sheath biopsies (LSB-PCR). This non-destructive technique samples leaf sheaths from 50 cane stalks, more than tripling the number of stalks routinely surveyed using the current evaporative-binding enzyme-linked immunoassay (EB-EIA) method, and does not require DNA extraction steps. In-field sample collection takes marginally longer, however primary sample processing takes less than a minute, contrasting favourably with the laborious scrubbing, cutting and pumping of cane stalks required by the EB-EIA method. A sample of 31 fields was selected for analysis after returning an EB-EIA absorbance result of >0.05, of which seven were confirmed as having RSD based on phase contrast microscopy (PCM). LSB-PCR detected RSD in six of these seven fields, but also in an additional seven fields presumed to be negative based on EB-EIA/PCM results. The causal agent of RSD, the bacterium Leifsonia xyli subsp. xyli, was confirmed as present by sequencing amplicons from three fields, two for those classed positive and one negative for RSD based on the EB-EIA/PCM method. The LSB-PCR is quicker and more sensitive than the EB-EIA method, and has the potential to provide insights into why RSD persists as a major productivity constraint in the Australian industry.