Detection and quantification of Botrytis cinerea incidence and severity in harvested pears for the development of a monitoring system to support commercial exports

A method to monitor Botrytis cinerea inoculum levels will greatly assist in the management of the Botrytis rot in pears. The objective of this project was to develop a system to continuously detect and quantify B. cinerea levels in orchards, packhouses and packed fruit. As a secondary aim, we attempted to correlate the inoculum levels present with decay levels measured at the end of storage and shelf.

During the first season, pear blossoms and fruit were harvested from orchards at full bloom and optimal harvest maturity respectively. These samples were either artificially inoculated with known amounts of B. cinerea spores or left un-inoculated. From these samples genomic DNA was extracted and used in a quantitative Real-Time PCR (qRT-PCR) reaction to determine the relative B. cinerea DNA content present in pear blossoms and the calyx-end of fruit at specific time-points. For the second season, fruit samples from several orchards were taken at various time-points during production and storage. In these samples the Botrytis cinerea DNA content was monitored and at the end of the storage period fruit was sampled to determine the incidence and severity of Botrytis rot.

• B. cinerea DNA was detectable in a) blossoms at full bloom and b) the calyx-end of fruit harvested at optimal maturity.
• There was a strong correlation between the cinerea DNA content and the number of spores artificially inoculated in both blossoms and the calyx-end of fruit.
• The minimum quantifiable limit of the test is between 10-100 spores for blossoms and 100 spores for calyx-end tissue.
• 15 Fruit per sample pool is an adequate sample size and produces reproducible results.
• In samples where Botrytis cinerea was artificially inoculated into the calyx-end, there was a significant reduction in the cinerea DNA content after chlorine flume treatment. This was not true for samples where no artificial inoculation was done.

• Botrytis cinerea DNA was detected at all time-points for all production areas analysed.
• The inoculum amount did not significantly change from the levels measured at the very first time-point. This shows the importance of infection early in production (possibly bloom) in determining the level of inoculum present during storage.
• Flume treatment during storage did not have a significant effect on the Botrytis cinerea inoculum within the calyx-end.
• The initial infection, most probably occurring during bloom, seems to be crucial in determining the level of Botrytis cinerea inoculum present during production and storage.