Population dynamics of Aspergillus section Nigri species on vineyard samples of grapes and raisins

Authors: Palumbo, Jeffrey - Jeff; O Keeffe, Teresa; Ho, Yvonne; FIDELIBUS, MATTHEW - University Of California - Cooperative Extension Service

Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/11/2015
Publication Date: 3/1/2016
Publication URL: https://handle.nal.usda.gov/10113/5298368
Citation: Palumbo, J.D., O Keeffe, T.L., Ho, Y.S., Fidelibus, M.W. 2016. Population dynamics of Aspergillus section Nigri species on vineyard samples of grapes and raisins. Journal of Food Protection. 79:448-453. doi: 10.4315/0362-028X.JFP-15-437.

Interpretive Summary: Black-spored Aspergillus fungi are commonly found in grape vineyards, and species of interest (particularly those that produce toxic compounds such as ochratoxins) are difficult to quickly identify. As a result, the amount of each species within a population of fungi is difficult to estimate. We used a quantitative PCR method to measure relative amounts of particular genes unique to each of four different Aspergillus species within the total fungi present on grapes, in order to estimate relative amounts of each species within the population. Grapes were sampled from two vineyards (W and D) at several stages of fruit development over two years. Traditional culturing showed that the amount of black-spored Aspergillus increased over the course of each growing season, and the quantitative PCR method showed that the relative amount of A. carbonarius (a fungal species responsible for ochratoxin production in grapes and raisins) increased within the population of black-spored Aspergillus as the season progressed. Comparing the two vineyards, harvest and raisin fruit samples in vineyard W contained more A. carbonarius within the total fungal population than vineyard D, and vineyard W fruit samples were also more frequently contaminated with ochratoxin than vineyard D. Together, this work shows that quantitative PCR methods are useful in measuring populations of potential toxin-producing fungi within total fungal populations present on fruits during the time of production.

Technical Abstract: Several species of Aspergillus section Nigri, including potential mycotoxin producers, are common residents of grape vineyards, but the relative population size of individual species throughout the growing season is difficult to determine using traditional isolation and identification methods. Using a quantitative droplet digital PCR (ddPCR) method in combination with dilution plating, total Aspergillus section Nigri populations and relative proportions of A. niger, A. welwitschiae, A. carbonarius and A. tubingensis were measured from vineyard samples without the need for identifying individual fungal isolates. Grapes were sampled from two raisin vineyards (vineyards D and W) at berry set, veraison, harvest and raisin stages in two consecutive years. Plate counts showed the total population of Aspergillus section Nigri present on the fruit increased from berry set to raisin, and became a larger component of the total recovered fungal population in both vineyards in both years. Results from ddPCR analysis showed that the relative proportion of A. carbonarius among the four species assayed increased later in the season (harvest and raisin) in comparison to earlier in the season (berry set and veraison). Total fungal and Aspergillus section Nigri plate counts were not significantly different between vineyards in either year. However, vineyard W generally showed higher proportions of A. carbonarius in harvest and raisin samples than vineyard D. This coincided with higher incidence and levels of ochratoxin A in vineyard W harvest and raisin fruit than in vineyard D fruit. This work demonstrates that this ddPCR method is a useful tool for culture-independent monitoring of populations of mycotoxigenic Aspergillus species during grape and raisin production.