Advances in Food Mycology

Black Aspergillus Species in Australian Vineyards 157
(Merck KGaA, Darmstadt, Germany) were enumerated on DRBC.
Samples were taken on nine occasions from air in the vineyard at
10 cm, 100 cm and 180 cm above the vineyard floor.
Statistical analyses were performed using Genstat (6th edition,
Lawes Agricultural Trust, Rothamsted, UK).
2.2. Survival of Aspergillus carbonarius Spores on the
Surface of Bunches Preharvest
A trial was conducted in the Hunter Valley, New South Wales,
Australia to examine the survival of A. carbonarius spores on the surface
of Chardonnay and Shiraz grapes (three replicate rows) during
the growing season in 2002-03. The vines were over 25 years old,
trained onto horizontal wires and under drip irrigation.
Spores of 7-14 day old cultures of A. carbonarius on Czapek Yeast
Agar (CYA) (Pitt and Hocking, 1997) plates were harvested into sterile
water containing Tween-80® (0.05% w/v; Merck, Victoria,
Australia), and diluted to 2-4 × 10 5 colony forming units per ml
(cfu/ml). Bunches were inoculated by immersion in 1 l of suspension
contained within a plastic bag. The same inoculum was used for up to
40 bunches without decrease in the spore concentration. Twelve
bunches in each row were inoculated at pre-bunch closure (berries
green and pea size), veraison and 11-16 days preharvest. Two bunches
were combined into a single sample, resulting in six samples per replicate
at each sampling stage. Inoculated bunches were sampled after
the inoculum had dried to give an initial value, at each of the subsequent
stages and at harvest.
Bunches were homogenised for 3 min in a stomacher (BagMixer,
Interscience, France) with the addition of sterile distilled water, and
serial dilutions of the suspension were plated onto DRBC. After incubation
for 3 days at 25°C, colonies of A. carbonarius were enumerated.
The average berry weight at each growth stage was calculated, and the
number of A. carbonarius colonies was expressed as cfu per berry, in
order to compare the number of viable spores present during each stage.
2.3. Winemaking
2.3.1. Inoculation and Vinification of Grapes
Berries were inoculated preharvest with a spore suspension of
A. carbonarius (prepared as described in 2.2) at approximately 10 7 cfu/ml.