Using ISTA techniques, a total number of 14 genera and 39 species of fungi viz., Alternaria alternata*, A. tenuissima*, Alternaria spp. *, Aspergillus candidus*, A. carneus*, A. clavatus*, A. fischeri* A. flavus, A. flavipes*, A. fumigacus*, A. glaucus*, A. jaul4S*, A. niger, A. ochraceus*, A. restrictus*, A. sydowi*, A. sulphureus*, A. terreus*, A. versicolor*, A. wentii*, Chaetomium spp. *, Cladiosporium fulvum*, Drechslera australiensis*, D. hawaiensis*, D. State of Cochliobolus spicifer*, Fusarium equiseti*, F. moniliforme*, F. solani*, Fusarium spp. *, Nigrospora sp. *, Paecilomyces liladnus*, Pe1icillium camemberti*, P. frequentans*, Penicillium spp. *, Rhizopus sp., Rhizoctonia solani*, Trichoderma harziallum*, Trichothecium roseum and Uloclodium sp. *, were isolated from 40 almond seed samples collected from different. localities of Pakistan. Of these only A. niger, A. flavus, Rhizopus sp., and Trichothecium roseum have previously been reported and the remaining 35 species of fungi marked with an asterisk are new records. Agar plate method yielded greater number of fungi (34 species) as compared to blotter (29 specie3) and deep freezing method (28). Component plating showed that seed coat was infected by greater number of fungi follower! by cotyledons, shell and axis. Almond seeds collected from Sindh showed precominace of mold fungi as compared to samples collected from Balochistan. At least 39 (97.5 %) seed samples were found infected with Aspergillus flavus showing a mean infection percentage of upto 49.7%.
Similarly 15 almond seed samples collected from different countries like Dubai, Abudhabi, Saudi Arabia (Mecca and Madina), Kuwait, Iran, Germany, England, Australia, USA and Egypt showed the occurrence of 7 genera and 21 species of fungi. Seeds collected from Iran were found more moldy (14 species) as compared to samples collected from U.K. (5 species of fungi) with very low intensity of infection.
A number of fungi isolated from almond seeds are known to produce mycotoxin harmful for human health. Aspergillus flavus which produces aflatoxin B1 and B2, G1 and G2 was detected in 49.7% almond seeds. Using AOAC method, out of 25 seed samples analysed, 5 were infected with aflatoxin B1 ranging from 5.8-140 µg/kg which included 2 samples showing the presence of aflatoxin B2 (7.0 and 140 µg /kg) as well. Using plug method and comparing with B1 standard, out of 29 different isolates of A. flavus only 9 were found to be toxigenic showing the presence of aflatoxin B\. The ratio of toxigenic and non-toxigenic strains was approximately 1 :2. Similarly using TLC technique, out of 25 seed samples, 5 were infected with zeralenol and only one sample with zeralenone toxin secreted by Fusarium sp.
In experiments on the control of mycoflora associated with almond seed where the effect of sorbic acid, propionic acid, NaCl, sunlight, colour rays and heat treatments from autoclave, even and microwave oven was examined, autoclaving was found to be most effective in killing all the fungi present on almond seed. Prorionic acid used @1000 ppm reduced infection of mold fungi by upto 90% and green colour rays reduced A. flavus infection by upto 80%. Propionic acid used @ 2000 ppm completely eliminated aflatoxin B1 in almond seed samples artificially inoculated with aflatoxin B1 whereas sorbic add was not effective. S Similarly propionic acid used @ 2000 ppm reduced aflatoxin collceatration by 50% in almond seeds artificially inoculated by a toxigenic strain of A. flavus while no aflatoxin could be detected in seeds where higher doses of propionic acid @ 4000,8000, and 16000 ppm were used. NaCl used @ 2% w/w reduced the at1atoxin by 20% while autoclaving showed complete control of A. flavus infection and upto 93 % reduction in aflatoxin concentration.