The Effect of Trichoderma harzianum on Honey Bee Survival

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METHODS

Fifteen nuc size honey bee, Apis mellifera, hives were placed in a five acre strawberry planting at the New York State Agricultural Experiment State in Geneva, NY. Hives were placed on the northern field edge of the field and separated from each other by 3 meters. Because plenty of pollen and nectar sources were available to the bees during the span this experiment, robbing activities between hives were minimal. Only 10 of the 15 hives were used in this experiment. Two treatments (hives treated with T. harzianum and an untreated check) were randomly assigned to the hives and replicated 5 times. A patent pending applicator device filled with T. harzianum strain T.39 (Trichodex, 1 x 1010 cfu/g, lot #395006, Makhetshim-Agan of N.A.) was placed in front of five hives on 8 July 1997. Trays were weighed and refilled twice a week until the experiement was terminated (8 August). As the bees entered and exited the hives, they were in constant contact with T. harzianum. Bees in the untreated hives walked through empty trays and were not exposed to T. harzianum. All hives were weighed at the initiation of the experiment, reweighed at the end, and the net change in weight was calculated. Prior to placement of the trays in front of the hives, 100 young bees in each hive were marked on the dorsal thorax with non toxic paint. Hives were then opened twice a week for 4 weeks and the number of live marked adults were counted. Counting was conducted early in the morning before a significant number of bees left the hive so a more accurate count could be obtained.

To determine the number of Trichoderma spores carried per bee, single bees were collected from the inside and outside of hives each week. After collection, individual bees were placed in 3 ml of a 40% detergent solution and shaken for 2 hours in a refrigerated room to remove and suspend Trichoderma spores. Two serial dilutions were made from this suspension and 100 µl of each suspension was applied to Trichoderma selective API media (Acid Potato Dextrose Agar with Igepal) and placed in an incubator at 30· C for 3 to 4 days. After removal, the number of colony forming units (cfu) were counted, and the number of spores per bee calculated using the formula [no. cfu per plate x (no. ml detergent/bee) x 10 x 10 dilution]. At the conclusion of the experiment, brood size was determined by removing a center frame, measuring the area containing brood and counting the number of the brood cells per frame. Survival data was statistically analyzed using nonparametric survival analysis (Kaplan-Meier estimator and rank tests) to evaluate differences in survival patterns between treatments and T-tests were used to analyze other data (Statview, Abacus Inc.).

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