1a. Objectives (from AD-416)
The overall objective of this cooperative research project is to minimize the impact of Banana Bunchy Top Virus (BBTV). Research partly funded by the previously funded SCA resulted in the development of transgenic banana lines that show initial resistance under greenhouse conditions. The objectives of this SCA are as follows: 1) clonally multiply transgenic banana lines, 2) test selected lines for resistance to BBTV under greenhouse and field conditions, 3) evaluate horticultural properties of resistant lines under field conditions, and 4) deregulate transgenic line(s) for potential commercialization.
1b. Approach (from AD-416)
Transgenic banana lines that showed resistance to BBTV in initial tests will be clonally propagated in tissue culture and serve as a source for further studies. Lines will be tested under greenhouse and confined field conditions to test for their resistance to BBTV via controlled inoculations or field inoculations with the aphid vector. The tested lines will also be allowed to fruit under field conditions and their fruit as well as other horticultural properties will be evaluated. Selected lines will be further tested in larger scale field trials, while the lines are being characterized molecularly. The best lines will be characterized in more detail to obtain information that will be used to develop petitions to have the transgenic banana deregulated by the USDA Animal Health and Inspection Service, Environmental Protection Agency, and for consultation with the Food and Drugs Administration. Formerly 5320-21000-011-06S (6/2009); SCA with UH Manoa.
3. Progress Report
The agreement was established in support of the Objective of the in-house project, the goal to develop new knowledge about the genetics, genomics, and transgenics of selected tropical crops. Results of 3 field trials showed that test aphids can continue to acquire BBTV from leaf of Roundup® injected plants up to 3 weeks after injection, whereas no aphid was able to acquire BBTV at 1 week after the spray of Provado® following Roundup® injection. This suppression did not persist and certain percentages of aphids were still able to acquire BBTV until 3 weeks after injection. Provado treatment significantly reduced percentages of test aphids acquiring BBTV from injected plants. Second trial showed that most plants died between 3-4 weeks after RoundUp injection. Results support our hypothesis that combination of RoundUp injection and Provado spray reduce the transmission of BBTV. Three cultivars of banana with distinct genotypes were planted to compare their susceptibility to BBTV. Half of 48 plants of each cultivar were inoculated with BBTV (viruliferous aphids) and the other half were not. Results showed the rate of BBTV infection was low. However, ‘Saba’ cultivar had the highest number of BBTV symptomatic plants, followed by ‘Largo’ and ‘Apple’ cultivars. This has contradicted the myth of many growers that banana varieties, such as ‘Saba’ cultivar, with BBB genome are more resistant to diseases. Microsatellite study showed that aphids were highly polymorphic across the colonies sampled and there were multiple aphid clonal lineages colonizing banana and alternative host plants in Hawaii. Analysis of aphid genotypes showed clusters of aphid populations across all the Islands within 3 distinct genetic groups. These genotypes seemed to have distinctive preference for host plants. Aphids of Group 3 were found mostly abundant on ginger plants but not on banana plants. Conversely, aphids of Group 2 were mostly found on banana but not on ginger. Aphids of Group 1 were equally present in banana and ginger. In addition, all the 3 groups were found on taro, and Groups 1 and 2 were also found in Heliconia, Costus sp, Xanthosoma, and Dieffenbachia. A total of 21 independently-transformed lines of ‘Dwarf Brazilian’ were produced. Plants of these resistant lines are being multiplied and field tested. Transgenic lines of Mexican lime resistant to Citrus Tristeza Virus (CTV) were produced. These lines have been evaluated in the greenhouse for resistance to CTV by exposure to viruliferous brown citrus aphids. Of these 52 lines, 17 remained uninfected by CTV as determined by tissue blot immunoassays. All three transgenic plants transformed with an empty vector became infected with CTV. However, only one of the three wild-type Mexican lime plants appeared to be infected with CTV. This project is monitored via progress reports, meetings, and telephone and email communications.