Transgenic Expression of dsRNA Targeting the Pentalonia nigronervosa acetylcholinesterase Gene in Banana and Plantain Reduces Aphid Populations.

The banana aphid, Pentalonia nigronervosa, is the sole insect vector of banana bunchy top virus (BBTV), the causal agent of banana bunchy top disease. The aphid acquires and transmits BBTV while feeding on infected banana plants. RNA interference (RNAi) enables the generation of pest and disease-resistant crops; however, its effectiveness relies on the identification of pivotal gene sequences to target and silence. Acetylcholinesterase (AChE) is an essential enzyme responsible for the hydrolytic metabolism of the neurotransmitter acetylcholine in animals. In this study, the AChE gene of the banana aphid was targeted for silencing by RNAi through transgenic expression of AChE dsRNA in banana and plantain plants. The efficacy of dsRNA was first assessed using an artificial feeding assay. In vitro aphid feeding on a diet containing 7.5% sucrose, and sulfate complexes of trace metals supported aphid growth and reproduction. When AChE dsRNA was included in the diet, a dose of 500 ng/µL was lethal to the aphids. Transgenic banana cv. Cavendish Williams and plantain cvs. Gonja Manjaya and Orishele expressing AChE dsRNA were regenerated and assessed for transgene integration and copy number. When aphids were maintained on elite transgenic events, there was a 67.8%, 46.7%, and 75.6% reduction in aphid populations growing on Cavendish Williams, Gonja Manjaya, and Orishele cultivars, respectively, compared to those raised on nontransgenic control plants. These results suggest that RNAi targeting an essential aphid gene could be a useful means of reducing both aphid infestation and potentially the spread of the disease they transmit.

RNAi technology for management of banana bunchy top disease.

Banana bunchy top disease (BBTD) is one of the world's most destructive viral diseases of banana and plantain, causing up to 100% yield loss in severe cases. The disease is vectored by banana aphids (Pentalonia nigronervosa) and carried long distances through the movement of infected plant materials. The banana aphids harboring banana bunchy top virus (BBTV) present in banana producing regions are the sole vector and the most efficient method of transmitting the virus to the healthy plants. Controlling the spread of BBTD has been very challenging since no known banana germplasm is immune to BBTV. The disease can be managed with the use of virus-free planting material and roguing. However, once BBTD is established in the field, it is very difficult to eradicate or manage it. Therefore, a more sustainable way of controlling the disease is developing host plant resistance against the virus and the vector. Biotechnological strategies via RNA interference (RNAi) could be used to target the banana aphid as well as BBTV to reduce virus-associated yield losses of banana and plantain, which feed over 500 million people around the world. This review discusses the status of BBTD and perspectives on effective RNAi technologies for controlling BBTV and the vector, banana aphid, transmitting the virus as sustainable management of the disease.

Phytochemical Analysis and Establishment of Embryogenic Cell Suspension and Agrobacterium-mediated Transformation for Farmer Preferred Cultivars of West African Plantain (Musa spp.).

Banana and plantain are among the foremost staple food crops providing food and livelihood to over 500 million people in tropical countries. Despite the importance, their production is hampered due to several biotic and abiotic stresses. Plant tissue culture techniques such as somatic embryogenesis and genetic transformation offer a valuable tool for genetic improvement. Identification and quantification of phytochemicals found in banana and plantain are essential in optimizing in vitro activities for crop improvement. Total antioxidants, phenolics, flavonoids, and tannins were quantified in various explants obtained from the field, as well as in vitro plants of banana and plantain cultivars. The result showed genotypic variation in the phytochemicals of selected cultivars. The embryogenic cell suspensions were developed for three farmer-preferred plantain cultivars, Agbagba, Obino l'Ewai, and Orishele, using different MS and B5-based culture media. Both culture media supported the development of friable embryogenic calli (FEC), while MS culture media supported the proliferation of fine cell suspension in liquid culture media. The percentage of FEC generated for Agbagba, Obino l'Ewai, and Orishele were 22 ± 24%, 13 ± 28%, and 9 ± 16%, respectively. Cell suspensions produced from FECs were successfully transformed by Agrobacterium-mediated transformation with reporter gene constructs and regenerated into whole plants.

Agromorphologic, genetic and methylation profiling of Dioscorea and Musa species multiplied under three micropropagation systems.

Plant in vitro vegetative propagation using classical semi-solid culture medium is limited due to the low degree of automation, suboptimal nutrient availability and induced physiological stress which often reduce its efficiency. Temporary Immersion System (TIS) emerged as an innovative approach to optimize and eliminate the drawbacks associated with the conventional system of micropropagation. In this study, both Dioscorea and Musa spp. were subjected to conventional semi-solid culture media, complete immersion in shaking liquid culture media and TIS using RITA bioreactor. In vitro grown plantlets were screened for possible vegetative changes using agro-morphological descriptors while genetic and methylation differences were assessed using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP). In vitro results showed that the number of shoots produced in Musa spp. varied significantly (P≤0.001) with the type of culture system. The highest mean shoot produced was observed with TIS (28.40) and the least using semi-solid culture medium (1.13). For Dioscorea spp., there was no significant interaction between the hormone combination and the culture system. However, the lowest mean shoot value (1.55) was observed in the semi-solid culture medium. Genetic analysis via AFLP using 15 primer pair combinations revealed that the 3 culture systems maintained genetic variation for Musa and Dioscorea spp. under in vitro and field conditions. Results showed 99% and 91% of the total bands were polymorphic under in vitro and field conditions respectively for Musa and 100% polymorphism for Dioscorea under in vitro and field conditions. Methylation investigation via MSAP using 12 primer pair combinations showed 25% and 46% polymorphic methylated-sensitive loci, 100% and 78% of non-methylated loci of the total bands generated under in vitro and field conditions respectively. Unmethylated (HPA+/MSP+) levels were highest in TIS (0.0842) as compared to CI (0.0227) and SS (0.0161) while full methylation or absence of target (HPA-/MSP-) was lowest in TIS (0.5890) and highest in SS (0.7138). For Dioscorea, 52% and 53% methylated sensitive loci and 100% non-methylated loci were polymorphic under in vitro and field conditions respectively. Although in vitro plant tissue culture techniques led to methylation at some loci of both species, there were no observable changes in the phenotype of both crops under field conditions. This also confirmed that not all methylation events lead to phenotypic changes.