Investigations on seed transmission of cacao swollen shoot virus (CSSV)

Cocoa swollen shoot virus disease (CSSVD) seriously affects cocoa production in Ghana, particularly in the areas of mass infection (AMI) in the Eastern Region whilst the disease continues spreading to areas with scatter outbreaks especially in the Western Region. Reports utilising visible diagnostic symptoms on whether the virus is seed-borne and transmissible to seedlings of cocoa (Theobroma cacao L.) have been conflicting. Studies show that visible viral symptoms are not always manifested on all affected parts of infected cocoa trees due to a period of CSSV latency. In this work, molecular diagnostic techniques involving DNA and RNA based polymerase chain reaction (PCR) methods were utilised with CSSV primers designed from conserved regions of published CSSV sequences to investigate whether CSSV is sexually transmissible from infected parents to developing seeds and seedlings through a series of pollination experiments. The study also evaluated whether pollen could be a source of CSSV infection into seeds and seedlings through cross pollination experiments by using pollen from CSSV infected cocoa trees onto healthy recipients. Robust precautions were taken to avoid cross contamination during duplicated DNA extractions and only those PCR results accompanied by effective positive and negative controls (from CSSV symptomatic and quarantine assured CSSV free trees respectively) were scored. Following DNA extraction all pods obtained from the self pollination experiments involving CSSV infected cocoa trees were found to contain seeds that were PCR positive for CSSV DNA. The pith (mesocarp) of each of the self pollination generated pods was similarly PCR positive for CSSV DNA, though not all seeds from the pods were positive for the PCR screening. Seed components showed diminishing frequencies of CSSV DNA presence in the order: testa> cotyledon> embryo. Sequential DNA samples from new leaves of some of the seedlings derived from the self-pollinated trees are consistently PCR positive for CSSV DNA over 24 months after germination. Nonetheless, a reverse transcription (RT) PCR screening test has been negative for all those PCR positive seedlings indicating absence of CSSV RNA transcripts in the seedlings. None of the seedlings have exhibited symptoms characteristic of CSSV and infectivity tests on them have been negative. Seeds and seedlings from the cross pollination experiments have also been negative for the PCR analysis implying that pollen could not to be a potential route for CSSV infection. Possible explanations for these events including viral DNA integration into the cocoa genome and possible methylation-directed inactivation of the CSSV DNA are discussed.EThOS - Electronic Theses Online ServiceGBUnited Kingdo