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The cytopathological and molecular studies of cacao swollen shoot virus (CSSV) infected cocoa plants, which was aimed at identifying structural and functional changes associated with CSSV infections that can be used as resistant markers has yielded interesting results. Nucleic-acid-rich spherical inclusion bodies, modified plastids, invagination of cell walls, apoptotic cells, massive deposition of phenolic substances in infected cells, in situ localisation of CSSV particles in novel cocoa tissues and the apparent aggregation of starch granules in infected tissues were changes that were found to be accompanying CSSV infection by microscopy. Since most of these discoveries were new in CSSV research, they were validated using PCR (conventional and quantitative (qPCR», flow cytomefry (FCM) and laser scanning confocal microscopy (LSCM). The qPCR assay, which was developed from the scratch to measure the CSSV content in infected cocoa plants, has since been used to start building profiles of CSSV resistant cocoa cultivars from Ghana. It was also used to· compare the viral load between the cotyledons and leaves of an infected cocoa plant, whereby the cotyledons were found to have 4-folds more CSSV than the leaves. The qPCR assay has also been used to start studying the movement of CSSV in infected cocoa plants. The FCM was used to confirm apoptotic outcome in CSSV infected plants. This was shown by a reduction in total nuclei DNA between infected and healthy plants at magnitudes which correspond to the microscopic observations. Results from the LSCM were inconclusive. Light microscopic analysis of nucleicacid- rich inclusion bodies, which would have added to building a more robust profile ofthe CSSV resistant cocoa cultivars from Ghana could not be completed due to time constraints. The significance of all these findings in breeding ~or CSSV resistant cocoa, and in CSSV research is discussed.

Since the late 1990's, crown dieback of young (approximately 5 to 10 years old) birch (Betula pendula and B. pubescens) has been causing concern at over 20 planted sites in Scotland. Preliminary studies showed that the endemic fungus Anisogramma virgultorum is pathogenic on birch and a suspected causal agent ofcrown dieback at affected sites. A survey conducted at nine Scottish Woodland Grant Scheme sites in 2004 and 2006 showed that disease incidence and severity was greater on B. pubescens than B. pendula (n=900) over all sites and over both survey years. Crown dieback was significantly greater on infected than uninfected B. pubescens. Anisogramma virgultorum was also present on trees surveyed at two (Dunkeld and Domoch) ofthe six birch provenanc(trials located in Scotland with 17.3% (n=2,741) and 3.7% (n=830) disease incidence, respectively. At the Dunkeld trial site, the presence of A. virgultorum contributed significantly to crown dieback. During a survey of 90 site-natural birch stands across Scotland (n=8,492) A. virgultorum was only present at 13 sites. Incidence ofdisease at each site was generally low (::;5%) except for three sites where 12, 21 and 52% incidence of A. virgultorum were recorded. An ascospore release experiment conducted over a 94-week period showed a strong periodicity with main spore release events occurring in spring (from April to June) of 2006 and 2007. This coincided with flushing of the vegetative buds and elongation of young birch shoots. Natural infection and artificial inoculation experiments showed that young birch seedlings are susceptible to infection with ·A. virgultorum from flushing of the vegetative buds through shoot elongation. In a histopathological study of A. virgultorum cankers different stages of canker development on birch shoots are described. Fully developed cankers with living perithecia were observed four to five months from initial infection. In a shoot dieback study, four branch systems of B. pubescens trees and four intact birch seedlings were monitored regularly over a two-year period. The majority of shoots with A. virgultorum cankers died back within one or two years after initial infection. A phylogenetic analysis of LSD sequences of A. virgultorum and the closely related A. anomala was conducted with 57 representative taxa of the order Diaporthales. The two fungi formed a well-supported monophyletic clade outside the Gnomoniaceae sensu strictu and were unrelated to the remaining eight families outlined in the Diaporthales.

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