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This study was carried out to work towards a better understanding of the interactions between ticks and <i>Metarhizium anisopliae, </i>an entomopathogenic fungus with the potential of being developed into a fungal biocontrol agent for this important pest of livestock and humans, and to perform preparatory work for field trials to be carried out by the group in the near future. A rapid and simple method of isolating and characterising <i>M. anisopliae</i> from soil samples has been developed, incorporating a spore floatation method of isolation, DNA extraction using FTAÒ cards and three PCR-RFLP methods of molecular characterisation, two of which have not yet been successfully applied in this area. During the development of the methods, a database of local <i>M. anisopliae </i>strains was constructed, all of which will collectively be essential for tracking and evaluating released strains. None of these local strains showed promise as potential tick biological control agents, but bioassays identified two tick-pathogenic strains that could be developed into biological control agents. The virulence of these and two other strains was found to differ greatly towards insect and tick species. Microscopy and biochemical studies have found that there are differences in spore attachment and germination on insect and tick cuticle that may influence virulence. Great variation was also seen in the properties of Pr1 enzymes produced by the four test strains of <i>M. anisopliae.</i> Although this may be expected to play an important role in virulence, results did not corroborate with those obtained in bioassays, revealing an area requiring further investigation.

The objective of the project is to improve the durability of calcium alginate gel, as an immobilizing matrix for yeast in aerobic culture, by increasing the alginate content of the gel in the range 4.5, 6.0, and 8.0 wt%, so that the immobilized yeast bioparticle can be used for at least 500 hours continuous culture, and to investigate the interaction between the gel matrix and the immobilized cells. The yeast, <i>S.cerevisiae</i> NCYC 1026, has been aseptically immobilized in thin calcium alginate gel discs (2.0 mm by 7.0 mm) of the required alginate content, and aerobically cultured at + 25°C and pH 4.5 in a chemically defined liquid culture medium. The immobilized yeast gel discs have been continuously cultured in an air lift bioreactor for at least 500 hours. The kinetic state of the immobilized cells and the immobilized biomass hold-up were measured independently, such that these data were available for the analysis of the continuous bioreactor, using mathematical models of the bioreaction system. The calcium alginate gel was found to reduce yeast growth and yield slightly, but to have a significant effect on glucose uptake by inducing active transport in the yeast. Otherwise the gel had no adverse action on the yeast, indeed the gel matrix tended to maintain yeast viability, when the immobilized cell gel discs were stored for up to 3 months with no glucose supply. During continuous culture the key operational parameters were found to be effective diffusivity of glucose in the gel matrix solid phase and immobilized cell biomass hold-up, possibly linked to the inconstant cell mass leakage rate from the gel. The 6.0 wt% alginate content gel was found to be the optimum gel for immobilized cell function and durability of the gel. Nevertheless the effective diffusivity even in this gel was only 1.0% of that in water, much lower than previously reported.

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