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The present study was carried out to determine the mechanisms of biological control of <i>Fusarium oxysporum </i>f.sp. <i>lycopersici </i>by <i>Brevibacillus brevis </i>under greenhouse conditions.  Dual culture techniques demonstrated that <i>B. brevis </i>is capable of inhibiting the growth of fungal mycelium and germination of fungal spores due to excretion of antifungal metabolite(s) into the substrate, while reciprocal <i>in vitro </i>tests demonstrated no impacts of <i>F. oxysporum </i>f.sp. <i>lycopersici </i>metabolites on viability of <i>B. brevis.  </i>When live tomato roots were introduced into microcosms containing <i>F. oxysporum </i>f.sp. <i>lycopersici </i>and <i>B. brevis,</i> however, the fungal pathogen showed a competitive advantage over controls without <i>B. brevis </i>in terms of host colonisation.  The steroidal glycoalkaloid α-tomatine, extracted from tomato, inhibited conidiospore germination and growth of <i>Fusarium oxysporum </i>f.sp <i>lycopersici,</i> but had no detectable effects on viability of <i>B. brevis.</i> The effects of preinoculation of tomato (<i>Lycopersicon esculentum, </i>variety Moneymaker) roots with <i>B. brevis </i>in presence and absence of <i>F. oxysporum </i>f.sp. <i>lycopersici</i> on the induction of defence enzymes was examined.  Varying activities of phenylalanine ammonia lyase (PAL), peroxidases (POX) and chitinases were observed in different treatments at different time points, suggesting that the genes encoding these enzymes play a role in defence against Fusarium wilt.  Although no direct evidence of induction of these enzymes by <i>B. brevis </i>alone was observed, plants which were preinoculated with the bacterium showed enhanced induction of PAL, POX and chitinases at different time points following subsequent inoculation with <i>F. oxysporum </i>f.sp. <i>lycopersici.</i> Complex interactions between the tomato plant, <i>B. brevis </i>and <i>F. oxysporum </i>f.sp. <i>lycopersici </i>occurred and changed with time, as indicated by variations in metabolites at different sampling times.

The effect of <I>L. terrestris </I>on the transport of marked bacterial inocula in the absence of percolating water was investigated using repacked soil microcosms over a 10 day period. The microcosms consisted of cylindrical cores containing loamy sand. The bacterial inoculum was applied in filter paper disks to the surface of soil cores containing <I>L. terrestris </I>and to control cores. Destructive analysis of cores was carried out 5 and 10 days after inoculation to facilitate enumeration of marked bacterial inocula at 3 depths in the soil. Significantly greater vertical transport of marked bacteria occurred in cores containing <I>L. terrestris. </I>The effect of <I>L. terrestris </I> on the dispersal of marked bacteria in the presence of percolating water was investigated using both repacked soil cores and larger intact cores over a one month period. Bacterial inocula were again applied in filter paper disks to the surface of cores and the cores subjected to simulated rainfall events at 3 day intervals. Concentrations of marked bacteria in leachate were determined and destructive harvests of cores carried out after 24 days. The results from repacked soil cores demonstrated that cores containing earthworms were associated with early breakthrough, higher percentage recovery and greater numbers of marked bacteria in the leachate than control cores without earthworms. Results from intact soil cores demonstrated that cores containing earthworms were associated with different bacterial leaching patterns, higher percentage recovery and greater numbers of bacteria in the leachate than control cores. For both repacked and intact cores, the presence of earthworms in cores led to transport of marked bacteria to greater depths in the soil. Plaster of Paris impregnation of repacked and intact cores highlighted the importance of earthworm burrowing in providing pathways for bypass water flow and hence bacterial transport through soil. Results from the use of interdisciplinary methods developed and adapted in this study identify the importance of earthworms in the fate of microbial inocula in soil and the potential for use of earthworms in biotechnology.

This study aims to investigate the link between genetic functioning and diversity, and to establish a relationship, if any, between that diversity and the stability of the community. Extreme transient perturbations were performed, using repeat chloroform fumigations, to reduce the community to its lowest genetic level (0.7% of the DNA of the control population). Severe reduction in community diversity and selection for bacilli was initially observed, the community subsequently recovered (6 months) to its previous diversity but was altered in community composition, bacilli were no longer dominant. Perturbations using benzene resulted in a reduced genetic diversity (16S rDNA and DGGE) which did not recover (after 4 weeks). However, full recovery was observed for culturable cell numbers, DNA concentration and community level physiological profile (Biolog)., suggesting high levels of genetic redundancy within soil. Investigation of the relationship between diversity and stability was performed by perturbing soils with naturally differing levels of diversity (Sourhope>Boyndie) to equivalent toxicity, using copper and benzene. Large decreases in biomass were observed for both soils after benzene perturbation. Assuming successful equivalent toxicity, the genetic diversity of the naturally more diverse soil was more resistant to benzene perturbation than the less diverse soil. The broad scale function of both benzene-treated soil communities was unaffected (14C-labelled wheat shoot mineralisation). Narrow niche function was impaired for both soils (14C-labelled 2,4-DCP mineralisation). The Sourhope soil recovered this function by the end of the experiment, but Boyndie did not. The genetic diversity of both soils was resistant to copper perturbation despite biomass decreases. Initial shifts in physiological profiles of both soils was observed but rapidly returned to that of the control. In conclusion, diversity-stability relationships may exist in soil communities, despite generally being an extremely robust system.

Survival of P. fluorescens 10586s FAC510 was studied in liquid culture and soil microcosms in the presence of the ciliate C. steinii and the flagellate Cercomonas sp. Both protozoa caused a reduction in bacterial viable cell concentration, however the ciliate caused a greater decrease in abundance. Starvation of the bacterial prey caused a reduction in grazing rates of both the ciliate and flagellate, indicating the lower nutrional quality of starved as opposed to non-starved cells. Manipulation of the physiological state of prey cells, by starvation, had much the same effect as observed when bacterial prey resources are reduced. Luminescence provided a valuable marker for monitoring P. fluorescens 10586s FAC510 in liquid culture and soil, since detection by luminometry provided a sensitive, rapid, and non-extractive technique for measurement of microbial activity. In the presence of C. steinii, bacterial activity increased, whilst predation by Cercomonas sp. caused a reduction. The different responses of the bacterial inoculum may reflect the divergent balance between bacterial turnover, leading to nutrient regeneration, and grazing strategies, which are selective of the two protozoa. The distribution of bacterial cells in the soil pore network was manipulated by adjustment of the antecedant matric potential prior to inoculation. The ability to predominantly place bacterial and protozan cells in specific pore size classes was confirmed by partial chloroform fumigation and resin impregnated sections of soil.

The evolution of antibiotic resistance presents an escalating problem in the treatment of various infectious diseases worldwide. Although the origin of antibiotic resistance genes is not generally clearly documented, it has been thought that they evolved from specific genetic elements which eventually managed to spread to other microorganism of different strains and species through mobile genetics elements, transposons and plasmids. Extensively studying all aspects of these genes and their impact on the development of new treatments and drugs is of extreme importance. This study focuses on evolving and understanding how novel antibiotic resistance develops. Error prone PCR (EP-PCR) was used to introduce random mutation in an arr gene which confers high level resistance to rifampicin in E. coli. The clones obtained from EP-PCR were screened on different antibiotics with varying concentration in an attempt to isolate a clone with an increased minimum inhibitory concentration (MIC) as compared to the wild type parent strain (pBstN49). Several clones showed decreased levels of resistance against rifampicin but however none showed any significant increase in any of the other antibiotic MICs tested.

by Lai-chu Wu. / Bibliography: leaves 212-241 / Thesis (M.Ph.)--Chinese University of Hong Kong, 1984

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