Düsseldorf, Universiẗat, Diss., 2002.

Variovorax paradoxus EPS is a gram-negative rod isolated from the sunflower rhizosphere at CSUSB. Preliminary research has shown that Variovorax paradoxus EPS has anti-staphylococcal activity in liquid and solid co-culture. Anti-staphylococcal activity of Wild type and V. paradoxus EPS 𝚫4519 on 0.5% YE agar with embedded S. aureus AH1710 supports the idea that a soluble molecule is responsible for this activity, as the agar acted as a physical barrier between V. paradoxus EPS and S. aureus colonies. Preliminary genetic analysis of V. paradoxus EPS identified three loci that suitable candidates for the synthesis of a potential anti-staphylococcal small molecule. Preliminary data failed to detect expression at two of the three identified loci and a strain with a mutation at the third locus continues to produce anti-staphylococcal activity. We hypothesize that the microbial agent is expressed at a different locus or loci that have not yet been identified. These gene products are responsible for the synthesis of the microbial agent and are controlled by exposure to Staphylococcus aureus. Optimal growth conditions were identified for V. paradoxus EPS and S. aureus to demonstrate the formation of a zone of inhibition on Tryptic Soy Agar. The use of a V. paradoxus EPS Δ 4519 transposon library at optimal growth conditions allowed us candidate mutants with altered antimicrobial activity phenotypes. We identified 28 insertion sites that resulted in altered antimicrobial activities, which will allow us to identify the genes involved in this biosynthetic pathway.

Prions are self-perpetuating and, in most cases, aggregation-prone protein isoforms that transmit neurodegenerative diseases in mammals and control heritable traits in yeast. Prion conversion requires a very high level of identity of the interacting protein sequences. Decreased transmission of the prion state between divergent proteins is termed "species barrier" and was thought to occur due to the inability of divergent prion proteins to co-aggregate. Species barrier can be overcome in cross-species infections, for example from "mad cows" to humans. We studied the counterparts of yeast prion protein Sup35, originated from three different species of the Saccharomyces sensu stricto group and exhibiting the range of prion domain divergence that overlaps with the range of divergence observed among distant mammalian species. Heterologous Sup35 proteins co-aggregated in S. cerevisiae cells. However, in vivo cross-species prion conversion was decreased and in vitro polymerization was cross-inhibited in at least some heterologous combinations, thus demonstrating the existence of prion species barrier. Our data suggests that species-specificity of prion transmission is controlled at the level of conformational transition rather than co-aggregation. We have shown the Sup35 prion domain is sufficient for the species barrier among the S. sensu stricto species, and constructed SUP35 chimeric prion domains, combining the subregions of various origins Our data demonstrated in different cross-species combinations, different modules of prion domain play a crucial role in the controlling of species-specificity of prion transmission. One essential amino acid position has been identified in S. cerevisiae and S. paradoxus system. Our data support a model suggesting that identity of the short amyloidogenic sequences is crucial for the species barrier. Sup35 originated from three different species of the S. sensu stricto group were capable of forming a prion in S. cerevisiae. However, it was not known whether they are capable of generating and maintaining the prion state in the homologous cell environment. We have constructed the S. paradoxus and S. bayanus strains with appropriate markers, and we were able to demonstrate de novo [PSI+] formation in S. paradoxus but not in S. bayanus. Our data show that [PSI+] formation is not a unique property of S. cerevisiae.

The weathering of shales is a globally important process affecting both natural and built environments. Shales form roughly 70 % of worldwide sedimentary rock deposits and therefore the weathering of these rocks has substantial effects on the geochemical cycling of elements such as carbon, iron and sulfur. Microbes have been shown to play a key role in weathering shales, primarily through the oxidation of the iron and sulfur of embedded pyrite and the resultant production of sulfuric acid. Despite significant interest in the microbial weathering of shales within industrial sectors such as biohydrometallurgy and civil engineering, comparatively few studies have investigated microbial shale weathering in natural environments. Furthermore, the role of microbes in natural shale weathering processes beyond iron oxidation has largely remained unexplored. In this thesis, the weathering capabilities of microbial communities from natural weathered shale was investigated. The North Yorkshire coastline was used as a study location, due to the abundance and diversity of natural cliffs and historic, disused industrial sites. Cliff erosion and recession on the North Yorkshire coastline is a major concern for local authorities and is the focus of current research. The aim of this work has been to evaluate microbial shale weathering processes within these environments, and hypothesise the possible contribution they may have to erosive processes. Phenotypic plate assays inoculated with weathered shale material were used to obtain rock weathering bacterial isolates that tested positive for a specific weathering phenotype, such as iron oxidation or siderophore production. Subsequent 16S rRNA sequencing enabled genera level identification, revealing 15 genera with rock weathering capabilities with several being associated with multiple weathering phenotypes including Aeromonas sp., Pseudomonas sp. and Streptomyces sp. Shale enrichment liquid cultures were incubated with shale rock chips to simulate natural biological weathering conditions, and the concentration of rock-leached elements in the fluid measured. No evidence of microbially-enhanced leaching was found consistently for any element, however the significant reduction in leachate iron concentration under biological conditions indicates that iron precipitation occurred via microbial iron oxidation. Enrichment cultures inoculated with weathered shale and containing organic matter (OM) rich rocks in water or M9 medium, both liquids lacking an organic carbon source, were grown over several months. The cultures yielded microbial isolates that could utilise rock bound OM sources and one bacterial isolate, Variovorax paradoxus, was taken forward for ecophysiological study. The shale rock that the organism was isolated from, along with other OM rich rocks (mudstones and coals), elicited complex responses from V. paradoxus including enhanced growth and motility. Finally, mineral microcosms in vitro and mesocosms in situ investigated microbial colonization and weathering of shale-comprising minerals (albite, calcite, muscovite, pyrite and quartz). Microcosms were established using iron oxidizing enrichment cultures, as based on the results of the simulated rock weathering experiments, while the in situ mesocosms were buried within weathered shale scree within a disused mine level. Levels of colonization significantly varied between minerals within the microcosms (pyrite > albite, muscovite > quartz > calcite). Although differences in mineral colonization were seen in the mesocosms, they did not match those in the microcosms and were not statistically significant. Pyrite incubated in the microcosms became significantly weathered, with extensive pit formation across the mineral surface that is consistent with microbial iron oxidation. In the mesocosms, pit formation was not identified on pyrite surfaces but dark etchings into the pyrite surface were found underneath fungi hyphal growth. The results of this thesis highlights that a range of microbial rock weathering mechanisms are abundant across weathered shale environments. Microbial iron oxidizing activity was a dominant biogeochemical process that altered rock-fluid geochemistry and weathered pyrite surfaces. However, the impact on rock or mineral weathering of other microbial mechanisms was not elucidated by this work. Given the known capabilities of these mechanisms, the conditions under which they are active may not have been met within the experimental setup used. Microbial iron oxidation in shale and shale-derived materials has previously been demonstrated to weaken rock structure through acid production and secondary mineral formation. From the results of this thesis, it is clear that microbial iron oxidation is an active process within some of the weathered shale environments studied, including cliff surfaces. Therefore, it can be hypothesised that microbial activity could play a role in structurally weakening shale rock within cliffs and accelerate their erosion. Future work should attempt to quantify the rate and extent of microbial iron oxidizing activity within shale cliff environments and investigate its contribution to erosive processes.

The motivation for this study was to examine the exploitation patterns of the M. capensis and M. paradoxus hake fisheries on the Southeast Coast, and determine the size and species of hake caught in each of the hand-line, long-line, and trawl methods. The handline hake fishery has increased substantially over the last several years and concerns are beginning to emerge about the impact this will have on the inshore resource collected on the South Coast between August 1998 and July 1999 was used to describe the hand-line method and estimate annual landings. Data on the size and species in longline catches of hake caught during 1997 were already available for this study. Size distributions in trawl catches were determined from commercial category landing data reported by catch weight and depth. The species composition in these catches determined by comparison using RV Afrikana survey data collected in the same depth regions. Location plays a significant role in determining the sizes and species of hake caught by each gear. Hand-lines catch smaller sizes on average than do long-lines, inshore trawls target mainly M. capensis while offshore trawls catch both hake species. A substantial amount of the hand-line hake caught on the South Coast is not reported. Examination of the exploitation patterns reveal that intense trawling pressure is directed at the smaller sized M. paradoxus inhabiting the depth region between 160-400-meters. Inshore trawls discard a large amount of small sized M. capensis within the 100-meter isobath. A preliminary stock assessment on the status of each hake species found that M. paradoxus appears to be over-exploited while M. capensis was in better shape. However, length-based pseudo-cohort analysis, used in this assessment, is critically reliant on having length frequency data from a steady state population in equilibrium. This limits the application of this model for management purposes and this finding is purely theoretical at this stage. Results suggest that each hake species is under a different pattern and level of exploitation and the multi-species nature of hake stocks on the South Coast should be considered in developing optimum management policies. Future work should focus on developing appropriate age/length keys so that an age-based VPA, which is more powerful than the length-based approach, can be applied towards stock assessments on the South Coast. Alternatively, length-data covering a longer period should be compiled and the equilibrium assumption further investigated so that the results from length-based models can be used with more confidence.

Staphylococcus aureus is one of the leading causes of fatal nosocomial infections. Often, S. aureus can grow as a biofilm which protects the population from the surrounding environment. Strains of S. aureus are resistant to virtually all known antibiotics on the market. Variovorax paradoxus is a soil microbe with many unusual metabolic activities. It has been previously observed that, V. paradoxus can inhibit the growth of S. aureus when in co-culture. In this work we report on inhibition of S. aureus biofilm formation by V. paradoxus due to a suspected inhibitory soluble factor.

Thesis (MScAgric)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: Cape hake (Merluccius capensis and M. paradoxus) is commercially the most important trawl-caught fish off the South African, coastline and due to current intensive fish processing procedures Cape hake contributes the most to the total fishwaste production. Besides its commercial importance fish is also regarded as one of the single most important consumable natural resources, either in the raw or frozen form. Most of South Africa's commercially trawled demersal fish has already been partially cleaned (i.e. headed and gutted) before landing with non-marketable bycatch and hake-waste normally disposed of as discards, resulting in a waste of a potential protein source. This study was thus aimed at fulfilling several objectives namely: observing the current large-scale commercial Cape hake harvesting procedure; constructing prediction models for several morphological parameters (whole hake mass, headed & gutted hake mass, hake head mass, hake head length, hake head breadth and hake head height) of Cape hake (Merluccius ssp.), using whole hake length as the independent variable; and determining the chemical composition (moisture, protein, fat, ash, macro and trace elements) of several hake head sections (clean head, neck flesh, tongue, tongue cartilage, jaw, gills, heart, intestines, gut, kidney, kidney & kidney bone and gut & gall); determining the effect that storage has on the fatty acid profile of both the clean head and neck flesh sections. The results obtained would supply necessary data required for techno-economic investigations in the use of hake heads. For each of the six prediction models constructed, there was an increase in the variance of the data points of categories 3 (64-80 cm) and 4 (>80 cm) as opposed to categories 1 (30-46 cm) and 2 (47-63 cm). This could be attributed to a smaller sample set for both categories 3 and 4 or due to an expected increase in the variance when investigating larger biological samples. There was also a clustering of data in the three areas for each prediction model namely, within category 1 and across categories 2 and 3 and 3 and 4. This emphasised the latitudinal stratification of the Cape hake population by age, hence their stratification by size. The prediction models constructed for both boat trips 2 and 3 differed significantly (p<0.01) from that of boat trip 1, with the exception of the hake head length (cm) prediction model. The constructed prediction models, for each of the three respective boat trips, showed good predictive abilities as was indicated by the low Mean Square Error (MSE) values for the test sets, and high Pearson's correlation coefficient (r) values. These prediction models can be used in the fishing industry with confidence for Cape hake within the time frame each respective boat trip was carried out. The neck flesh could be regarded as the most important concerning chemical composition whereas the jaw could be seen as the most important when one considers mineral content. This therefore means that the jaw section, once appropriately processed is a potential Ca, Na and Fe source for supplementing diets of people suffering from a Ca, Na or Fe deficient diet. With regard to chemical status the neck flesh section is seen as a good potential source of both protein and fat, which could be attributed to the fact that hake muscle constitutes a major portion of this section. This section could thus be used to supplement the protein and fat of an existing food product, which is protein and fat deficient for people suffering from a protein and fat deficient diet. Similarly, a market could be created for the production of an economical food product with the neck flesh section being the main ingredient. Once this have been accomplished, fishing vessels may be persuaded to retain their Cape hake fish-waste for further processing due to the value of the prepared food products and thereby maintain profitability while abiding to governmental law. In conclusion non-government scientists should have more input in the decision-making process concerning matters affecting South Africa's marine biodiversity in order for future key policy and legislation drafts to be effective. Improvement of current fish preservation techniques and the known chemical composition of currently discarded material will result in informed decisions of future matters concerning its disposal. / AFRIKAANSE OPSOMMING: Kaapse stokvis (Merluccius capensis en M. paradoxus) is kommersieel Suid-Afrika se belangrikste vis spesie. Aangesien die Suid-Afrikaanse visprosesseringsbedryf baie intensief is, dra die Kaapse stokvis verwerkingsindustrie die grootste gedeelte by tot die totale visafval produksie. Die meeste van Suid-Afrika se visvangste word gedeeltelik skoongemaak voor landing terwyl nie-kommersiële byvangste en visafval gewoonlik oorboord gegooi word tydens die vangproses. Dit lei tot die vermorsing van 'n potensïele proteïen bron. Hierdie studie was dus gemik om: die huidige grootskaalse kommersiële Kaapse stokvis visvangsproses waar te neem; voorspellingsmodelle vir verskeie morfologiese parameters (heel vis massa, vis massa sonder kop en binnedele, stokvis kop massa, stokvis kop lengte, stokvis kop breedte en stokvis kop hoogte) vir Kaapse stokvis (Merluccius ssp.) te ontwikkel deur die hele lengte van die vis te gebruik as die onafhanklike veranderlike; die chemiese samestelling (vog, proteïen, vet, as, makro en spoor elemente) van verskillende dele van die viskop (skoonkop, nekweefsel, tong, tong kraakbeen, kaak, kiewe, hart, ingewand, derm, nier, nier & nierbeen en derm & gal); sowel as die effek van opberging op die vetsuurprofiel van beide die skoonkop en nekweefsel dele van die Kaapse stokvis kop. Hierdie resultate sal dan gebruik word vir die tegnies-ekonomies ondersoek in die gebruik van Kaapse stokvis koppe. Vir elk van die ses voorspellingsmodelle ontwikkel, was daar 'n vermeerdering in die variansie van die datapunte vir kategorieë 3 (64-80 cm) en 4 (>80 cm) teenoor kategorieë 1 (30-46 cm) en 2 (47-63 cm). Dit kan moontlik wees as gevolg van die kleiner monster trekking vir beide kategorieë 3 en 4 of as gevolg van verwagte toename in variansie wanneer groter biologiese monsters ondersoek word. Daar was ook 'n groepering van data in drie plekke vir elke voorspellingsmodel naamlik; binne in kategorieë 1 en oor kategorieë 2 en 3 en 3 en 4. Dit beklemtoon die geografiese breedte van die Kaapse stokvis populasie op grond van ouderdom, en dus die geografiese breedte op grond van grootte. Die voorspellingsmodelle ontwikkel vir beide die tweede en derde bootvangs het betekenisvol verskil (p<0.01) van die eerste bootvangs, behalwe die vir die stokvis kop lengte (cm) voorspellingsmodel. Die voorspellingsmodelle vir elk van die bootvangste het goeie voorspellingsvermoë getoon wat bewys is deur die lae Gemiddelde Kwadraat Fout waardes vir toetsgroepe en hoë Pearson's korrelasie koeffisiënt (r) waardes. Hierdie voorspellingsmodelle wat ontwikkel is, kan dus met vertroue in die Kaapse stokvis visvangsbedryf gebruik word mits dit ooreenstem met die periode waarin elke bootvangs uitgevoer was. Die nekweefsel gedeelte is die mees belangrikste met betrekking tot chemiese samestelling en die kaak die belangrikste in terme van minerale samestelling van die verskeie viskop dele. Die kaak is dus, as dit voldoende geprosesseer word, 'n goeie potensïele bron van Ca, Na en Fe en kan dus gebruik word om die dieet van mense wat 'n gebrek het aan hierdie minerale aan te vul. Met betrekking tot die chemiese samestelling van die nekweefsel gedeelte kan dit beskou word as 'n goeie potensiële bron van beide proteïen en vet, wat toegeskryf kan word aan die feit dat spierweefsel 'n groot deel uitmaak van hierdie viskop gedeelte. Hierdie viskop gedeelte sal dus uitstekend wees om die proteïen- en vetinhoud van 'n voedselproduk wat van nature 'n lae proteïen- en vetinhoud het te verhoog en hierdie produk sou dan geteiken word op daardie gedeelte van die gemeenskap wat 'n proteïen en vet tekort in hul dieet het. As dit eers alles in plek is, dan sal die visvangs bedryf hul Kaapse stokvis afval behou vir verdere prosessering deurdat dit gebruik word om die voedingsinhoud van bestaande voedsel soorte sal verbeter en terselfdertyd sal hulle aan wetgewing voldoen. Gevolglik sal nie-regerings navorsers meer betrokke moet wees by die besluitnemingsproses met betrekking tot sake wat die Suid-Afrikaanse mariene lewe affekteer en wat toekomstige wetgewing meer effektief sal maak. Die verbetering van huidige vis preserveringstegnieke gepaardgaande met die kennis van die chemiese samestelling van die Kaapse stokvis koppe sal lei na beter toekomstige besluite oor die afset daarvan.