Thesis (M.Tech.: Biotechnology)- Dept. of Biotechnology, Durban University of Technology, 2007 xv, 127 leaves / The human gut microbiota play a major role in host health, and attempts are being made to manipulate the composition of the gut microbiota-increase the composition of bacterial groups, such as lactobacilli and bifidobacteria that are perceived as exerting health promoting properties. These bacteria defined as food supplements (probiotics) beneficially affect the host by improving the intestinal microbial balance, and have been used to change the composition of the colonic microbiota. However, such changes may be transient, and the implantation of exogenous bacteria therefore becomes limited. In contrast, prebiotics are naturally occurring carbohydrates that are classified as non-digestible oligosaccharides present in edible plants. These carbohydrates enter the colon as intact compounds, elicit systemic physiological functions and act as fermentable substrates for colonic microflora-influencing the species composition and metabolic characteristics of intestinal microflora providing important health attributes. Currently, a widely marketed prebiotic, inulin is extracted from plants of the family Asteraceae. There are many unexploited plants that are regularly consumed and that may have a prebiotic effect or can have very high levels of inulin which could make them commercially viable. In this study, we investigated prebiotic compounds, especially inulin from locally growing, non-commercialised leafy plants. The aqueous extracts of 22 plants from the families Asparagaceae, Alliaceae, Asteraceae, Solanaceae, Cucurbitaceae, Amaranthaceae, Acanthaceae, Polygonaceae, Portulaceae, Fabaceae, Chenopodiaceae, Pedaliaceae and Apiaceae from Kwa-Zulu Natal were investigated for a prebiotic effect using a modified batch-culture technique with Lactobacillus bulgaricus, Lactobacillus lactis, Lactobacillus reuteri and Bifidobacterium longum, four common probiotics and the inulin content of the plants was determined using high performance liquid chromatography. Of the 22 plants studied, Solanum nigrum, Amaranthus spinosus, Amaranthus hybridus, Asystasia gangetica, Senna occidentalis, Cerathoteca triloba, Asparagus sprengeri, Tulbaghia violacea, Sonchus oleraceus and Taraxacum officinale exhibited a prebiotic effect. The prebiotic effect of the Taraxacum officinale, Sonchus oleraceus and Asparagus sprengeri extracts on L. lactis and L. reuteri was higher than or equivalent to inulin-a commercial prebiotic. In this study, Sonchus oleraceus exhibited the best prebiotic effect-was the only plant to stimulate all the probiotics including B. longum. Of all the plants analysed, Asparagus sprengeri tuber contained the highest amount of inulin (3.55%).

The Pacific Northwest is known for its once-abundant wild salmonid populations that have been in decline for more than 50 years due to habitat destruction and commercial overexploitation. To compensate, federal and state agencies annually release hundreds of thousands of hatchery-reared fish into the wild. However, accumulating data indicate that hatchery fish have lower fitness in natural environments, and that hatchery rearing negatively influences return rates of anadromous salmonids. Recently, mounting evidence revealed that the richness and diversity of intestinal microbial species influence host health. We examined the gut microbiota of pre-migratory hatchery-reared steelhead (Oncorhynchus mykiss) to assess microbial community diversity. The Cascade Mountains serve as an allopatric border between two distinct clades of steelhead that show significant differences in genomic and mitochondrial diversity. We identified differences in core microbiota of hatchery-reared fish that correlate with this divergent phylogeographic distribution. Steelhead sampled from hatcheries east of the Cascades had overall greater core gut microbiota diversity. These differences were found despite similarities in diet and rearing conditions. In addition to taxonomic variation across the geographic divide, we identified significant differences in metabolic pathways using PICRUSt gene prediction software. Our analysis revealed significant enrichment of genes associated with lipid metabolism in the gut microbiome of western fish. 8 of 19 individual lipid metabolism pathways were more prominent in western populations. Lipids are a vital nutritional component for teleost species involved in migration and subsequent return for spawning in natal environments. We hypothesize that the observed differences in lipid metabolism across this phylogenetic divide results from an increased ability of eastern Cascade (O. m. gairdneri) fish to utilize lipids taken in via the diet. This increased absorption and utilization would make lipids less available for the intestinal microbiota of the eastern fish, as evidenced by the lower abundance of lipid metabolism genes in the east. Our research utilizes information from the microbiome to understand the phenotypic implications occurring in segregated populations of hatchery-reared steelhead, further confirming elements of coevolution between an organism and its internal environment.

Thesis (M.Tech.: Biotechnology)- Dept. of Biotechnology, Durban University of Technology, 2007 xv, 127 leaves / The human gut microbiota play a major role in host health, and attempts are being made to manipulate the composition of the gut microbiota-increase the composition of bacterial groups, such as lactobacilli and bifidobacteria that are perceived as exerting health promoting properties. These bacteria defined as food supplements (probiotics) beneficially affect the host by improving the intestinal microbial balance, and have been used to change the composition of the colonic microbiota. However, such changes may be transient, and the implantation of exogenous bacteria therefore becomes limited. In contrast, prebiotics are naturally occurring carbohydrates that are classified as non-digestible oligosaccharides present in edible plants. These carbohydrates enter the colon as intact compounds, elicit systemic physiological functions and act as fermentable substrates for colonic microflora-influencing the species composition and metabolic characteristics of intestinal microflora providing important health attributes. Currently, a widely marketed prebiotic, inulin is extracted from plants of the family Asteraceae. There are many unexploited plants that are regularly consumed and that may have a prebiotic effect or can have very high levels of inulin which could make them commercially viable. In this study, we investigated prebiotic compounds, especially inulin from locally growing, non-commercialised leafy plants. The aqueous extracts of 22 plants from the families Asparagaceae, Alliaceae, Asteraceae, Solanaceae, Cucurbitaceae, Amaranthaceae, Acanthaceae, Polygonaceae, Portulaceae, Fabaceae, Chenopodiaceae, Pedaliaceae and Apiaceae from Kwa-Zulu Natal were investigated for a prebiotic effect using a modified batch-culture technique with Lactobacillus bulgaricus, Lactobacillus lactis, Lactobacillus reuteri and Bifidobacterium longum, four common probiotics and the inulin content of the plants was determined using high performance liquid chromatography. Of the 22 plants studied, Solanum nigrum, Amaranthus spinosus, Amaranthus hybridus, Asystasia gangetica, Senna occidentalis, Cerathoteca triloba, Asparagus sprengeri, Tulbaghia violacea, Sonchus oleraceus and Taraxacum officinale exhibited a prebiotic effect. The prebiotic effect of the Taraxacum officinale, Sonchus oleraceus and Asparagus sprengeri extracts on L. lactis and L. reuteri was higher than or equivalent to inulin-a commercial prebiotic. In this study, Sonchus oleraceus exhibited the best prebiotic effect-was the only plant to stimulate all the probiotics including B. longum. Of all the plants analysed, Asparagus sprengeri tuber contained the highest amount of inulin (3.55%).

Thesis (MSc)--University of Stellenbosch, 2007. / ENGLISH ABSTRACT: The tolerance of Enterococcus mundtii ST4SA to stressful gastro-intestinal conditions in humans and animals is vital to its success as a probiotic. The need for new effective probiotics with stronger inhibitory (bacteriocin) activity has arisen due to the increasing number of antibiotic resistant pathogens. Enterococci are used in the fermentation of sausages and olives, cheese making and as probiotics. Their role as opportunistic pathogens in humans makes them a controversial probiotic (Moreno et al., 2005). Enterococci occur naturally in the gastro-intestinal tract which renders them intrinsic acid and bile resistance characteristics. E. mundtii ST4SA produces a 3950 Da broad-spectrum antibacterial peptide active against Gram-positive and Gram-negative bacteria, and viruses. The bacteria include Enterococcus faecalis, Streptococcus spp., Pseudomonas aeruginosa, Klebsiella pneumoniae, Streptococcus pneumoniae and Staphylococcus aureus. E. mundtii ST4SA inactivates the herpes simplex viruses HSV-1 (strain F) and HSV-2 (strain G), a measles virus (strain MV/BRAZIL/001/91, an attenuated strain of MV), and a polio virus (PV3, strain Sabin). This study focuses on the genetic stability of E. mundtii ST4SA genes when exposed to stress factors in the human and animal gastrointestinal tract. Based on results obtained by real-time PCR, the expression of genes encoding bacST4SA, RecA, GroES and 23S rRNA by E. mundtii ST4SA were not affected when the cells were exposed to acid, bile and pancreatic juice. This suggests that these genes of E. mundtii ST4SA will remain stable in the intestine. This could indicate that other genes of E. mundtii ST4SA could remain stable in the host. Further studies on the stability of genes encoding antibiotic resistance and virulence factors should be conducted to determine their stability and expression in the host in stress conditions. Concluded from this study, E. mundtii ST4SA is an excellent probiotic strain. / AFRIKAANSE OPSOMMING: Enterococcus mundtii ST4SA se weerstandsvermoë teen stresvolle gastrointestinale kondisies is essensieel vir die sukses van hierdie organisme as ‘n probiotikum. Die aanvraag vir nuwe, meer effektiewe probiotika met sterker inhibitoriese (bakteriosien) aktiwiteit is as gevolg van die toename in antibiotikum weerstandbiedende patogene. Enterococci word algemeen gebruik as probiotika, sowel as in die fermentasie van worse, olywe en kaas. Hulle rol as oppertunistiese patogene in mense veroorsaak kontroversie as gevolg van hul toenemende gebruik as probiotika. Enterococci is deel van die natuurlike mikroflora in die gastrointestinale weg van mense en diere. Dit verleen aan hierdie spesies ‘n natuurlike weerstandsvermoë teen maagsure, galsoute en pankreatiese afskeidings. E. mundtii ST4SA produseer ‘n 3950 Da wye spektrum anti-bakteriese peptied, aktief teen Gram positiewe en Gram negatiewe bakterieë sowel as virusse. Hierdie bakterieë sluit Enterococcus faecalis, Streptococcus spp., Pseudomonas aeruginosa, Klebsiella pneumoniae, Streptococcus pneumoniae en Staphylococcus aureus in. E. mundtii ST4SA inaktiveer die herpes simpleks virus HSV-1 en HSV-2, ‘n masels virus (MV/BRAZIL/001/91), en ‘n polio virus (PV3, stam Sabin). Hierdie studie fokus op die genetiese stabiliteit van E. mundtii ST4SA gene, wanneer hulle blootgestel word aan stress faktore in die mens en dier gastrointestinale weg. “Intydse” PKR data gebasseer op die uitdrukking van die bacST4SA, RecA, GroES en 23S rRNA gene in stresvolle kondisies dui aan dat E. mundtii ST4SA nie geaffekteer word wanneer die sel blootgestel word aan suur, gal en pankreatiese vloeistowwe nie. Hierdie resultate dui aan dat hierdie gene van E. mundtii ST4SA stabiel sal bly in die intestinale weg van die mens en dier. Dit kan aandui dat ander gene van E. mundtii ST4SA soos die wat kodeer vir virulensie faktore en antibiotikum se weerstandsvermoë stabiel mag bly in die gasheer. Verdere studies wat fokus op die stabiliteit van gene wat kodeer vir antibiotikum weerstandbiedendheid en virulensie faktore moet uitgevoer word om hulle stabiliteit en uitdrukking in die gasheer te bepaal. Bevindings van hierdie studie dui aan dat E. mundtii ST4SA goeie potensiaal het as ‘n probiotikum.