Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: In order to test the effect of different plant growth-promoting strategies on Triticum aestivum L. (wheat), we investigated the ability of biochar and a grain-associated soil yeast, to improve the growth of this crop. Our first goal was to study the effect of biochar amendments to sandy soil on the growth and nutrition of wheat in the presence of mycorrhizal fungi. This was accomplished by amending soil with 0%, 1%, 2.5%, 5% and 10% (w/w) biochar and cultivating wheat plants in these soil-biochar mixtures. After harvesting, plant growth and mycorrhizal colonization of roots were measured. In addition, we studied the nutritional physiology of these plants with regards to nitrogen (N), phosphorous (P) and potassium (K) concentrations, as well as the growth efficiencies and uptake rates of these nutrients. We found that wheat growth was improved by biochar amendments to soil, probably as a result of elevated K levels in the plant tissues supplied by the biochar amendments. The second goal of this study was to obtain a soil yeast from the rhizosphere of another monocot in the family Poaceae, i.e. Themeda triandra Forssk. (red grass), and then evaluate this isolate for its ability to improve wheat performance. Three different Cryptococcus species were isolated from the rhizosphere of wild grass, i.e. Cryptococcus zeae, Cryptococcus luteolus and Cryptococcus rajasthanensis. Since C. zeae was previously isolated from maize, an isolate representing this species was selected to be used in further experimentation. With the ultimate goal of testing the ability of this yeast to improve wheat growth, its effect on wheat germination was investigated and compared to that of two other soil yeasts, i.e. Cryptococcus podzolicus CAB 978 and Rhodotorula mucilaginosa CAB 826. These three yeasts were subsequently tested for their ability to improve wheat growth in pot cultures in a greenhouse. After one and two months of growth, the culturable yeasts present in the rhizosphere and bulk soil were enumerated. The effects of these yeasts were elucidated by measuring wheat growth in terms of dry weight, as well as root and shoot relative growth rates (RGR). Changes in wheat nutrition were evaluated by determining the concentrations, growth efficiencies and uptake rates for P, K, zinc (Zn) and iron (Fe). During this study, it was found that only C. zeae CAB 1119 and C. podzolicus CAB 978 were able to enhance seed germination. Similarly, it was shown that C. zeae CAB 1119 was able to improve wheat growth during the first and second month of cultivation, whereas C. podzolicus CAB 978 only improved growth during the first month, and R. mucilaginosa CAB 826 had no effect on growth. This improved growth could be attributed to C. zeae CAB 1119 improving the P, K, Zn and Fe growth efficiency of wheat, which positively influenced the root and shoot RGR, and subsequently wheat growth. Our final goal was to test whether C. zeae CAB 1119 could affect wheat growth and nutrition when cultivated in sandy soil, which contained natural microbial consortia and 10% (w/w) biochar. Plants treated with viable or autoclaved cells of C. zeae CAB 1119, were subsequently cultivated in soil only or soil amended with biochar. After one month, plants were harvested and growth was measured with regards to dry weight, root RGR and shoot RGR. In addition, the concentrations of P, K, Zn and Fe were analyzed for these plants, where after the growth efficiencies and uptake rates were calculated for these four nutrients. Results indicated that plants growing in soil amended with biochar, and treated with viable C. zeae CAB 1119, showed the best growth. The increased root and shoot RGR witnessed in these plants was probably due to increased concentrations of P and K in the plants. This study opens new avenues of research with regards to the bio-fertilizers of wheat. / AFRIKAANSE OPSOMMING: Die uiteindelike doel van die studie was om die effek van verskillende plantgroei bevorderende metodes op die groei van Triticum aestivum L. (koring) te ondersoek. Dus het ons die vermoë van houtskool en ‘n graan-geassosieerde grondgis getoets om die groei van dié plant te bevorder. Die eerste doel van die studie was om die effek van houtskool toedienings tot sanderige grond te evalueer. Dit is bewerkstellig deur 0%, 1%, 2.5%, 5% en 10% (w/w) van die houtskool by die sand toe te voeg en koring in die houtskool-sand mengsels te kweek. Na die verlangde groei tydperk is die koring geoes en die mikorrizale kolonisasie op en in die koring wortels bepaal. Gedurende hierdie studie is die effek van bogenoemde toedienings op die fisiologie van die plante ondersoek deur die konsentrasies, opname tempo’s, en groei ekonomie van die plante vir stikstof (N), fosfaat (P) en kalium (K) te bepaal. Ons het gevind dat die groei van koring deur die toediening van houtskool bevorder is en dit blyk dat dié effek weens die teenwoordigheid van hoë K vlakke in die plantweefsel is. Die tweede doel van ons studie was om ‘n gis vanuit die risosfeer van ‘n monokotiel wat aan die familie Poacea behoort, naamlik Themeda triandra Forssk. (rooigras) te isoleer. Die vermoë van die isolaat om die groei van koring te bevorder was daarna getoets. Drie verskillende Cryptococcus spesies was vanuit die risosfeer van rooigras geïsoleer, nl. Cryptococcus zeae, Cryptococcus luteolus en Cryptococcus rajasthanensis. Omdat C. zeae in ‘n vorige studie vanaf mielies geisoleer was, is ‘n isolaat van hierdie spesie gebruik in verdere eksperimente. Met die doel om te bepaal of dié gisspesie koringgroei kan bevorder, was die effek van C. zeae op die ontkieming van koring bestudeer en vergelyk met dié van twee ander grond giste, nl. Cryptococcus podzolicus CAB 978 en Rhodotorula mucilaginosa CAB 826. Hierdie drie giste is ook ondersoek om die groei van koring in ‘n glashuis te bevorder. Na een en twee maande se groei was die getalle van giste teenwoordig in die risosfeer en grond verder weg van die wortels bepaal. Die effek van dié giste op die groei van koring is bepaal in terme van droë gewig asook die relatiewe wortel en halm groei tempos. Veranderinge in die nutrient status van koring is ondersoek deur die konsentrasies, groei-ekonomie en tempo van opname vir P, K, sink (Zn) en yster (Fe) te bepaal. Ons het gedurende dié studie gevind dat C. zeae CAB 1119 en C. podzolicus CAB 978 die ontkieming van koring kon verbeter. Ons het ook gevind dat C. zeae CAB 1119 die groei van koring gedurende die eerste en tweede maand van groei kon bevorder, terwyl C. podzolicus CAB 978 dit net gedurende die eerste maand kon vermag en R. mucilaginosa CAB 826 geen effek gehad het nie. Die verbeterde groei kon aan C. zeae CAB 1119, wat die P, K, Zn en Fe groei effektiwiteit van die plante verbeter het, toegeskryf word. Die verbetering van groei effektiwiteit het ‘n positiewe invloed op die relatiewe groeisnelheid van die wortels en halms gehad, en dus op koringgroei. Die laaste doel van die studie was om te bepaal of C. zeae CAB 1119 die groei van koring kon bevorder wanneer die koring in sand wat natuurlike mikrobiese populasies bevat en met houtskool aangevul is, gekweek word. Plante is met lewensvatbare of nielewensvatbare selle van C. zeae CAB 1119 behandel en gekweek in sanderige grond, en/of grond waarby 10% (w/w) houtskool toegevoeg is. Die plante is na een maand geoes en die groei bepaal in terme van droë massa en die relatiewe wortel en halm groei tempos. Die konsentrasies van P, K, Zn en Fe in die plante, asook die fisiologie van die plante, nl. groei ekonomie en tempo van opname, met betrekking tot P, K, Zn en Fe is bepaal, Ons het gevind dat plante wat in die houtskool-grond mengsel gekweek is en met lewensvatbare selle van C. zeae CAB 1119 behandel is die beste groei getoon het. Die verbeterde relatiewe groei tempos van die wortels en halms was mees waarskynlik die gevolg van verhoogde P en K konsentrasies in die plante. Hierdie studie toon nuwe resultate in verband met die gebruik van biologiese alternatiewes tot kunsmis.

Yeasts form significant and important part of pedosphere microbiota. They keep nature balance, participate in cycles of elements and nutrients, are antagonists of various pathogens and as important decomposers, they produce the whole spectrum of different extracellular enzymes. The aim of this study was to determine the ability of yeasts, isolated from the soil adjacent to the fruit trees in Southwest Slovakia as well as from the contaminated soil (Pernek area, Slovakia), to produce extracellular enzymes. In total, 68 strains belonging to 45 different species were tested for the production of starch-like polysaccharide and for extracellular enzyme activities: polygalacturonases, lipases, proteases, cellulases, chitinases, -glucosidases and -amylases. This work was also focused on optimization of method for the yeast chitinase assay. Four methods were proved; two of them utilized liquid medium with chitin (colloidal and insoluble) as the sole carbon source and two others used solid plate methods with agar medium containing chitin. Based on results, cultivation in colloidal liquid chitin medium, terminated by the chitinase assay according to Ehrlich, was evaluated as the best method for detection of predominant exochitinase activity of yeasts. More than 75 % of tested yeasts exhibited some extracellular activity. Generally, the yeasts isolated from the soil under the fruit trees showed broader spectrum of enzyme activities than those originated from contaminated soils. Lipases, proteases and -glucosidases were found to be the most common activities. Only small proportion of yeasts was able to produce chitinases and/or cellulases. Aureobasidium pullulans, CCY 27-1-134, from the soil adjacent to the apple tree, showed the widest range of activities from all tested strains and it possessed all examined activities. On the other side, it did not produce starch-like polysaccharide. Tausonia (Trichosporon) pullulans and Cystofilobasidium macerans were the second most active producers of extracellular enzymes with variations in production of cellulases and -amylases. Representatives of the former polyphyletic genus Cryptococcus exhibited lipases, -glucosidases, -amylases and they were producers of the starch, but the interspecies differences were also noted. All strains of the genus Galactomyces were positive for polygalacturonases and the genera Candida and Cyberlindnera were positive for -glucosidases. All strains of Galactomyces candidum were tested for the production of polygalacturonases during 168 hours long cultivation on pectin media. Strain CCY 16-3-4 showed very stable growth on this medium and simultaneously exhibited significant amount of extracellular polygalactouronases. It has a potential to be very suitable producer of these enzymes but particular characterization of properties is necessary for its future use. Results of the screening showed that the production of extracellular enzymes is mostly strain-dependent and not species-dependent.