Agricultural production in Monterey County, California is a multi-billion dollar industry. Near the coast, seawater intrusion has threatened to degrade the groundwater quality due to over-pumping of the aquifer. The Monterey Regional Water Pollution Control Agency (MRWPCA), in partnership with the Monterey County Water Resources Agency, has provided recycled water since 1998 to over 12,000 acres of prime agricultural farmland in the northern Salinas Valley in an effort to reduce groundwater removal. The dominant soil types in the region are clay loam and clay soils, which are both susceptible to sodium (Na) accumulation and water infiltration problems. Recycled water blended with well water is used to irrigate cool season vegetables (i.e., artichokes, broccoli, Brussels sprouts, celery, cauliflower, and lettuce) and strawberries. A long-term study was implemented by MRWPCA to monitor salinity levels in commercial vegetable fields because of grower concerns that salts in the recycled water would have long term effects on soil quality. Accumulation of salts over time would make the soil less productive. Soil salinity levels were monitored at three Control and three Test Sites beginning in the spring of 2000. The Control Sites received well water, and the adjacent Test Sites received an approximate 2:1 blend of recycled and well water, respectively. Control and Test Sites were paired based on location to compare the same soil, crop, drainage systems, and farming practices. The soil was sampled three times per year from all sites: spring (before planting), mid-summer after harvest of the first crop, and late fall after the second crop harvest. Composites of four cores were collected at each site from the zero to 36-inch depth at 12-inch intervals. Each 12-inch interval soil sample was analyzed for pH, electrical conductivity (ECe), extractable cations (Na+, Ca2+, Mg2+, and K+) and extractable anions (Cl-, NO3-, and SO4-). After 10 years of monitoring, the data showed that using recycled water blended with well water at the Test Sites increased the ECe of the soil profile from 2.1 to 2.5 dS/m and increased the sodium adsorption ratio (SAR) from 3.0 to 3.9. The data also showed that using well water at the Control Sites increased the ECe of the soil profile from 1.4 to 2.6 dS/m and the change in SAR was negligible. The Test and Control Sites were significantly different for ECe and SAR, which was expected considering a higher salt content in the recycled water compared to the well water. The significant differences for ECe and SAR were associated with the significant differences in soil Na+ levels between the Test and Control Sites. The SAR and ECe of soil samples from all sites were in a range acceptable for vegetable production. The use of recycled water for irrigation of cool season vegetables and strawberries in the study area has not shown an indication of degraded soil productivity. Based on vegetable production and the slow increase of salts in the soil, recycled water can be used for long-term irrigation with proper management.

High levels of elements such as sodium (Na), potassium (K) and phosphorus (P) in iron ore minerals are known to reduce the quality and price of these minerals. South Africa, as one of the world largest exporter of iron ore, is affected by this problem. Both potassium (K) and phosphorus (P) are peculiar to South African iron ore. The present study has therefore focussed on developing an environmentally friendly biological method for lowering the levels of K and P in iron ore minerals. Short and long term experiments were set up to isolate, identify, screen and test potential bioleaching bacteria and fungi from different environmental samples. The study started by investigating the possible relationship that exists between weathering and bioleaching processes. The investigation was intended to provide relevant information on the natural role of microorganisms such as ectomycorrhizal (ECM) fungi in the mining environment. The experiments involved the use of both mycorrhizal and non-mycorrhizal Pinus patula seedlings for the weathering of iron ore minerals. Four types of ECM fungi were used, namely Pisolithus tinctorius (PT), Paxillus involutus (PI), Laccaria bicolor (LB) and Suillus tomentosus, (ST). From the results, ectomycorrhizal weathering can be said to be species-specific and significantly influenced by fungal type and particle size. In addition, it was also discovered that both mycorrhizal and non-mycorrhizal roots can participate in weathering processes. Further investigations of ECM fungi when not in symbiosis, were carried out to know how or if they can be potential candidates to mobilise K and P from iron ore minerals. The experimental set up involved in vitro pure cultures of four different ECM fungi, namely Pisolithus tinctorius (PT), Paxillus involutus (PI), Phialocephala fortini (PFR), and Suillus tomentosus (ST). In addition, the treatments involved the use of five different particle sizes of each ore type. The results obtained indicated the potential of the ECM fungi to mobilise P and K from the two iron ore types though at different levels. Factors such as ore type, particle size, organic acid production and attachment of the fungi to the iron ore were all found to influence the mobilisation of nutrients from these ores. Another experiment that addressed some of the limitations encountered with the use of pure cultures of ECM fungi was conducted. Isolated indigenous fungal pure cultures from the surfaces of iron ore minerals were screened for their abilities to solubilise minerals by lowering the levels of K and P. These isolates were identified molecularly as close relatives of three genera that included Penicillium, Alternaria (2 isolates) and Epicoccum for isolates FO, SFC2/KFC1 and SFC2B respectively. The identified Penicillium sp. turned out to be the only phosphate solubiliser among these isolates. Direct bioleaching capability of the fungus was compared to that of its metabolite. At the end, the metabolite showed better K removal than the direct use of the fungi. Interpretation of these results indicates possible relationship between K and P removal, and the organic acids production by this fungus. Other factors such as particle size and mineral type were also found to significantly influence the leaching process. Additional experiment was conducted to investigate the indigeous bacteria and their potentials in reducing the K and P contents of iron ore minerals. A total of 23 bacterial strains that belong to Proteobacteria, Firmicutes, Bacteroidetes and Actinobateria were isolated from the iron ore minerals and identified with molecular methods. All the bacterial isolates were screened for their potential as mineral solubilisers. Only eight of the isolates were selected and used in shake flask experiments that contained both KGT and SK mineral types as their sources of K and P. The experiment showed that all the eight isolates have potentials to produce organic acids especially high levels of gluconic acid but lower quantities of acetic, citric and propanoic acid. Scanning electron microscopy (SEM) and fourier transform infrared (FITR) analyses also helped to uncover the role that biofilm and extracellular polymeric substances could play in mineral solubilisation. Finally, an investigation of a new method for reduction of K and P levels of iron ore minerals was carried out, focussing on the use of cheap resources as well as septic conditions. The study involved the use of fermented spoilt grape fruits (Vitis sp.) and the solution from the product utilised in shake-flask experiments. Treatments involved two types of iron ore minerals (KGT and SK) and two different particle sizes. The result suggests the significant effect of particle size, time and organic acids on the reduction of K and P from the iron ore minerals. The important part of this finding is the discovery of a cheap microbial energy source (spoilt grape) that can be further exploited for full biobeneficiation of iron ore minerals. Another advantage of this method is the fact that the experiment can be conducted under non–sterile conditions, making it a system that can be operated outdoor. / Thesis (PhD)--University of Pretoria, 2010. / Microbiology and Plant Pathology / unrestricted

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El trabajo realizado en la Tesis Doctoral llega a las siguientes conclusiones:1.- T. halophila muestra una relación en peso raíz/parte aérea mayor que A. thaliana, y esta diferencia se ve incrementada en condiciones de estrés salino, lo que podría suponer una ventaja para afrontarlo.2.- El estrés salino produce en A. thaliana mayores reducciones en la absorción y en las concentraciones internas de K+ que en T. halophila, a la vez que T. halophila presenta menor absorción de Na+ y transporte a la parte aérea que A. thaliana. Ambas circunstancias resultan en una mayor relación K+/Na+ en T. halophila, lo que puede suponer una mayor tolerancia a la salinidad.3.- El gen ThHAK5 codifica para un transportador que media un transporte de K+ de alta afinidad en levaduras similar al observado en las plantas de T. halophila lo que sugiere que este transportador juega un papel fundamental en la absorción de K+ en el rango de la alta afinidad en esta especie vegetal.4.- Aunque AtHAK5 y ThHAK5 presentan una gran homología de secuencia y unas características funcionales similares, la regulación de los genes que los codifican difieren en condiciones salinas. Así, la salinidad reduce en menor medida la inducción de ThHAK5 por ayuno de K+. En consecuencia, la absorción de K+ de alta afinidad está menos afectada por la presencia de NaCl en el medio externo en T. halophila.5.- La mutagénesis al azar permite encontrar aminoácidos importantes para la función de las proteínas y ésta ha permitido identificar dos versiones mutantes del transportador de K+ de alta afinidad AtHAK5 más eficientes, capaces de transportar K+ a concentraciones externas de Na+ muy elevadas (0.1 mM K+ y 800 mM Na+). / The work done in this Thesis provides some interesting conclusions:1.- Thellungiella halophila show a weight ratio root/shoot bigger than Arabidopsis thaliana, and this difference arise under salt stress, what might provide an effective mechanism of salt tolerance to T. halophila.2.- In A. thaliana, salt stress induces a bigger reduction of K+ uptake and tissue concentrations than in T. halophila, and at the same time T. halophila shows a reduced Na+ uptake and Na+ transport to the shoot. Both properties enable a higher ratio K+/Na+ in T. halophila which might be another mechanism of salt tolerance. 3.- The ThHAK5 gene isolated in this Thesis, encode a K+ transporter that mediates high affinity K+ transport in Saccharomyces cerevisiae similar to the observed in intact plants of T. halophila, which suggest a key role of this transporter in the high affinity range of concentrations.4.- Although AtHAK5 and ThHA5 shows high sequence homology and similar functional properties, gene regulation is different under salt stress. Thus, salinity reduces to a lesser extent the K+-starvation ThHAK5 induction. As a consequence, high affinity K+ uptake is less affected by NaCl in T. halophila. 5.- Random mutagenesis allows the identification of important aminoacids for protein function, and with this technique two more efficient mutant versions of AtHAK5 have been isolated. The evolved AtHAK5 mutant versions are able to transport K+ at high Na+ external concentrations (0.1 mM K+ and 800 mM Na+) in yeast.