Masters Degrees (Microbiology)

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Microbial biotransformation of kimberlite ores.
(2008) Ramcharan, Karishma.
Microbial leaching plays a significant role in the natural weathering of silicate containing ores such as diamond-bearing kimberlite. Harnessing microbial leaching processes to pre-treat mined kimberlite ores has been proposed as a means of improving diamond recovery efficiencies. The biomineralization of kimberlite is rarely studied. Therefore, this study investigated the feasibility of exploiting both chemolithotrophic and heterotrophic leaching processes to accelerate the weathering of kimberlite. Preliminary investigations using mixed chemolithotrophic leaching cultures were performed on four finely ground kimberlite samples (<100μm) sourced from different mines in South Africa and Canada. Mixed chemolithotrophic cultures were grown in shake flasks containing kimberlite and inorganic basal media supplemented either with iron (Fe2+, 15g/l) or elemental sulfur (10g/l) as energy sources. Weathering due to dissolution was monitored by Inductive Coupled Plasma (ICP) analyses of Si, Fe, K, Mg and Ca in the leach solutions at known pH. Structural alterations of kimberlite after specified treatment times were analyzed by X-ray Powder Diffraction (XRD). The results of the preliminary investigation showed that weathering can be accelerated in the presence of microbial leaching agents but the degree of susceptibility and mineralogical transformation varied between different kimberlite types with different mineralogical characteristics. In general, the results showed that the kimberlite sample from Victor Mine was most prone to weathering while the sample from Gahcho Kue was the most resistant. It was therefore deduced that kimberlite with swelling clays as their major mineral component weathered relatively more easily when compared to kimberlite that consisted of serpentine and phlogopite as their major minerals. Gypsum precipitates were also distinguished indicating that a partial alteration in the kimberlite mineralogical structure occurred. Both energy sources positively influenced the dissolution process, with sulfur producing superior results. This was attributed to the generation of sulfuric acid which promotes cation dissolution and mineral weathering. Success in the preliminary investigations led to further experimental testing performed to determine the effect of particle size and varying energy source concentrations on the biotransformation of kimberlite. It was observed that although weathering rates of the larger kimberlite particles (>2mm<5mm) were lower than that of the finer particles, slight changes in their mineralogical structures represented by the XRD analyses were seen. Optimisation studies of energy source concentration concluded that although the highest concentration of elemental sulfur (20% w/w) and ferrous iron (35% w/w) produced the most pronounced changes for each energy source tested, the leaching efficiency at these concentrations were not drastically greater than the leaching efficiency of the lower concentrations, as expected. Following the success of batch culture shake flasks weathering tests, the effect of continuous chemolithotrophic cultures on the biotransformation of larger kimberlite particles (>5mm<6.7mm) was investigated. A continuous plug-flow bioleach column was used to model the behaviour of chemolithotrophic consortia in a dump- or heap leaching system. Two sequential columns were setup, in which the first consisted of kimberlite mixed with sulfur and the second purely kimberlite. Inorganic growth medium was pumped to the first column at a fixed dilution rate of 0.25h-1 and the leachate from the first column dripped into the second. After an 8 week investigation period, the ICP and XRD data showed that weathering did occur. However, the pH results showed that the leaching process is governed by the amount of acid produced by the growth-rate independent chemolithotrophic consortia. Data from pH analyses also showed that the leaching bacteria reached ‘steady state’ conditions from day 45 onwards. The pH also remained higher in the second column than in the first column highlighting the alkaline nature of the kimberlite ores and its ability to act as a buffering agent and resist weathering. This important factor, as well as further optimisation studies in process operating conditions and efficiency, needs to be considered when establishing heap-leaching technology for these kimberlite ores. In the preliminary heterotrophic investigation, Aspergillus niger was used to produce organic metabolites to enhance kimberlite mineralization. The results demonstrated that the organic acid metabolites generated caused partial solubilization of the kimberlite minerals. However, it was deduced that for more significant changes to be observed higher amounts of organic acids need to be produced and maintained. The results obtained in this study also showed that the type of kimberlite presents a different susceptibility to the dissolution process and the presence of the fungal cells may improve the leaching efficiency. The results in this study provided an optimistic base for the use of microbial leaching processes in accelerating the weathering of kimberlite. These findings may also serve to supply data to formulate recommendations for further and future column microbial leach tests as well as validation and simulation purposes.
Application of bacterial bioflocculants for wastewater and river water treatment.
(2008) Buthelezi, Simphiwe P.; Pillay, Balakrishna.
Dyes are often recalcitrant organic molecules that produce a colour change and contribute to the organic load and toxicity of textile industrial wastewater. Untreated effluent from such sources is harmful to aquatic life in the rivers and lakes due to reduced light penetration and the presence of highly toxic metal complex dyes. The use of alum as flocculant/coagulant in wastewater treatment is not encouraged as it induces Alzheimer’s disease in humans and results in the production of large amounts of sludge. Therefore, the development of safe and biodegradable flocculating agents that will minimize environmental and health risks may be considered as an important issue in wastewater treatment. Bioflocculants are extracellular polymers synthesized by living cells. In this study, bacterial bioflocculants were assessed for their ability to remove dyes from textile wastewater as well as reducing the microbial load in untreated river water. The bacteria were isolated from a wastewater treatment plant and identified using standard biochemical tests as well as the analysis of their 16S rDNA gene sequences. Six bacterial isolates were identified viz. Staphylococcus aureus, Pseudomonas plecoglossicida, Pseudomonas pseudoalcaligenes, Exiguobacterium acetylicum, Bacillus subtilis, and Klebsiella terrigena. The flocculating activities of the bioflocculants produced by these isolates were characterized. The effect of temperature, pH, cations and bioflocculant concentration on the removal of dyes, kaolin clay and microbial load was also determined. The amount of bioflocculants produced by the bacterial isolates ranged between 5 and 27.66 g/l. According to the findings of the present study, bacterial bioflocculants were composed of carbohydrates, proteins, uronic acid, and hexosamine in varying quantities. The bioflocculants were effective to varying degrees in removing the dyes in aqueous solution, in particular whale dye, medi-blue, fawn dye and mixed dyes, with a decolourization efficiency ranging between 20-99.9%. Decolourization efficiency was influenced by the bioflocculant concentration, pH, temperature, and cations. The bacterial bioflocculants were also capable of reducing both the kaolin clay and the microbial load from river water. The flocculating activity ranged between 2.395–3.709 OD-1 while up to 70.84% of kaolin clay and 99% of the microbial load from the river water was removed. The efficiency of kaolin clay flocculation increased with higher concentration of bacterial bioflocculants. The optimum pH for the flocculating activity was observed between 6 and 9. The best flocculating activity was observed at 28oC. Divalent cations such as Mg2+ and Mn2+ improved the flocculation while salts such as K2HPO4, CH2COONa, and Na2CO3 did not. The findings of this study strongly suggest that microbial bioflocculants could provide a promising alternative to replace or supplement the physical and chemical treatment processes of river water and textile industry effluent.
Optimizing biocontrol of purple nutsedge (Cyperus rotundus).
(2006) Brooks, Michael John.; Laing, Mark Delmege.
Cyperus rotundus L. CYPRO (purple nutsedge) and Cyperus esculentus L. CYPES (yellow nutsedge) are problematic weeds on every continent. At present there is no comprehensive means of controling these weeds.. The primary means of control is herbicides, although the weeds are becoming more resistant. Bioherbicide control of purple and yellow nutsedge is an important avenue of research, with much of the focus being to increase the virulence of current fungal pathogens of C. rotundus and C. esculentus. The primary aim of this study was to increase the virulence of a fungal pathogen of C. rotundus and C. esculentus, with the objective of creating a viable bioherbicide. A possible means of increasing the virulence of a pathogen would be to increase the amount of amino acid produced by the fungus. This was proposed as a means of increasing the virulence of Dactylaria higginsii (Luttrell) M. B. Ellis. Overproduction of amino acids such as valine and leucine result in the feedback-inhibition of acetolactate synthase (ALS), an enzyme which is a target for many herbicides currently on the market. By applying various amino acids to tubers of purple nutsedge and comparing the results with a reputable herbicide, glyphosate, it was possible to determine the success of the amino acid applications. Only glutamine treatment at 600 mg.r1 resulted in significantly less (P
Humic acid pretreatment for enhancing microbial removal of metals from a synthetic 'wastewater'.
(2004) Desta, Tsegazeab Goje.; Wallis, Frederick Michael.
The presence of heavy metal ions in waste streams is one of the most pervasive environmental issues of present times. A rotating biological contactor (RBC) was used to investigate the potential capacity of microbial biofilms in remediation of the metal ion species from a mixed metal contaminated effluent solution containing Cr+3 , Pb+2 and Cu+2 , each at a concentration of 200 mg r1 • In the first part of this study the effectiveness of various support materials for the development of microbial biofilms capable of removing heavy metals from a synthetic effluent was investigated. EDX analysis showed that none of the support matrices investigated, viz. gravel, polyester batting and sand, adsorbed metal ions on their surfaces; hence, metal adsorption was due purely to microbial activities. The biofilms attached more firmly and uniformly to polyester batting than to gravel and sand. The characteristics of polyester batting which made it a superior support matrix were its surface roughness and porous hydrophilic nature, which provided a larger surface area for the adhesion of microorganisms and attraction of nutrients during the biofilm development process. The selective accumulation of metal ion specIes by various microbial populations grown as biofilm using polyester batting as support matrix in separate compartments of a single-stage RBC bioreactor was examined. Lead ions were readily accumulated by almost all the microbial biofilms tested. Fungus-dominated biofilms selectively accumulated chromium ions whereas biofilms comprising mainly bacteria more readily accumulated copper ions from the mixed metal contaminated effluent solution. However, where interactions between the bacterial and fungal components were encouraged the mechanical stability of the biofilms was enhanced so that large amounts of all three metal ion species were removed by this biofilm. The combined effect of a series of bench-scale columns containing liquid humic acid and a three stage RBC bioreactor on the removal of metal ion species from a mixed metal contaminated effluent was investigated. After seven days of treatment the combined system had removed approximately 99% of the Cr+3, 98% of the Pb+2 and 90% of the Cu+2 ions from the mixed metal contaminated synthetic effluent. Complexation of the metal ions with humic acid was the predominant factor accounting for approximately 68-86% Cr+3 , 70-86% Pb+2 and 53-73% Cu+2 removal levels within the columns. A large proportion of the remaining Cr+3 and Pb+2, but not of the Cu+2, was removed in compartment 1 of the RBC. This suggested that the presence of the former two metals in solution might have reduced the removal of the Cu+2 ions from the system. The removal of substantially large amounts of the competing ions chromium and lead during the initial stages of the treatment process meant that copper was successfully taken up in the second and third RBC compartments. Hence, the economy of the treatment process was improved as larger quantities of the metal ions were removed in a shorter period of time than was possible when using the individual treatments (humic acid-metal complexation and biofilm adsorption) separately. More than 75%,92% and 86% of the adsorbed Cr+3 , Pb+2 and Cu+2 ions, respectively, were recovered from the three RBC bioreactor compartments following repeated washing of the biofilms with 0.1 M HCI. This relatively easy desorption suggested that the metal ions were simply adsorbed onto the surfaces of the biofilm cells rather than being taken into the cytoplasm of the cells.
The effect of water treatment residues on soil microbial and related chemical properties.
(2003) Pecku, Shantel.; Hunter, Charles Haig.; Hughes, Jeffrey Colin.
Water treatment residue (WTR), a by-product of the water treatment process, consists primarily of precipitated hydroxides of the coagulants used in the water treatment process, along with sand, silt, clay, humic compounds, and dissolved organic matter. It is usually disposed of by landfill, a technology with numerous problems that include dwindling landfill capacity, extensive dewatering requirements for the WTRs, high costs of transportation, and potential liability for landfill clean-up. Therefore, land disposal (or land treatment) presents a popular alternative disposal method based on the principle that the physical, chemical, and microbial properties of the soil can be used to assimilate applied waste without inducing any negative effects on soil quality. The objective of this study was to investigate the effects of land disposal of the WTR generated by Umgeni Water, a local water treatment authority, on soil quality. These effects were investigated using depth samples from soil profiles of Westleigh and Hutton soil forms at field trials located at Ukulinga Research Farm, near Pietermartizburg and Brookdale Farm, Howick, KwaZulu-Natal, South Africa, respectively. Four rates of WTR (0, 80, 320, and 1280Mg ha-1 incorporated into the soil) were investigated at both trials, in addition to mulched treatments at rates of 320 and 1280Mg ha-1 at Brookdale only. Sampling of plots was carried out in September 2001 and May 2002, and all treatments were investigated under fallow and grassed cultivation. Laboratory measurements used to assess soil quality included pH, electrical conductivity (EC), organic carbon (QC), and microbial activity using f1uorescein diacetate (FDA) hydrolysis. At both trials in September 2001 WTR-amended plots displayed higher pH in the 0-200mm soil in comparison to the controls, whereas by May 2002 pH had returned to the condition of the controls. Addition of WTR at Ukulinga resulted in higher QC in September 2001, but in May 2002 this was similar to the controls. However, at Brookdale QC was unaffected by WTR. At Ukulinga and Brookdale the effect of WTR on EC was variable, and microbial activity in the soil profile was unaffected by WTR addition. Observations at Ukulinga and Brookdale reflected long term changes (3 and 5 years, respectively) to soil quality following WTR addition. To examine the initial changes in soil quality a laboratory experiment was set up using the field trial soils. Research objectives were also extended to include WTRs from Rand Water (Johannesburg), Midvaal Water Company (Stilfontein), Amatola Water (East London), and two samples from the Faure Water Treatment Plant (near Cape Town). The second Faure sample (Faure2 ) was collected when blue green algal problems were experienced at the plant. The measurements used to investigate these short term effects on soil quality were soil pH, EC, and microbial activity as indicated by respiration rate. Each of the WTRs added to the Hutton and Westleigh soils increased soil pH by varying increments, and the higher the WTR application rate, the higher was the pH recorded. With the exception of the Rand and Umgeni WTRs that clearly increased soil EC, the effect of the otherWTRs on EC was variable. The Faure1 and Amatola WTRs appeared to have no effect on microbial activity, whereas the Umgeni, Rand, Midvaal, and Faure2 WTRs stimulated microbial activity by Day 2 following the addition of WTR, but this had declined by Day 14. As for pH, higher microbial activity was recorded at higher WTR application rates. Changes in microbial community structure of the Hutton soil only, following the addition of WTR were examined using denaturing gradient gel electrophoresis (DGGE) analysis. Community profiles of the different WTRs proved to be markedly different. However, WTR-amended soil retained banding patterns consistent with the control soil indicating that dominant populations in the Hutton soil had been retained. The field trials indicated that long term effects of land disposal of WTR were not detrimental to the measured indicators of soil quality namely, pH, EC, QC, and microbial activity. The laboratory assessments of the short term response of the Hutton and Westleigh soil forms to WTR addition suggested that the tested variables were altered by WTR, but not significantly changed to the detriment of soil quality. Microbial community analysis indicated that the community structure of the Hutton soil was not significantly altered by WTR amendments. Present findings provide no evidence to suggest that land disposal of WTR is detrimental to soil quality. It is therefore regarded as a feasible disposal option although there are some aspects that should be investigated further. These include investigations into rhizosphere/microbial interactions and the feasibility of growing cash crops.
Characterization of selected Bacillus isolates exhibiting broad spectrum antifungal activity.
(2004) Tewelde, Teklehaimanot Weldeslasie.; Hunter, Charles Haig.; Beukes, Mervyn.
The genus Bacillus is comprised of Gram-positive, rod-shaped, spore-forming bacteria which are well known for their ability to produce a diverse array of antimicrobial compounds. Ofparticular interest is the ability of certain strains to produce antifungal compounds. Such organisms have the potential for application in agriculture where they can be used as biocontrol agents against selected plant pathogenic fungi. A study was undertaken to further characterize selected Bacillus isolates that exhibit broad spectrum antifungal activity. Dual culture bioassays were used to screen seven selected Bacillus isolates for activity against four plant pathogenic fungi in vitro. All isolates were able to inhibit the pathogens to varying degrees. Two isolates, R29 and B81, were selected for further testing and characterization. Further bioassays were performed on five complex nutrient media which were adjusted to pH S.S and 7, and both incubated at 2SoC and 30°C" respectively. It was found that pH and media composition showed significant influences on the antifungal activities of the isolates tested, but that a SoC temperature difference in incubation temperature did not. Tryptone soy agar was found to give rise to the largest inhibition zones. Both isolates were tentatively identified using standard biochemical and morphological tests. Based on its phenotypic characteristics, R29 was identified as a strain of B. subtilis. B81 proved to be more difficult to assign to a specific group or species of Bacillus, though B. subtilis and B. licheniformis were considered to be the nearest candidates. Genomic DNA was extracted from both isolates and a portion of each of their 16s rDNA genes were amplified and sequenced for homology testing against the GeneBank database. Homology testing confirmed that both isolates were members of the genus Bacillus and most probably strains of B. subtilis. The DNA fragment used for sequencing proved to be too small to give conclusive identification of the isolates. Isolate R29 was selected for further characterization of its antifungal compound/so Growth curve studies using a defined synthetic medium showed that antifungal activity arose during the stationary phase and appeared to be closely linked to sporulation. The antifungal component of cell free culture supematant was extracted using various methods including thin layer chromatography, acid precipitation, hydrophobic interaction chromatography and methanol extractions. High performance liquid chromatography (HPLC) analysis of extracts from acid precipitation and hydrophobic interaction chromatography revealed two active peaks indicating that at least two antifungal compounds were produced. Methanol extracted samples produced the cleanest sample extract but only revealed one active peak from the HPLC fraction . Nuclear magnetic resonance analysis of purified samples indicated that the antifungal compound/s have aromatic complex and peptide structures. The extracted antifungal compounds were Protease K resistant and found to be thermostable at temperatures ranging 80-121oC, and, were active at pH ranges of 3-13. The antifungal compounds were found to exhibit similar properties to known antifungallipopeptides i.e. iturin A and fengycin A and B. Further characterization and identification of the active compounds is recommended usmg methods such as liquid chromatography mass spectrometer and matrix-assisted laser desorption ionisation time-of- flight. The results presented in this dissertation provide a basis from which antifungal compounds produced by strains ofBacillus can be further characterized.
Isolation and identification of antibiotic producing microorganisms from natural habitats in the KwaZulu-Natal midlands.
(2001) Okudoh, Vincent Ifeanyi.; Wallis, Frederick Michael.
The search for new antibiotics continues in a rather overlooked hunting ground. In the course of screening for new antibiotic-producing microorganisms, seventy-nine isolates showing antimicrobial activity were isolated from soil samples from various habitats in the KwaZulu-Natal midlands, South Africa. Existing methods of screening for antibiotic producers together with some novel procedures were reviewed. Both modified agar-streak and agar-plug methods were used in the primary screens. The use of selective isolation media, with or without antibiotic incorporation and/or heat pretreatment, enhanced the development of certain actinomycete colonies on the isolation plates. Winogradsky's nitrite medium (Winogradsky, 1949), M3 agar (Rowbotham and Cross, 1977), and Kosmachev's medium (Kosmachev, 1960), were found to be selective for actinomycetes. Statistical analysis showed highly significant interactions between isolates, assay media and the test organisms. The diameters of inhibition zones were found to be larger on Iso-sensitest agar (ISTA)[Oxoid, England] than in nutrient agar plates. Of the 79 isolates that showed antimicrobial activity, 44 isolates were selected for confirmatory screening. Of these, 13 were selected for secondary screening. Criteria for selection were based on significant inhibition of at least two test organisms and/or the inhibition of the specifically targeted organisms, Pseudomonas and Xanthomonas species. Following secondary screening eight isolates were considered for further investigation. The isolates were tentatively identified . on the basis of morphological features, using both light microscopy and scanning electron microscopy(SEM); their ability to utilize various carbon sources; and selected physiological and staining tests. Suspected actinomycetes were further characterized on the basis of selected chemical properties using thin layer chromatography (TLC) and high pressure liquid chromatography (HPLC) techniques. High pressure liquid chromatography analysis (Beckman 6300 analyzer) detected the presence of diaminopimelic acid (DAP) in whole-cell hydrolysates of six of the isolates while TLC analysis confirmed the type ofDAP present. The isolates N2, N12, N16, N19 and N35 were tentatively identified as Thermomonospora, Saccharopolyspora, Nocardiodes, Corynebacterium and Promicromonospora, respectively. Isolate N30 was identified as belonging to the coryneform group ofbacteria, possibly an Arthrobacter species. Isolate, N8, tentatively identified as Actinosynnema, was unique among the isolates tested as it showed good antimicrobial activity against all the Gram- positive and Gram-negative bacteria, and yeasts used as test organisms in the present investigation.
Cost-benefit analysis of the environmental impacts of Darvill Wastewater Works, Pietermaritzburg, KwaZulu-Natal.
(2002) Sikhakhane, Sindisiwe S.; Ahmed, Fethi B.; Darroch, Mark Andrew Gower.
Darvill Wastewater Works (DWWW) receives and treats both domestic and industrial wastewater from the city of Pietermaritzburg, in KwaZulu-Natal. Sludge from the wastewater treatment is sprayed onto surrounding lands, causing odour and fly problems. The plant also discharges treated effluent into the Msunduzi River, compromising water quality. This study uses several economic valuation techniques to estimate the value of the benefits of improving air and water quality to overcome these problems caused by DWWW. The benefits. are then compared with the costs of upgrading DWWW to see whether or not upgrading DWWW to improve air and water quality would be worthwhile. The Contingent Valuation Method (CVM) was used to elicit people's willingness to pay (WTP) for improvements in air quality due to the elimination of odours and flies caused by sludge deposited by DWWW. The WTP estimates reflect individual's preferences for improvements in air quality. The stated WTP amounts were positively related to household income, but negatively related to the age and gender of the respondent and the number of dependants in the household. The mean monthly WTP for the surveyed households is higher for those that are closer to the pollution source (R23.00 and R29.00 for Zones land 2) and less for those further away (RI4.00 for Zone 3). Sobantu residential area had the lowest mean monthly WTP (R18.00), followed by Lincoln Meade (R27.00) and Hayfields (R54.00). This is expected, as Sobantu has relatively high levels of unemployment and lower household incomes. Strategic, hypothetical and free rider bias may have led to the unexpected signs of some estimated regression coefficients in linear regression models used to estimate WTP. The mean WTP was estimated as R307.20 per annum per household, and when this is aggregated over the total population in the residential areas impacted by odours and flies (37192 households), the benefits of eliminating odours and flies are estimated as R11 425 382.00 per annum. A hedonic price method was used to quantify the decline in property values as a result of odours and flies caused by sludge deposited by DWWW. Properties experienced a R6650.08 decline in selling price if the distance from them to DWWW is decreased by one kilometre. Properties that are closer to DWWW were worth RI5 953.90 less than those further away from DWWW. Aggregating these values over all estimated impacted households in the study, gives an estimated benefit of improving air quality of R28 480 518.00 per annum. The impact of water pollution was quantified by estimating the revenue (R3 744 975.00) that would be lost by Pietermaritzburg if the Duzi Canoe Marathon were to be cancelled due to incidences of diarrheoa reported during the race. A cost of illness procedure was adopted to quantify the effect of water pollution on the health of communities that use the Msunduzi River as a source of potable water supply. A value of R1 243 372.50 was estimated as the annual cost of water-related illnesses in these rural areas. This value represents the costs of the river pollution to those communities. Both of these exercises indicated that improving water quality of the Msunduzi River would be beneficial to society. The effect of nutrient enrichment of the Msunduzi River was quantified by estimating the cost of removing water hyacinth from the Inanda Dam, treatment cost at Wiggins water treatment works and the value of recreation at Mahlabathini Park (Inanda Dam). The annual cost of removing water hyacinth was estimated from the direct costs of chemicals and labour as R47 202.15. The increased treatment costs at Wiggins attributable to DWWW were estimated as R1 104 999.20 and R956 924.15 per annum for removal of algae, and tastes and odours, respectively. The value of R706.90 per annum was estimated as the consumer surplus accruing to recreationists, and, therefore, the value of recreation at Mahlabathini Park to an individual. These annual benefits, when aggregated over the total study population (296 590) were over two hundred million rands (R209 659 470.00). The estimated total benefits (R256 662 840.00) of eliminating odours and flies and effluent problems were compared to the actual costs of two alternative methods of upgrading DWWW using cost-benefit analysis. These alternatives were co-disposal option (R170 473 320) and a land disposal option (R168 809377). Benefit-cost ratios of 1.51 and 1.52 suggest that from society's standpoint, it would be beneficial to upgrade the plant in order to eliminate its adverse environmental impacts. The study results have important implications for policy makers, both the DWWW management and the Pietermaritzburg-TLC municipality. At present DWWW is operating beyond its design capacity, and this problem, together with the poor status of Pietermaritzburg's reticulation system, causes overflow of untreated or compromised final effluent into the Msunduzi River during rainy seasons. These problems also impact on the efficient operation of the plant as the sludge is not properly digested before being sprayed onto surrounding land. Thus to prevent further environmental degradation, a fundamental basis of the National Environmental Management Act, DWWW would need to address these issues. Upgrading DWWW would be a short-term solution if the problems with the storm-runoff into the plant is not addressed.
Molecular and physiological characterization of thiosulphate-oxidizing microbial associations prior to use in hydrogen sulphide biofiltration.
(2000) Laughlin, Jamie B. A.; Senior, Eric.
Interacting microbial associations capable of utilizing thiosulphate as an energy source were enriched/isolated from activated sludge, landfill site [mal covering soil and soil from an acid mine water drainage site. The isolates were designated Lf-I, Ws-2 and Am-3, respectively. Although hydrogen sulphide was the target molecule for gas biofiltration, thiosulphate, which is a key oxidized intermediate, was used in this study due to the difficulty of working with a toxic gas. Together with thiosulphate oxidation, the microbial associations were assessed for their abilities to oxidize dissolved sulphide to elemental sulphur. Physiological analyses (temperature, pH and substrate concentration optimization) were made with closed and open cultures while morphological characterization and species compositional changes were monitored by light and scanning electron microscopy (SEM). To investigate further functional and structural responses to physiological changes, denaturing-gradient gel electrophoresis (DGGE) separation of PCR-amplified 16S DNA gene fragments and Biolog GN microtitre plates were used. The associations were found to be active metabolically between 0 and 35°C, 15 and 50°C, and 15 and 45°C, with optimum temperatures of 25, 40 and 35°C for Lf-l, Ws-2 and Am-3, respectively. The optimum pH range for microbial association Lf-l was between 3 and 4. The maximum specific growth rates of associations Lf-l , Ws-2 and Am-3 were 0.08, 0.06 and 0.03 h~l , respectively. Components of all three Gram negative rod-dominated associations were motile and displayed anaerobiosis. During open culture cultivation the species complement of Lf-l , as determined by morphological analysis, changed. The same association oxidized sulphide (40 ppm) to sulphur although Ws-2 and Am-3 did not have this capacity. Biolog GN plates detected pH-effected species compositional changes in Lf-l and these were confirmed by DGGE. The same technique showed that enrichment had occurred in the Biolog GN wells. Species composition changes also resulted in response to different pH values (2 to 9), temperatures (5 to 40°C) and dilution rates (0.003 to 0.09 h-1 ), but activity changes were not always accompanied by population profile changes.
The development of an in vitro system to assess the effect of arbuscular mycorrhizal fungi on cereal crops in KwaZulu-Natal, South Africa.
(2010) Govender, Avrashka.; Jamal-Ally, Sumaiya Faizal.
Cereal crops such as maize and sorghum are economically important in South Africa (SA) as a staple food diet. In order to meet the needs of South Africa’s growing population, higher yields in crop production need to be attained. However, the two major stress factors that affect yield production and require primary attention are nutrient deficiencies and pest infestations. Research is now being focused on certain endophytes that have become a valuable tool for agriculture as they protect crops against the above-mentioned stresses. The endophyte focused on in this study was Arbuscular Mycorrhizal fungi (AMF). This research was aimed at developing an in vitro culture system for SA cereal crops to enable interaction studies of endophytes. This dissertation is divided into two parts; the first part focused on the development of an in vitro culture system, the assessment of sorghum plant growth and exudate production in the presence of the Glomus intraradices strain. The results indicated that sorghum produces the required root exudates in the second stage of growth. Using high pressure liquid chromatography with mass spectrometry (HPLC/MS), it was noted that sorghum produced phytochemicals as chemoattractants for the respective endophytes. However, it was documented that when the plant underwent certain stresses they produced exudates, which acted as phytotoxic compounds that destroyed symbiotic organisms around sorghum rhizophere. The second part focused on optimization of the surface sterilization of maize seeds. The results indicated that maize contained unidentified endophytes, which negatively affected plant development. Surface sterilization of maize seeds was accomplished. The successful in vitro development can be used for future use to study plant development. Understanding plant development and interaction with symbiotic endophytes would not only be of great benefit but would also make it easier to create a biocontrol agent in vitro, which would bring about high crop yields at cost-effective prices and would be less labour intensive.
Molecular characterisation and detection of xanthomonas albilineans, the sugarcane leaf scald pathogen.
(1994) Permaul, Kugenthiren.; Pillay, Balakrishna.; Pillay, D.
No abstract available.
Application of image analysis in microecophysiology research : methodology development.
(1998) Dudley, B. T.; Wallis, Frederick Michael.
Rehabilitation of landfill sites is important for successful land utilization. Revegetation is one key element of the process since it can overcome aesthetic problems. The inimical challenges of landfill leachate and gas are largely responsible for the difficulties associated with the revegetation of completed sites. Many components of landfill leachate can be catabolized by microbial associations thereby reducing their impacts on the environment. The importance of research on interactions between pollutants, microorganisms and soil is its applicability in environmental risk assessment and impact studies of organic pollutants which enter the soil either accidentally or intentionally. The application of image analysis with microscopy techniques to landfill soil-pollution interactions provides a means to study surface microbiology directly and to investigate microbial cells under highly controlled conditions. This research focused on the development of a method to study the real time processes of attachment, establishment, growth and division of microbial cells/associations in site covering soils. Image analysis provides a powerful tool for differential quantification of microbial number, identification of morphotypes and their respective responses to microenvironment changes. This minimal disturbance technique of examining visually complex images utilizes the spatial distributions and metabolic sensitivities of microbial species. It was, therefore, used to examine hexanoic acid catabolizing species, both free-living and in a biofilm, with respect to obviating the threat of hexanoic acid to reclamation strategies. The three sources of inoculum (soil cover, soil from the landfill base liner and municipal refuse) were compared for their ability to provide associations which catabolized the substrate rapidly. During the enrichment programme the inocula were challenged with different concentrations of hexanoic acid, a common landfill intermediate. From the rates at which the substrate was catabolized conclusions were drawn on which concentration of hexanoate facilitated the fastest enrichment. The results of initial batch culture enrichments confirmed that the soil used contained microbial associations capable of catabolizing hexanoic acid at concentrations < 50mM, a key leachate component. Exposing the landfill top soil microorganisms to a progressive increase in hexanoic acid concentration ensured that catabolic populations developed which, in situ, should reduce the phytotoxic threat to plants subsequently grown on the landfill cover. The analysis of surface colonization was simplified by examining the initial growth on newly-exposed surfaces. The microbial associations generated complex images which were visually difficult to quantify. Nevertheless, the dimensional and morphological exclusions which were incorporated in the image analysis software permitted the quantification of selected components of the associations although morphology alone was inadequate to confirm identification. The effects of increasing the dilution rate and substrate concentration on the growth of surface-attached associations in Continuous Culture Microscopy Units (CCMUs) were examined. Of the five dilution rates examined the most extensive biofilm development (9.88 jum2) during the selected time period (72h) resulted at a dilution rate of 0.5h' (at 10mM hexanoic acid). The highest growth (608 microorganisms.field"1) was recorded in the presence of 50mM hexanoic acid (D = 0.5h"1). To ensure that the different morphotypes of the associations were able to multiply under the defined conditions a detailed investigation of the component morphotypes was made. Numerically, after 60h of open culture cultivation in the presence of 50mM hexanoic acid, rods were the predominant bacterial morphotypes (43.74 field'1) in the biofilms. Both rods and cocci were distributed throughout the CCMUs whereas the less numerous fungal hyphae (0.25 field'1) were concentrated near the effluent port. The specific growth rates of the surface-attached associations and the component morphotypes were determined by area (//m2) colonized and number of microorganisms.field"' and compared to aerobic planktonic landfill associations. From area determinations ( > 0.16 h'1) and the number of microorganisms.field"1 10mM hexanoic acid was found to support the highest specific growth rate ( > 0.05 h"1) of the surfaceattached association isolated from municipal refuse. With optical density determinations, the highest specific growth rate (0.01 h'1) was recorded with 25mM hexanoic acid. The surface-attached microbial associations component species determinations by area and number showed that the hyphae had the highest specific growth rate ( > 0.11 h"1). The surface-attached microbial association specific growth rate determinations from the discriminated phase (0.023 h'1), area colonized (0.023 h"1) and number of microorganisms (0.027 h"1) calculated from the results of the component species rather than the association should give more accurate results. The specific growth rate obtained differed depending on the method of determination. Any one of these may be the "correct" answer under the cultivation conditions. Depending on the state (thickness) of the association (free-living, monolayer or thick biofilm) the different monitoring methods may be employed to determine the growth. As a consequence of the results of this study, the kinetics of microbial colonization of surfaces in situ may be subjected to the same degree of mathematical analysis as the kinetics of homogeneous cultures. This type of analysis is needed if quantitative studies of microbial growth are to be extended to surfaces in various natural and artificial environments.
Installation, commissioning and preliminary microbiological and operational investigations of full-scale septic tank digestion of sewage.
(1997) Taylor, Michelle Anne.; Senior, Eric.
This study investigated the commissioning and maintenance of a Pennells two-tank bioreactor system with specific reference to its application in rural areas of KwaZulu- Natal, South Africa to treat sewage and generate biogas. The septic tank configuration was installed in a community which lacked electricity and domestic waste disposal. An artificial wetland was constructed at the outlet of the system to facilitate further treatment. Inefficient operation and maintenance of the system occurred due to various social/community-related problems which are typical of a field- and community-based project of this nature in a rural region of a Third World African country. These problems affected both maintenance and digester performance. The Pennells system was characterized by incomplete anaerobiosis which limited methanogenesis. Despite this, and attendant problems of low temperatures and elevated pH values, COD removal resulted. Laboratory-scale batch cultures, in conjunction with fluorescence and scanning electron microscopy, were used to identify a suitable anaerobic digester sludge for inoculation purposes. Perturbation experiments with locally used detergents and toxic compounds demonstrated the inimical effects of these agents. In contrast, low concentrations of penicillin and tetracycline promoted methanogenesis. Further analysis with light, fluorescence and scanning electron microscopy identified the acidogens as the predominant bacterial species, whilst fluorescence microscopy confirmed the absence of methanogens in the bioreactor.
Exploitation of indigenous fungi in low-cost ex situ attenuation of oil- contaminated soil.
(1997) McGugan, Brandon Ross.; Senior, Eric.
The central aim of this study was to determine if indigenous fungi of an oil-contaminated soil could be effectively used in a low-cost bioremediation of the soil. Since some of the contaminant had been present at the site for over two decades, the indigenous microbial species had been subjected to specific selection pressures for a protracted period, thus facilitating key enzymatic capabilities for hydrocarbon degradation. Analysis of the pertinent influential parameters of soil bioremediation indicated that an ex situ technique, utilising the catabolic activities of the indigenous soil fungi, was a feasible low-cost option. Fungi were isolated from the contaminated soil through a variety of techniques. The abilities of these isolates to degrade the contaminant oil and a range of representative hydrocarbon molecules was evaluated by a systematic screening programme. Sixty-two isolates were initially examined for their growth potential on hydrocarbon-supplemented agar. A bioassay, utilising hydrocarbon-impregnated filter paper discs, was then used to examine the abilities of 17 selected isolates to catabolise three representative hydrocarbon molecules (hexadecane, phenanthrene and pristane) in different concentrations. In the same bioassay, the influence of a co-metabolite (glucose) on growth potential was also examined. Eight fungal species: Trichophyton sp.; Mucor sp.; Penicillium sp.; Graphium sp.; Acremoniwn sp.; Chaetomium sp.; Chrysosporium sp.; and an unidentified basidiomycete were then selected. Liquid batch cultures with a hydrocarbon mixture of hexadecane, phenanthrene, pristane and naphthalene facilitated quantitative analysis (HPLC) of the hydrocarbon catabolic abilities of the selected isolates. Ex situ bioremediation was evaluated at laboratory-scale by both bioaugmentation and biostimulation in soil microcosm trials. During the course of the study, total petroleum hydrocarbon (TPH) concentration (U.S. EPA Method 418.1) was used as a simple and inexpensive parameter to monitor hydrocarbon disappearance in response to soil treatments. Soil microbial activities were estimated by use of a fluorescein diacetate hydrolysis bioassay. This was found to be a reliable and sensitive method to measure the activity of respiring heterotrophs as compared with the unreliable data provided by plate counts. In the bioaugmentation trial, the eight selected isolates were individually used to inoculate (30% v/v) the contaminated soil. The highest rate of biodegradation (50.5% > than the non-sterile control) was effected by an Acremonium species after 50 days incubation (25°C). The second highest rate of biodegradation (47% > than the non-sterile control) was achieved with a soil treatment of sterile barley/beer waste only. Comparable rates of hydrocarbon degradation were achieved in simple biostimulation trials. Thus, due to its lower cost, biostimulation was the preferred remediation strategy and was selected for further laboratory investigation. Common agricultural or industrial lignocellulosic wastes such as: wood chips; straw; manure; beer brewery waste; mushroom compost; and spent mushroom substrate were used as soil treatments, either alone or in combination. The effect of the addition of a standard agricultural fertiliser was also examined. The highest level of biodegradation (54.4% > the non-sterile control) was recorded in a microcosm supplemented (40% v/v) with chicken manure. Finally, an ex situ bioremediation technique was examined in a pilot-scale field trial. Wood chips and chicken manure were co-composted with the contaminated soil in a low-cost, low-maintenance bioremediation system know as passive thermal bio venting. Extensive monitoring of the thermal environment within the biopile was made as an indirect measure of microbial activity. These data were then used to optimise the composting process. Three-dimensional graphical representations of the internal temperatures, in time and space, were constructed. From these graphs, it was determined that an inner core region of approximately 500 cm3 provided a realistic simulation of conditions within a full-scale biopile. During this trial a TPH reduction of 68% was achieved in 130 days. The findings of this research suggested that the utilisation of fungal catabolism is applicable to soils contaminated with a wide range of hydrocarbon contaminants. Passive thermal bioventing offers a bioremediation strategy which is highly suitable for South African conditions in terms of its low level of technological sophistication, low maintenance design and, most importantly, its relatively low cost.
An Epidemiological study of gentamicin resistant gram negative bacteria with particular reference to pseudomonas aeruginosa at King Edward V111 Hospital, Durban
(1985) Bhana, Ratilal Hargovind.
The sources of gentamicin resistant pseudomonads and enterobacteria were studied in detail. A total of 1703 gentamicin resistant gram negative bacilli (GRGNB) isolated from patients, staff and their immediate environment were studied over a 6 month period . Of these 954 were isolated from clinical specimens obtained from patients and 540 from their immediate environment. A furthur 209 stains were isolated from the staff members who were responsible for the care of these patients. Pseudomonas aeruginosa; pyocin type 1 phage type F7 and .serotype 11 was the commonest isolate. It constituted 24,9% of all isolates in this study. This organism was distributed in all the wards investigated and was isolated throughout the 6 month study period. This strain, therefore, appears to be part of the "resident'' flora of King Edward Vlll Hospital for it was found on patients, staff and their immediate environment. Among the Enterobacteriaceae, Klebsiella pneumoniae was the commonest isolate and made up 13,6 % of all isolates. All the isolates obtained in this study were resistant to five of more antibiotics tested (gentamicin, tobramycin, kanamycin, streptomycin, carberricillin, polymyxin B amikacin and sisomicin). Of 310 staff members screened 25,2% harboured GRGNB on their hands. Among patients the commonest source of GRGNB was stool which yielded 141 (14,8 %) of the clinical isolates. Of the environmental sources studied, sinks harboured 87 (14%) GRGNB. The isolates from the environment and staff members were identical to patient strains. The significance of these findings is discussed.
Rapid prediction of multi-drug resistance in clinical specimens of Mycobacterium tuberculosis.
(2011) Ndimande, Bongiwe Olga.; Pillay, Manormoney.
Conventional drug susceptibility testing techniques, the ‘gold standard’ for M. tuberculosis are slow, requiring about 3-6 weeks from a positive culture. This diagnostic delay, before initiation of appropriate treatment, contributes to increased transmission rates. Molecular techniques provide rapid results and therefore present an alternative to conventional tests. The aim of this project was to develop an inhouse reverse line blot hybridization assay (RIFO assay) that could detect mutations associated with Rifampicin resistance directly in clinical specimens of patients in KwaZulu Natal. A 437 bp region of the rpoB gene was sequenced to ascertain the most frequently occurring mutations conferring resistance to rifampicin in isolates in KwaZulu-Natal. Wildtype and mutant probes designed to target these mutations, were immobilized on a Biodyne C membrane. Hybridization conditions were optimized using biotin labeled PCR products from culture. Detection was performed with peroxidase labeled streptavidin using enhanced chemiluminescence. Four DNA extraction methods were evaluated on sputum specimens to determine the one with the least inhibitory effect on amplification. A total of 11 mutations were found in 236 clinical isolates: 531TTG (109, 58.3%), 516GTC (26, 13%), 533CCG/516GGC (20, 10%), 533CCG (18, 9.6%), other mutations < 5% each. The chelex extraction method was found to be optimal for removing inhibitors in sputum specimens. Sputum specimens of 404 patients hospitalized at King George V Hospital between 2005 and 2006 were rifoligotyped. The RIFO assay was optimised on clinical isolates and then applied to sputum specimens. The RIFO assay on culture and sputum correlated well with the DST (sensitivity 92% and 94% respectively). However, the specificity was very low in both culture and sputum specimens compared to DST (38% and 35% respectively). This could be attributed to the presence of silent mutations, mixed infections, mixed populations of bacteria or the small number of susceptible strains used in this study. The in-house RIFO assay can be used directly on sputum specimens to predict Rifampicin resistance and therefore MDR-TB in less than a week compared to the gold standards. A total of 43 samples can be tested simultaneously at low cost and the membrane is reusable compared to commercial kits such as the Hains test that is expensive and strips are not reusable. A similar assay can be designed to target mutations for the detection of XDR-TB. Future studies should be conducted in a clinical setting on patients with sensitive strains to increase the specificity.
Investigation into the diversity of antifungal aerobic endospore-forming bacteria associated with bulk and crop rhizosphere soil.
(2011) Musoke, Jolly.; Hunter, Charles Haig.
Members of the genus Bacillus are mainly Gram positive, aerobic rod shaped, endospore-forming bacteria that are increasingly being recognised for their ability to promote plant growth and antagonise fungal pathogens. From a biological control perspective, Bacillus spp. strains that produce antifungal compounds are of particular interest. In this study, aerobic endospore-formers were isolated from an undisturbed indigenous grassland soil and screened for antifungal activity and other plant growth promoting traits. Endospore-formers were also isolated from rhizosphere soil associated with the roots of maize, wheat and kale grown in pots containing soil from the same grassland site. Microbial diversity amongst isolates showing antifungal activity was investigated using different molecular fingerprinting methods, namely, intergenic transcribed spacer–PCR (ITS-PCR), random amplified polymorphic DNA-PCR (RAPD-PCR) and 16S rRNA gene amplification and sequencing. Characterization of the active antimicrobial compound(s) associated with selected isolates was also attempted. Prior to isolating from bulk and rhizosphere soils, samples were pre-heated to eliminate heat sensitive vegetative cells. Mean endospore counts were; wheat rhizosphere, Log 6.03 c.f.u g-1 soil; maize rhizosphere, Log 5.88 c.f.u g-1 soil; kale rhizosphere Log 5.90 c.f.u g-1 soil; and bulk soil Log 5.67 c.f.u g-1soil. A total of three hundred and eighty-four isolates were screened for antagonism towards Rhizoctonia solani using dual-culture plate bioassays. Thirty four of the isolates (~9%) mostly isolated from the bulk soil inhibited R. solani at varying degrees. Differences in antimicrobial interactions were apparent in in vitro bioassay; supposedly due to different concentrations and/or types of antimicrobial compounds. Biochemical tests for amylase, cellulase, chitinase, and proteinase activity, siderophore production and inorganic phosphate solubilisation were conducted. None of the isolates possessed all of these attributes and only a few showed multiple traits. Ninety-one percent of the isolates exhibited proteinase activity, 76% were able to hydrolyze starch whereas only four displayed cellulase activity. Only four isolates from the bulk-soil were capable of solubilising inorganic phosphate. ITS-PCR and 16S rRNA gene sequence analysis showed high levels of genetic homology amongst isolates and the majority were closely associated with representatives of the B. cereus group. Isolate C76 was the exception, being closely matched with B. subtilis. ITS-PCR banding profile was useful for distinguishing between species but did not distinguish within species. RAPD-PCR distinguished finer levels of genetic diversity between and within sample sets, with primer OPG-11 showing the greatest levels of heterogeneity. DNA extraction methods and the influence of template DNA dilution were investigated to determine their influence on RAPD-PCR analysis reproducibility. Prominent bands were comparable for crude template- and kit-extracted DNA but slight changes in band intensity and in some instances, additional faint bands were observed. At the highest DNA concentrations tested (7 μg/ml), further bands with molecular weights above 2.5 kbp were apparent. Strict standardization of PCR conditions greatly reduced variability of the RAPD-PCR analysis. Isolates from the different sample sets were screened for the presence of genetic markers associated with the biosynthesis of zwittermicin A, an aminopolyol antibiotic produced by some members of the B. cereus group. In an initial screen only one isolate, W96, yielded PCR amplicons consistent with those previously reported in the literature for the zwittermicin A genes. Later a further sixteen isolates grouped with W96 on the basis of the RAPD-PCR fingerprinting profiles, were screened for the presence of these genes. Of these, only six showed PCR amplification products similar to W96. Sequence homology testing against the GenBank database confirmed the presence of the zwittermicin A genes in these isolates. Isolate W96 was selected for further extraction and characterization of its antifungal compound(s). However, after culturing in various broth media cell free supernatants of W96 failed to show antifungal activity in vitro even when the supernatants were concentrated 20-fold. These findings provide a general overview of the diversity of aerobic endospore-forming bacteria present in an undisturbed indigenous grassland soil that exhibited antifungal activity in vitro and the limited influence tested crop rhizospheres have on this diversity. Combined use of ITS-PCR, 16S rRNA sequencing and RAPD-PCR techniques served as a rapid and effective means of grouping isolates for further investigations of their potential use as biocontrol agents and plant growth promoting rhizobacteria.
Quantification of fungal degradation of pinus patula and eucalyptus grandis.
(1992) Singh, Vahunth.; Baecker, Albin A. W.
Previous studies of fungal decay have mainly examined long term effects of wood decay. In contrast, the present work, was designed to quantify fungal degradation of wood during incipient decay. Three facultatively anaerobic, dimorphic fungi were isolated from the rumen of sheep. These fungi were identified as Mucor racemosus, Candida tropicalis and Geotrichum capitatum. Scanning electron microscopy showed that these fungi colonised Pinus patula and Eucalyptus grandis extensively but did not appear to degrade the wood. The obligate anaerobe Neocallimastix frontalis colonised wood very sparsely, whereas the white rot bas id iomycetes Cori 01 us versicolor, and Phanaerochaete chrysosporium, and the brown rotters Coniophora puteana and Lentinus lepideus, colonised wood under both aerobic and anaerobic conditions. The extents of colonisation were greater under aerobic conditions. The work then quantified the effects of the basidiomycetes C. versicolor, P. chrysosporium, C. puteana and L .lepideus, and the non-decay mould, M. racemosus in individual and coculture experiments. Wood colonisation was quantified by Kjeldahl nitrogen determinations converted to biomass assays, and degradation was quantified by weight losses, and Klason lignin determinations. Furthermore, the degraded wood samples were also analysed by HPLC analysis of hydrolysates and their sugar contents were determined to establish whether the glucose of cellulose and xylose + mannose of hemicellulose had been utilised by the respective fungi. The extent and nature of sugar utilisation by monocultures and cocultures in wood were then compared with the biomass and degradation data. statistical analyses of' these comparisons correlated the extents of colonisation, degradation, and the patterns of wood sugars predominantly utilised by each fungus. The results of the corresponding glucose, xylose and 'lignin analyses confirmed the brown rot physiological capacity of C.puteana in both'woods. The white rot fungi behaved as simultaneous rotters and,
Molecular and biochemical characterisation of ethanolic D-xylose fermenting Pichia stipitis, Candida shehatae and their fusants.
(1994) Govinden, Roshini.; Pillay, Basil Joseph.; Pillay, D.
No abstract available.
Analysis of microbial populations associated with a sorghum-based fermented product used as an infant weaning cereal.
(1999) Kunene, Nokuthula F.; Hastings, John W.; Von Holy, Alexander.
The incidences of diarrhoeal episodes in infants and children have mostly been associated with the consumption of contaminated weaning foods. This is especially true in developing countries where factors such as the lack of sanitation systems and electricity have been found to contribute to an increase in the incidence of microbiologically contaminated weaning foods. The process of fermentation has been found to reduce the amount of microbiological contamination in such foods as a result of the production of antimicrobial compounds such as organic acids, peroxides, carbon dioxide and bacteriocins. In this study, microbiological surveys were conducted on sorghum powder samples and their corresponding fermented and cooked fermented porridge samples collected from an informal settlement of the Gauteng Province of South Africa. The process of fermentation was found to result in significant decreases (P>0.05) in Gram-negative counts and spore counts, while aerobic plate counts decreased slightly. Lactic acid bacteria counts, however, increased significantly (P>0.05). The cooking process was found to result in further significant decreases (P>0.05) in all counts. Sorghum powder samples and fermented porridge samples were found to be contaminated with potential foodborne pathogens, including Bacillus cereus, Clostridium perfringens and Escherichia coli, however, none of the pathogens tested for were detected in any of the cooked fermented porridge samples. SDS-PAGE and phenotypic analysis of 180 lactic acid bacteria isolated from sorghum powder samples and their corresponding fermented and cooked fermented porridge samples showed that a majority of the isolates were lactobacilli and leuconostocs, however, some isolates were identified as pediococci and lactococci. These results demonstrated the heterogeneity of the lactic acid bacteria isolates that were associated with fermentation processes in this study. Of the lactic acid bacteria identified, Lactobacillus plantarum and Leuconostoc mesenteroides strains were found to have the highest distribution frequencies, being distributed in 87% and 73% of the households, respectively. Analysis of Lactobacillus plantarum (58) and Leuconostoc mesenteroides (46) strains isolated from sorghum powder samples and corresponding fermented and cooked fermented porridge samples by AFLP fingerprinting showed that they originated from a common source, which was sorghum powder. There was, however, evidence of strains that may have been introduced at household level. Antimicrobial activity of selected lactic acid bacteria was found to be mainly due to a decrease in pH in fermented and cooked fermented porridge samples. None of the lactic acid bacteria tested seemed to produce bacteriocins.

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