Changes in the aerobic saprophytic microbial flora during biltong production with special reference to the micrococcaceae
- Authors: Taylor, M B
- Date: 1976
- Subjects: Micrococcaceae , Saprophytism
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4120 , http://hdl.handle.net/10962/d1013308
- Description: Ninety-four presumptive Micrococcus and Staphylococcus strains isolated from both commercial beef biltong and game biltong, were identified using a scheme based on the system used by Baird-Parker. The changes occurring in both the aerobic, saprophytic microbial flora and the environmental factors, during conversion of beef to biltong, were examined. The predominantly Gram-negative, halo-sensitive flora initially present on the meat, was replaced by Gram-positive, halo-tolerant staphylococci and micrococci, which form the dominant component of the microflora of the final product. This replacement was attributed to changing environmental factors, principally to the increasing sodium chloride concentration and associated decline in water activity. The presence of the antifungal antibiotic, pimaricin, during processing did not influence the bacterial flora of the product. However, the addition of potassium sorbate altered the microbial profile of the product significantly. The presence of these two preservatives, at the concentrations used, could not be detected organoleptically. The importance of the saprophytic microflora of the product ln relation to the environmental factors during processing, is also discussed.
- Full Text:
- Date Issued: 1976
- Authors: Taylor, M B
- Date: 1976
- Subjects: Micrococcaceae , Saprophytism
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4120 , http://hdl.handle.net/10962/d1013308
- Description: Ninety-four presumptive Micrococcus and Staphylococcus strains isolated from both commercial beef biltong and game biltong, were identified using a scheme based on the system used by Baird-Parker. The changes occurring in both the aerobic, saprophytic microbial flora and the environmental factors, during conversion of beef to biltong, were examined. The predominantly Gram-negative, halo-sensitive flora initially present on the meat, was replaced by Gram-positive, halo-tolerant staphylococci and micrococci, which form the dominant component of the microflora of the final product. This replacement was attributed to changing environmental factors, principally to the increasing sodium chloride concentration and associated decline in water activity. The presence of the antifungal antibiotic, pimaricin, during processing did not influence the bacterial flora of the product. However, the addition of potassium sorbate altered the microbial profile of the product significantly. The presence of these two preservatives, at the concentrations used, could not be detected organoleptically. The importance of the saprophytic microflora of the product ln relation to the environmental factors during processing, is also discussed.
- Full Text:
- Date Issued: 1976
Studies on the completely mixed activated sludge treatment of fellmongery and tannery lime-sulphide effluents
- Authors: Rawlings, Douglas Eric
- Date: 1977
- Subjects: Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purificiation -- Biological treatment , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4112 , http://hdl.handle.net/10962/d1013052
- Description: Industries producing highly polluted waste waters are having to purify their effluents to meet with ever increasing requirements laid down by water authorities. The South African Water Act of 1956 has prescribed a very high standard to which waste waters must conform before discharge into a South African water course. Enforcement of these standards falls under the jurisdiction of government authorities such as the Department of Water Affairs. Similarly, municipalities and other local authorities set standards with which trade effluents must comply before discharge into public sewers for treatment in a municipal sewage works. These local authorities are empowered to recover from the trader the additional costs incurred in treating trade effluents. Costs are usually levied in respect of volume, oxygen demand, settleable solids and the production of secondary sludge. In recent years, these standards have been enforced to an extent where the survival of several industries has become dependant on whether these industries are able to purify or dispose of their effluents in a manner acceptable to the water authorities. Chap. 1, p. 1.
- Full Text:
- Date Issued: 1977
- Authors: Rawlings, Douglas Eric
- Date: 1977
- Subjects: Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purificiation -- Biological treatment , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4112 , http://hdl.handle.net/10962/d1013052
- Description: Industries producing highly polluted waste waters are having to purify their effluents to meet with ever increasing requirements laid down by water authorities. The South African Water Act of 1956 has prescribed a very high standard to which waste waters must conform before discharge into a South African water course. Enforcement of these standards falls under the jurisdiction of government authorities such as the Department of Water Affairs. Similarly, municipalities and other local authorities set standards with which trade effluents must comply before discharge into public sewers for treatment in a municipal sewage works. These local authorities are empowered to recover from the trader the additional costs incurred in treating trade effluents. Costs are usually levied in respect of volume, oxygen demand, settleable solids and the production of secondary sludge. In recent years, these standards have been enforced to an extent where the survival of several industries has become dependant on whether these industries are able to purify or dispose of their effluents in a manner acceptable to the water authorities. Chap. 1, p. 1.
- Full Text:
- Date Issued: 1977
Hybridization studies within the genus Kluyveromyces van der Walt emend. van der Walt
- Authors: Johannsen, Elz̀bieta
- Date: 1979
- Subjects: Yeast fungi -- Biotechnology , Yeast fungi -- Genetics , Yeast fungi -- Hybridization
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4123 , http://hdl.handle.net/10962/d1013400
- Description: Hybridization studies based on the prototrophic selection technique, involving the use of auxotrophic mutants of strains of all accepted species of the genus Kluyveromyces, are reported. Two main groups of mutually interfertile taxa were established within the genus. The first group comprises Kluyveromyces bulgaricus, Kluyveromyces cicerisporus, Kluyveromyces dobzhanskii, Kluyveromyces drosophilarum, Kluyveromyces fragilis, Kluyveromyces lactis, Kluyveromyces marxianus, Kluyveromyces phaseolosporus, Kluyveromyces vanudenii and Kluyveromyces wikenii. The second group consists of Kluyveromyces dabzhanskii, Kluyveromyces drosophilarum, Kluyveromyces laotis, Kluyveromyces vanudenii and Kluyveromyces wiokerhamii. Hybrids were also detected in crosses involving Kluyveromyces drosophilarum and Kluyveromyces waltii as well as Kluyveromyces marxianus and Kluyveromyces thermotolerans. In terms of the concept of the biological species and in compliance with the requirements of the International Code of Botanical Nomenclature, taxa which hybridize with Kluyveromyces marxianus and form fertile recombinants at frequencies observed in intraspecific crosses, are accepted as varieties of Kluyveromyces marxianus. Hybridization was observed between Kluyveromyces marxianus var. lactis and the presumed imperfect forms of some Kluyveromyces species, namely Candida kefyr, Candida macedoniensis and Torulopsis sphaerica. Recombination was not detected in crosses involving Kluyveromyces marxianus var. marxianus and representatives of other yeast genera, i.e. Pichia, Saccharomyces, Torulaspora and Zygosaccharomyces. Conclusions regarding the relationship between members of the genus Kluyveromyces, reached on the basis of this investigation are compared with those reported by other workers, who based their investigations on phenotypic characteristics as well as on the determinations of mol % G+C and DNA-DNA homology studies.
- Full Text:
- Date Issued: 1979
- Authors: Johannsen, Elz̀bieta
- Date: 1979
- Subjects: Yeast fungi -- Biotechnology , Yeast fungi -- Genetics , Yeast fungi -- Hybridization
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4123 , http://hdl.handle.net/10962/d1013400
- Description: Hybridization studies based on the prototrophic selection technique, involving the use of auxotrophic mutants of strains of all accepted species of the genus Kluyveromyces, are reported. Two main groups of mutually interfertile taxa were established within the genus. The first group comprises Kluyveromyces bulgaricus, Kluyveromyces cicerisporus, Kluyveromyces dobzhanskii, Kluyveromyces drosophilarum, Kluyveromyces fragilis, Kluyveromyces lactis, Kluyveromyces marxianus, Kluyveromyces phaseolosporus, Kluyveromyces vanudenii and Kluyveromyces wikenii. The second group consists of Kluyveromyces dabzhanskii, Kluyveromyces drosophilarum, Kluyveromyces laotis, Kluyveromyces vanudenii and Kluyveromyces wiokerhamii. Hybrids were also detected in crosses involving Kluyveromyces drosophilarum and Kluyveromyces waltii as well as Kluyveromyces marxianus and Kluyveromyces thermotolerans. In terms of the concept of the biological species and in compliance with the requirements of the International Code of Botanical Nomenclature, taxa which hybridize with Kluyveromyces marxianus and form fertile recombinants at frequencies observed in intraspecific crosses, are accepted as varieties of Kluyveromyces marxianus. Hybridization was observed between Kluyveromyces marxianus var. lactis and the presumed imperfect forms of some Kluyveromyces species, namely Candida kefyr, Candida macedoniensis and Torulopsis sphaerica. Recombination was not detected in crosses involving Kluyveromyces marxianus var. marxianus and representatives of other yeast genera, i.e. Pichia, Saccharomyces, Torulaspora and Zygosaccharomyces. Conclusions regarding the relationship between members of the genus Kluyveromyces, reached on the basis of this investigation are compared with those reported by other workers, who based their investigations on phenotypic characteristics as well as on the determinations of mol % G+C and DNA-DNA homology studies.
- Full Text:
- Date Issued: 1979
Bacteriophage growth on stationary phase achromabacter strains
- Authors: Robb, Susan Mary
- Date: 1980
- Subjects: Bacteriophages , Strains and stresses
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4125 , http://hdl.handle.net/10962/d1014131
- Description: Achromobacter w.t. and strain 14 both support phage α3a growth in stationary phase, but unlike the w.t. strain, exponential phase cultures of strain 14 block phage development. A standard method was developed for determining phage growth in stationary phase cultures. Lyophilised cells were used to eliminate variations due to the unstable phenotype of Achromobacter strain 14 cells. Phage α3a growth in stationary phase was characterized by a long and variable latent period of 6 to 9 h and an increased burst size of 709 p.f.u. per cell as compared with 153 p.f.u. per cell in exponential wild type cells. During the latent period the infected cells were very sensitive to changes in growth conditions and in particular, dilution. Pre-conditioning of the bacterial cells by allowing them to stand for 24 h after shaking for 3 days was an important aspect of the stationary phase phage growth system. Cells which had been allowed to stand retained the ability to be infected and to support phage growth for at least 16 days. Shaking cultures gradually lost the ability to support phage growth but the phage could persist in the host cell for 10 days until removal from shaking when the lytic cycle could proceed after allowing the cultures to stand. In comparison the latent period and burst size in Achromobacter w.t. stationary phase cells were reduced to less than 2 h and less than 200 respectively. Stationary phase cultures differed physiologically and morphologically depending on the aeration conditions. In comparison with non-aerated standing cultures, vigorously aerated cultures showed a decrease in viability, RNA synthesis, membrane transport, intracellular ATP levels, UV resistance and heat resistance but had markedly higher protein synthesis levels. Aerated cells were small non-motile rods which did not support phage growth. They developed into large motile rods under conditions of limited aeration and were able to propagate phage. It was proposed that changes in the host control mechanisms for macromolecular synthesis may be instrumental in either blocking or permitting phage development. A spontaneous mutant of Achromobacter strain 14 (14x) which liberated phage and was resistant to superinfection was isolated. The phage-host relationship was unstable and similar to the phage carrier state. The liberated phage were able to grow in exponential strain 14 cells. It was proposed that strain 14 was a defective lysogen and that an immunity phase shift model may account for the differential phage growth in exponential and stationary phase cells. Host transcriptional control appears to be implicated in control of phage development in exponential and stationary phase cells. Achromobacter Lp only supported phage in exponential phase but a rifampicin resistant mutant of this strain was able to propagate phage in stationary phase. In vitro RNA synthesis assays showed that the rifampicin resistance was caused by an alteration in the RNA polymerase. Preliminary experiments to determine intracellular phage macromolecular synthesis were carried out using exponential Achromobacter w.t. cells which had been irradiated with UV prior to infection. In irradiated cells, infection with phage resulted in stimulation of DNA synthesis but no stimulation of protein synthesis. Phage production was drastically reduced in cells which had been treated with very low UV doses. It was proposed that α3a development may rely heavily on host cell functions which are destroyed by UV. Achromobacter mutants with defective leucine transport systems were isolated. Mutants which lost the leucine uptake system completely were totally resistant to phage infection and were unable to adsorb phage α3a. This is the first report to implicate an amino-acid transport system in phage adsorption.
- Full Text:
- Date Issued: 1980
- Authors: Robb, Susan Mary
- Date: 1980
- Subjects: Bacteriophages , Strains and stresses
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4125 , http://hdl.handle.net/10962/d1014131
- Description: Achromobacter w.t. and strain 14 both support phage α3a growth in stationary phase, but unlike the w.t. strain, exponential phase cultures of strain 14 block phage development. A standard method was developed for determining phage growth in stationary phase cultures. Lyophilised cells were used to eliminate variations due to the unstable phenotype of Achromobacter strain 14 cells. Phage α3a growth in stationary phase was characterized by a long and variable latent period of 6 to 9 h and an increased burst size of 709 p.f.u. per cell as compared with 153 p.f.u. per cell in exponential wild type cells. During the latent period the infected cells were very sensitive to changes in growth conditions and in particular, dilution. Pre-conditioning of the bacterial cells by allowing them to stand for 24 h after shaking for 3 days was an important aspect of the stationary phase phage growth system. Cells which had been allowed to stand retained the ability to be infected and to support phage growth for at least 16 days. Shaking cultures gradually lost the ability to support phage growth but the phage could persist in the host cell for 10 days until removal from shaking when the lytic cycle could proceed after allowing the cultures to stand. In comparison the latent period and burst size in Achromobacter w.t. stationary phase cells were reduced to less than 2 h and less than 200 respectively. Stationary phase cultures differed physiologically and morphologically depending on the aeration conditions. In comparison with non-aerated standing cultures, vigorously aerated cultures showed a decrease in viability, RNA synthesis, membrane transport, intracellular ATP levels, UV resistance and heat resistance but had markedly higher protein synthesis levels. Aerated cells were small non-motile rods which did not support phage growth. They developed into large motile rods under conditions of limited aeration and were able to propagate phage. It was proposed that changes in the host control mechanisms for macromolecular synthesis may be instrumental in either blocking or permitting phage development. A spontaneous mutant of Achromobacter strain 14 (14x) which liberated phage and was resistant to superinfection was isolated. The phage-host relationship was unstable and similar to the phage carrier state. The liberated phage were able to grow in exponential strain 14 cells. It was proposed that strain 14 was a defective lysogen and that an immunity phase shift model may account for the differential phage growth in exponential and stationary phase cells. Host transcriptional control appears to be implicated in control of phage development in exponential and stationary phase cells. Achromobacter Lp only supported phage in exponential phase but a rifampicin resistant mutant of this strain was able to propagate phage in stationary phase. In vitro RNA synthesis assays showed that the rifampicin resistance was caused by an alteration in the RNA polymerase. Preliminary experiments to determine intracellular phage macromolecular synthesis were carried out using exponential Achromobacter w.t. cells which had been irradiated with UV prior to infection. In irradiated cells, infection with phage resulted in stimulation of DNA synthesis but no stimulation of protein synthesis. Phage production was drastically reduced in cells which had been treated with very low UV doses. It was proposed that α3a development may rely heavily on host cell functions which are destroyed by UV. Achromobacter mutants with defective leucine transport systems were isolated. Mutants which lost the leucine uptake system completely were totally resistant to phage infection and were unable to adsorb phage α3a. This is the first report to implicate an amino-acid transport system in phage adsorption.
- Full Text:
- Date Issued: 1980
Genetic studies and physiological responses to ultraviolet radiation in the Bacteroides fragilis group
- Authors: Jones, David Todman
- Date: 1980
- Subjects: Bacteroides Ultraviolet radiation
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4072 , http://hdl.handle.net/10962/d1007047
- Description: The contents of this thesis have been divided into 2 parts . The first part deals with genetic studies carried out on 36 strains belonging to the Bacteroides fragilis group. A number of mutants were isolated from several of the strains. A notable feature of the methods used was the low yield of mutants obtained and the marked sensitivity of these organisms to the mutagenic agents. Variations in colonial morphology was found to be a common feature amongst these organisms. In a few strains this phenomenon was clearly visible, in the remainder it was much weaker, and often could only be seen with the aid of a microscope . Colonial variation was found to be due to the ability of a proporti on of the cells to pruduce capsules or slime layers. The variants were found to segregate at high frequency and different growth conditions were found to have little effect on the segregation frequency or capsule formation . A number of phages specific for B. fragilis and B. t hetaiotaomicron were isol ated. All these phages were virulent and attempts to induce lysogenic phages were unsuccesful . The use of these phages in attempts to obtain transduction proved unsuccessful. A phage carrier state was found to occur in the majority of the phage-host cell systems, which seemed to be due to the presence of phage-resistant encapsulated cells in the population. Bacteriocins were produced by about half the strains, these inhibited the growth of a high proportion of the 36 strains tested. The bacteriocins were released into the growth media at the end of the growth period in the 2 bacteriocins tested. A link between the mode of action of one bacteriocin and rifampicin-resistance was investigated. All the bacter iocins tested were found to be inactive against some rifampicin-resistant mutants of a susceptible strain, suggesting a common mode of action. The presence of capsules in some cells appeared to confer bacteriocin-resistance on these variants. The second part of the thesis deals with a study of the physiological responses of a single strain of B.fragilis to ultraviolet radiation. This strain was found to be more sensitive to ultraviolet radiation under aerobic conditions. The amount of pyrimidine dimers formed after irradiation under anaerobic and aerobic conditions, was not found to differ significantly, indicating that the increase in sensitivity under aerobic conditions was not due to an increase in DNA damage. The use of repair inhibitors and the survival characteristics indicate that this difference was due to decreased repair capabilities under aerobic conditions. Liquid holding recovery in B.fragiZis was found to occur under aerobic conditions . This process was brought about by excision repair and appeared to be due to a decrease in repair efficiency under aerobic conditions. Under anaerobic conditions, where full repair capabilities were present, liquid holding recovery was inhibited. Both minimal medium recovery and fluence dependent filament formation were found to occur in irradiated B.fragiZis cells. The survival kinetics of a number of irradiated B.fragiZis phages were determined and a number of phage reactivation processes were investigated. Little or no host cell reactivation appeared to occur in the strains investigated, however, some ultraviolet reactivation and multiplicity reactivation was found to occur, but only under anaerobic conditions. Photoreactivation was found to be absent in this organism, but an excision repair system was present . The excision repair system was partially characterized and was found to resemble short patch excision repair in E.coli. Evidence was found which suggested that a second mode of repair which was sensitive to oxygen, also occurred in this strain. This repair system which appeared to be responsible for error-prone repair, and the systems which were responsible for ultraviolet reactivation and multiplicity reactivation, seemed to be dependent on a recombination function' which was inhibited by oxygen. The significance of this finding for future genetic studies was discussed.
- Full Text:
- Date Issued: 1980
- Authors: Jones, David Todman
- Date: 1980
- Subjects: Bacteroides Ultraviolet radiation
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4072 , http://hdl.handle.net/10962/d1007047
- Description: The contents of this thesis have been divided into 2 parts . The first part deals with genetic studies carried out on 36 strains belonging to the Bacteroides fragilis group. A number of mutants were isolated from several of the strains. A notable feature of the methods used was the low yield of mutants obtained and the marked sensitivity of these organisms to the mutagenic agents. Variations in colonial morphology was found to be a common feature amongst these organisms. In a few strains this phenomenon was clearly visible, in the remainder it was much weaker, and often could only be seen with the aid of a microscope . Colonial variation was found to be due to the ability of a proporti on of the cells to pruduce capsules or slime layers. The variants were found to segregate at high frequency and different growth conditions were found to have little effect on the segregation frequency or capsule formation . A number of phages specific for B. fragilis and B. t hetaiotaomicron were isol ated. All these phages were virulent and attempts to induce lysogenic phages were unsuccesful . The use of these phages in attempts to obtain transduction proved unsuccessful. A phage carrier state was found to occur in the majority of the phage-host cell systems, which seemed to be due to the presence of phage-resistant encapsulated cells in the population. Bacteriocins were produced by about half the strains, these inhibited the growth of a high proportion of the 36 strains tested. The bacteriocins were released into the growth media at the end of the growth period in the 2 bacteriocins tested. A link between the mode of action of one bacteriocin and rifampicin-resistance was investigated. All the bacter iocins tested were found to be inactive against some rifampicin-resistant mutants of a susceptible strain, suggesting a common mode of action. The presence of capsules in some cells appeared to confer bacteriocin-resistance on these variants. The second part of the thesis deals with a study of the physiological responses of a single strain of B.fragilis to ultraviolet radiation. This strain was found to be more sensitive to ultraviolet radiation under aerobic conditions. The amount of pyrimidine dimers formed after irradiation under anaerobic and aerobic conditions, was not found to differ significantly, indicating that the increase in sensitivity under aerobic conditions was not due to an increase in DNA damage. The use of repair inhibitors and the survival characteristics indicate that this difference was due to decreased repair capabilities under aerobic conditions. Liquid holding recovery in B.fragiZis was found to occur under aerobic conditions . This process was brought about by excision repair and appeared to be due to a decrease in repair efficiency under aerobic conditions. Under anaerobic conditions, where full repair capabilities were present, liquid holding recovery was inhibited. Both minimal medium recovery and fluence dependent filament formation were found to occur in irradiated B.fragiZis cells. The survival kinetics of a number of irradiated B.fragiZis phages were determined and a number of phage reactivation processes were investigated. Little or no host cell reactivation appeared to occur in the strains investigated, however, some ultraviolet reactivation and multiplicity reactivation was found to occur, but only under anaerobic conditions. Photoreactivation was found to be absent in this organism, but an excision repair system was present . The excision repair system was partially characterized and was found to resemble short patch excision repair in E.coli. Evidence was found which suggested that a second mode of repair which was sensitive to oxygen, also occurred in this strain. This repair system which appeared to be responsible for error-prone repair, and the systems which were responsible for ultraviolet reactivation and multiplicity reactivation, seemed to be dependent on a recombination function' which was inhibited by oxygen. The significance of this finding for future genetic studies was discussed.
- Full Text:
- Date Issued: 1980
Studies on an autolysin produced by clostridium acetobutylicum
- Authors: Webster, Jocelyn Rowena
- Date: 1981
- Subjects: Clostridium acetobutylicum , Autolysis , Bacteriocins , Proteins -- Synthesis , DNA -- Synthesis , RNA -- Synthesis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3893 , http://hdl.handle.net/10962/d1003724
- Description: An extracellular bacteriocin-like substance produced by Clostridium acetobutylicum was detected during studies on an industrial fermentation process. The bacteriocin-like substance was not inducible by either ultraviolet light or mitomycin C, and its production was not associated with the induction of a protease. Studies on the mode of action of the bacteriocin-like substance indicated that it had no significant effect on DNA, RNA, or protein synthesis, and it did not cause the loss of intracellular ATP. However, the bacteriocin-like substance was able to lyse SDS-treated cells and cell walls of C. acetobutylicum and was identified as an autolysin. Some of the characteristics of this extracellular autolysin were determined, and after purification it was shown to be a glycoprotein with a molecular weight of 28 000.
- Full Text:
- Date Issued: 1981
- Authors: Webster, Jocelyn Rowena
- Date: 1981
- Subjects: Clostridium acetobutylicum , Autolysis , Bacteriocins , Proteins -- Synthesis , DNA -- Synthesis , RNA -- Synthesis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3893 , http://hdl.handle.net/10962/d1003724
- Description: An extracellular bacteriocin-like substance produced by Clostridium acetobutylicum was detected during studies on an industrial fermentation process. The bacteriocin-like substance was not inducible by either ultraviolet light or mitomycin C, and its production was not associated with the induction of a protease. Studies on the mode of action of the bacteriocin-like substance indicated that it had no significant effect on DNA, RNA, or protein synthesis, and it did not cause the loss of intracellular ATP. However, the bacteriocin-like substance was able to lyse SDS-treated cells and cell walls of C. acetobutylicum and was identified as an autolysin. Some of the characteristics of this extracellular autolysin were determined, and after purification it was shown to be a glycoprotein with a molecular weight of 28 000.
- Full Text:
- Date Issued: 1981
Investigation of the causative agents of the 1982 Gazankulu poliomyelitis outbreak, using four biochemical techniques
- Authors: Gibson, Katherine Margaret
- Date: 1989
- Subjects: Poliomyelitis -- Analysis , Poliomyelitis -- History -- South Africa , Poliomyelitis vaccine -- Analysis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3878 , http://hdl.handle.net/10962/d1001612
- Description: Comparison of poliovirus strains was carried out to determine the origin of the virus in two isolates obtained during the 1982 outbreak of poliomyelitis in Gazankulu. Comparisons of the outbreak isolates with vaccine and wild-type strains of the same poliovirus type were carried out using four biochemical techniques. SDS-polyacrylamide gel electrophoresis (SDS-PAGE), two-dimensional thin-layer chromatography (TLC) and reversed-phase high-performance liquid-chromatography (RP-HPLC) were used for comparing viral capsid proteins. Comparison of poliovirus strains at a genetic level was carried out using two-dimensional oligonucleotide mapping of viral RNA. Results showed the type 1 poliovirus isolate, 5061, to be a novel wild-type poliovirus. The type 2 isolate, 5068, was closely related to the poliovirus type 2 Sabin vaccine strain, P712. It was concluded that the intrinsic variability of poliovirus strains was responsible for the appearance of isolate 5068
- Full Text:
- Date Issued: 1989
- Authors: Gibson, Katherine Margaret
- Date: 1989
- Subjects: Poliomyelitis -- Analysis , Poliomyelitis -- History -- South Africa , Poliomyelitis vaccine -- Analysis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3878 , http://hdl.handle.net/10962/d1001612
- Description: Comparison of poliovirus strains was carried out to determine the origin of the virus in two isolates obtained during the 1982 outbreak of poliomyelitis in Gazankulu. Comparisons of the outbreak isolates with vaccine and wild-type strains of the same poliovirus type were carried out using four biochemical techniques. SDS-polyacrylamide gel electrophoresis (SDS-PAGE), two-dimensional thin-layer chromatography (TLC) and reversed-phase high-performance liquid-chromatography (RP-HPLC) were used for comparing viral capsid proteins. Comparison of poliovirus strains at a genetic level was carried out using two-dimensional oligonucleotide mapping of viral RNA. Results showed the type 1 poliovirus isolate, 5061, to be a novel wild-type poliovirus. The type 2 isolate, 5068, was closely related to the poliovirus type 2 Sabin vaccine strain, P712. It was concluded that the intrinsic variability of poliovirus strains was responsible for the appearance of isolate 5068
- Full Text:
- Date Issued: 1989
An investigation into the use of anaerobic digestion for the treatment of tannery wastewaters
- Authors: Jackson-Moss, Clive Alan
- Date: 1991
- Subjects: Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purification -- Biological treatment -- South Africa , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4059 , http://hdl.handle.net/10962/d1004120 , Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purification -- Biological treatment -- South Africa , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Description: The anaerobic digestion of tannery wastewaters was investigated with a view to using this form of treatment in the tanning industry. As these wastewaters are extremely complex and contain high concentrations of both inorganic and organic compounds, the effect of these individual compounds on the anaerobic digestion process was investigated in detail, in order to ascertain the fate of these compounds during the digestion process. The experiments comprising the initial toxicity study were carried out as adaptation experiments using a synthetic wastewater. It was found that the heavy metals such as chrome, aluminium and iron precipitated and accumulated in the sludge bed of the digesters . The soluble ions such as sodium and chloride were not retained and passed through the digesters. Approximately 20 % of the calcium ions were removed through precipitation, with the remainder being present in the digester effluent . Under the anaerobic conditions, ammonification of the organic nitrogen occurred, and influent sulphates were reduced to sulphides . These sulphides were present as either H2S, HS or insoluble sulphides. As these compounds under investigation on caused no inhibition of the anaerobic digestion process at the concentrations found in tannery wastewaters, the anaerobic treatment of these wastewaters appeared to be possible, provided the bacteria were given sufficient time to adapt to the potentially toxic compounds. However, despite the findings of the synthetic study, the successful anaerobic digestion of the tannery effluents could not be achieved. Although the use of acid was found to be essential in order to control the digester pH in the optimum range, the metabolism of the methanogenic bacteria was inhibited by the presence or absence of unknown compounds. Neither the addition of essential trace nutrients, nor the prevention of the competition between the methanogens and the sulphate-reducing bacteria were able to reverse this inhibition. As tannery effluents contain very low concentrations of phosphorous, it is possible that the methanogens were inhibited by a lack of phosphorous, which is essential during methanogenesis. In contrast to the results obtained from the effluent experiments, the anaerobic digestion of tannery sludge was found to be possible. Of the organic solids present in the sludge, 60 % were degraded and converted into biogas, which had a methane content greater than 70 %. The degradation of the organic solids ensured that COD and PV reductions of greater than 90 % were achieved, and the fate of the compounds in the digesters were in agreement with the findings of the v synthetic study. Efforts to improve the efficiency of the digestion process through the addition of trace nutrients and the use of a two-stage process were only successful in bringing about a minor improvement in digester performance. The overall results of this investigation show, therefore, that although the anaerobic treatment of the tannery effluent was not achieved, the successful anaerobic digestion of tannery sludge is possible at low loading rates. As many difficulties still need to be solved, a great deal of further research is necessary if anaerobic digestion is to be used on an industrial scale for the treatment and disposal of tannery wastewaters.
- Full Text:
- Date Issued: 1991
- Authors: Jackson-Moss, Clive Alan
- Date: 1991
- Subjects: Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purification -- Biological treatment -- South Africa , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4059 , http://hdl.handle.net/10962/d1004120 , Tanneries -- Waste disposal , Sewage sludge -- South Africa -- Management , Sewage -- Purification -- Biological treatment -- South Africa , Water quality management -- South Africa , Water -- Purification -- Biological treatment -- South Africa
- Description: The anaerobic digestion of tannery wastewaters was investigated with a view to using this form of treatment in the tanning industry. As these wastewaters are extremely complex and contain high concentrations of both inorganic and organic compounds, the effect of these individual compounds on the anaerobic digestion process was investigated in detail, in order to ascertain the fate of these compounds during the digestion process. The experiments comprising the initial toxicity study were carried out as adaptation experiments using a synthetic wastewater. It was found that the heavy metals such as chrome, aluminium and iron precipitated and accumulated in the sludge bed of the digesters . The soluble ions such as sodium and chloride were not retained and passed through the digesters. Approximately 20 % of the calcium ions were removed through precipitation, with the remainder being present in the digester effluent . Under the anaerobic conditions, ammonification of the organic nitrogen occurred, and influent sulphates were reduced to sulphides . These sulphides were present as either H2S, HS or insoluble sulphides. As these compounds under investigation on caused no inhibition of the anaerobic digestion process at the concentrations found in tannery wastewaters, the anaerobic treatment of these wastewaters appeared to be possible, provided the bacteria were given sufficient time to adapt to the potentially toxic compounds. However, despite the findings of the synthetic study, the successful anaerobic digestion of the tannery effluents could not be achieved. Although the use of acid was found to be essential in order to control the digester pH in the optimum range, the metabolism of the methanogenic bacteria was inhibited by the presence or absence of unknown compounds. Neither the addition of essential trace nutrients, nor the prevention of the competition between the methanogens and the sulphate-reducing bacteria were able to reverse this inhibition. As tannery effluents contain very low concentrations of phosphorous, it is possible that the methanogens were inhibited by a lack of phosphorous, which is essential during methanogenesis. In contrast to the results obtained from the effluent experiments, the anaerobic digestion of tannery sludge was found to be possible. Of the organic solids present in the sludge, 60 % were degraded and converted into biogas, which had a methane content greater than 70 %. The degradation of the organic solids ensured that COD and PV reductions of greater than 90 % were achieved, and the fate of the compounds in the digesters were in agreement with the findings of the v synthetic study. Efforts to improve the efficiency of the digestion process through the addition of trace nutrients and the use of a two-stage process were only successful in bringing about a minor improvement in digester performance. The overall results of this investigation show, therefore, that although the anaerobic treatment of the tannery effluent was not achieved, the successful anaerobic digestion of tannery sludge is possible at low loading rates. As many difficulties still need to be solved, a great deal of further research is necessary if anaerobic digestion is to be used on an industrial scale for the treatment and disposal of tannery wastewaters.
- Full Text:
- Date Issued: 1991
The pineal gland as a model to elucidate the primary mode of action of sympathoactive agents
- Authors: Welman, Alan David
- Date: 1991
- Subjects: Pineal gland , Cythochemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3876 , http://hdl.handle.net/10962/d1001610
- Description: An attempt was made to use the pineal gland as a model for the study of the primary mode of action of sympathoactive agents. Two drugs were investigated, viz. alpha-methyldopa and ephedrine whose mode of action is not entirely clear. Organ cultures of pineal glands from rats treated chronically with alpha-methyldopa showed enhanced conversion of radioactive serotonin to melatonin (aMT) , as well as its precursor Nacetylserotonin (aHT). This treatment was also found to raise Nacetyltransferase (NAT) activity. These increases associated with alpha-methyldopa treatment were further enhanced by the beta-adrenergic agonist, isoproterenol, suggesting a supersensitivity-type effect occurring at the level of the beta-receptor. A subsequent binding study, however, showed a decrease in beta-receptor binding with exposure to alpha-methyldopa, providing mitigating evidence against the occurrence of a supersensitivity phenomenon. It is possible that a metabolite of alpha-methyldopa acts as an alpha 1 and beta-adrenergic agonist, resulting in greater melatonin (aMT) and N-acetylserotonin (aHT) synthesis than by a beta-adrenergic agonist, isoproterenol. Combined treatment of pineals with alpha-methyldopa and an alphareceptor blocker, phentolamine, resulted in melatonin (aMT) , Nacetylserotonin (aHT) , and N-acetyltransferase (NAT) activity levels which were lower than those obtained with alpha-methyldopa treatment alone, thus confirming the alpha-adrenergic activity of the metabolite of alpha-methyldopa. Additional pineal metabolites were isolated and measured simultaneously in the organ culture experiments. Organ cultures of rat pineal glands treated with ephedrine showed raised levels of melatonin (aMT) and N-acetylserotonin (aHT). Treatment with ephedrine also produced raised N-acetyltransferase activity. A further enhancement of these parameters was induced by norepinephrine, suggesting a supersensitivity-type effect occurring at the level of the beta-adrenergic receptor. Rats were treated with reserpine (a norepinephrine depleter) and the pineals exposed to ephedrine. Endogenous norepinephrine normally released by the action of ephedrine was thus absent, and under these conditions, levels of melatonin (aMT) and N-acetylserotonin (aHT) were reduced. N-acetyltransferase (NAT) activity was also reduced, but maintained levels pointing to substantial adrenergic activity of ephedrine as well as norepinephrine released by virtue of the drug's action. A subsequent binding study showed a decrease in beta-adrenergic receptor binding with exposure to ephedrine and a further decrease in ephedrine treated pineals from reserpine treated rats, thus ruling out the occurrence of a supersensitivity phenomenon. It is possible that both ephedrine and released norepinephrine have alpha- and beta-receptor activity. Additional pineal metabolites were isolated and measured in the organ culture experiments. A 16-hour time profile of the production of melatonin (aMT) and N-acetylserotonin (aHT) with norepinephrine and ephedrine treatment provided useful information regarding the course of action of the two agents. A pineal cell-culture system was developed and exposed to ephedrine and norepinephrine. N-acetyltransferase (NAT) activity levels measured after exposure to these agents were raised, confirming the adrenergic activity of both in the model. Finally, an HPLC system coupled to a UV detector was used in an attempt to measure melatonin (aMT) extracted from pineal organ culture media. The results showed that melatonin could be measured by this method, however, a more sensitive detection system was recommended for future work.
- Full Text:
- Date Issued: 1991
- Authors: Welman, Alan David
- Date: 1991
- Subjects: Pineal gland , Cythochemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3876 , http://hdl.handle.net/10962/d1001610
- Description: An attempt was made to use the pineal gland as a model for the study of the primary mode of action of sympathoactive agents. Two drugs were investigated, viz. alpha-methyldopa and ephedrine whose mode of action is not entirely clear. Organ cultures of pineal glands from rats treated chronically with alpha-methyldopa showed enhanced conversion of radioactive serotonin to melatonin (aMT) , as well as its precursor Nacetylserotonin (aHT). This treatment was also found to raise Nacetyltransferase (NAT) activity. These increases associated with alpha-methyldopa treatment were further enhanced by the beta-adrenergic agonist, isoproterenol, suggesting a supersensitivity-type effect occurring at the level of the beta-receptor. A subsequent binding study, however, showed a decrease in beta-receptor binding with exposure to alpha-methyldopa, providing mitigating evidence against the occurrence of a supersensitivity phenomenon. It is possible that a metabolite of alpha-methyldopa acts as an alpha 1 and beta-adrenergic agonist, resulting in greater melatonin (aMT) and N-acetylserotonin (aHT) synthesis than by a beta-adrenergic agonist, isoproterenol. Combined treatment of pineals with alpha-methyldopa and an alphareceptor blocker, phentolamine, resulted in melatonin (aMT) , Nacetylserotonin (aHT) , and N-acetyltransferase (NAT) activity levels which were lower than those obtained with alpha-methyldopa treatment alone, thus confirming the alpha-adrenergic activity of the metabolite of alpha-methyldopa. Additional pineal metabolites were isolated and measured simultaneously in the organ culture experiments. Organ cultures of rat pineal glands treated with ephedrine showed raised levels of melatonin (aMT) and N-acetylserotonin (aHT). Treatment with ephedrine also produced raised N-acetyltransferase activity. A further enhancement of these parameters was induced by norepinephrine, suggesting a supersensitivity-type effect occurring at the level of the beta-adrenergic receptor. Rats were treated with reserpine (a norepinephrine depleter) and the pineals exposed to ephedrine. Endogenous norepinephrine normally released by the action of ephedrine was thus absent, and under these conditions, levels of melatonin (aMT) and N-acetylserotonin (aHT) were reduced. N-acetyltransferase (NAT) activity was also reduced, but maintained levels pointing to substantial adrenergic activity of ephedrine as well as norepinephrine released by virtue of the drug's action. A subsequent binding study showed a decrease in beta-adrenergic receptor binding with exposure to ephedrine and a further decrease in ephedrine treated pineals from reserpine treated rats, thus ruling out the occurrence of a supersensitivity phenomenon. It is possible that both ephedrine and released norepinephrine have alpha- and beta-receptor activity. Additional pineal metabolites were isolated and measured in the organ culture experiments. A 16-hour time profile of the production of melatonin (aMT) and N-acetylserotonin (aHT) with norepinephrine and ephedrine treatment provided useful information regarding the course of action of the two agents. A pineal cell-culture system was developed and exposed to ephedrine and norepinephrine. N-acetyltransferase (NAT) activity levels measured after exposure to these agents were raised, confirming the adrenergic activity of both in the model. Finally, an HPLC system coupled to a UV detector was used in an attempt to measure melatonin (aMT) extracted from pineal organ culture media. The results showed that melatonin could be measured by this method, however, a more sensitive detection system was recommended for future work.
- Full Text:
- Date Issued: 1991
The pineal gland as a model to elucidate the primary mode of action of sympathoactive agents
- Authors: Welman, Alan David
- Date: 1991
- Subjects: Pineal gland , Cythochemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4279 , http://hdl.handle.net/10962/d1002005 , Pineal gland
- Description: An attempt was made to use the pineal gland as a model for the study of the primary mode of action of sympathoactive agents. Two drugs were investigated, viz. alpha-methyldopa and ephedrine whose mode of action is not entirely clear. Organ cultures of pineal glands from rats treated chronically with alpha-methyldopa showed enhanced conversion of radioactive serotonin to melatonin (aMT) , as well as its precursor Nacetylserotonin (aHT). This treatment was also found to raise Nacetyltransferase (NAT) activity. These increases associated with alpha-methyldopa treatment were further enhanced by the beta-adrenergic agonist, isoproterenol, suggesting a supersensitivity-type effect occurring at the level of the beta-receptor. A subsequent binding study, however, showed a decrease in beta-receptor binding with exposure to alpha-methyldopa, providing mitigating evidence against the occurrence of a supersensitivity phenomenon. It is possible that a metabolite of alpha-methyldopa acts as an alpha 1 and beta-adrenergic agonist, resulting in greater melatonin (aMT) and N-acetylserotonin (aHT) synthesis than by a beta-adrenergic agonist, isoproterenol. Combined treatment of pineals with alpha-methyldopa and an alphareceptor blocker, phentolamine, resulted in melatonin (aMT) , Nacetylserotonin (aHT) , and N-acetyltransferase (NAT) activity levels which were lower than those obtained with alpha-methyldopa treatment alone, thus confirming the alpha-adrenergic activity of the metabolite of alpha-methyldopa. Additional pineal metabolites were isolated and measured simultaneously in the organ culture experiments. Organ cultures of rat pineal glands treated with ephedrine showed raised levels of melatonin (aMT) and N-acetylserotonin (aHT). Treatment with ephedrine also produced raised N-acetyltransferase activity. A further enhancement of these parameters was induced by norepinephrine, suggesting a supersensitivity-type effect occurring at the level of the beta-adrenergic receptor. Rats were treated with reserpine (a norepinephrine depleter) and the pineals exposed to ephedrine. Endogenous norepinephrine normally released by the action of ephedrine was thus absent, and under these conditions, levels of melatonin (aMT) and N-acetylserotonin (aHT) were reduced. N-acetyltransferase (NAT) activity was also reduced, but maintained levels pointing to substantial adrenergic activity of ephedrine as well as norepinephrine released by virtue of the drug's action. A subsequent binding study showed a decrease in beta-adrenergic receptor binding with exposure to ephedrine and a further decrease in ephedrine treated pineals from reserpine treated rats, thus ruling out the occurrence of a supersensitivity phenomenon. It is possible that both ephedrine and released norepinephrine have alpha- and beta-receptor activity. Additional pineal metabolites were isolated and measured in the organ culture experiments. A 16-hour time profile of the production of melatonin (aMT) and N-acetylserotonin (aHT) with norepinephrine and ephedrine treatment provided useful information regarding the course of action of the two agents. A pineal cell-culture system was developed and exposed to ephedrine and norepinephrine. N-acetyltransferase (NAT) activity levels measured after exposure to these agents were raised, confirming the adrenergic activity of both in the model. Finally, an HPLC system coupled to a UV detector was used in an attempt to measure melatonin (aMT) extracted from pineal organ culture media. The results showed that melatonin could be measured by this method, however, a more sensitive detection system was recommended for future work
- Full Text:
- Date Issued: 1991
- Authors: Welman, Alan David
- Date: 1991
- Subjects: Pineal gland , Cythochemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4279 , http://hdl.handle.net/10962/d1002005 , Pineal gland
- Description: An attempt was made to use the pineal gland as a model for the study of the primary mode of action of sympathoactive agents. Two drugs were investigated, viz. alpha-methyldopa and ephedrine whose mode of action is not entirely clear. Organ cultures of pineal glands from rats treated chronically with alpha-methyldopa showed enhanced conversion of radioactive serotonin to melatonin (aMT) , as well as its precursor Nacetylserotonin (aHT). This treatment was also found to raise Nacetyltransferase (NAT) activity. These increases associated with alpha-methyldopa treatment were further enhanced by the beta-adrenergic agonist, isoproterenol, suggesting a supersensitivity-type effect occurring at the level of the beta-receptor. A subsequent binding study, however, showed a decrease in beta-receptor binding with exposure to alpha-methyldopa, providing mitigating evidence against the occurrence of a supersensitivity phenomenon. It is possible that a metabolite of alpha-methyldopa acts as an alpha 1 and beta-adrenergic agonist, resulting in greater melatonin (aMT) and N-acetylserotonin (aHT) synthesis than by a beta-adrenergic agonist, isoproterenol. Combined treatment of pineals with alpha-methyldopa and an alphareceptor blocker, phentolamine, resulted in melatonin (aMT) , Nacetylserotonin (aHT) , and N-acetyltransferase (NAT) activity levels which were lower than those obtained with alpha-methyldopa treatment alone, thus confirming the alpha-adrenergic activity of the metabolite of alpha-methyldopa. Additional pineal metabolites were isolated and measured simultaneously in the organ culture experiments. Organ cultures of rat pineal glands treated with ephedrine showed raised levels of melatonin (aMT) and N-acetylserotonin (aHT). Treatment with ephedrine also produced raised N-acetyltransferase activity. A further enhancement of these parameters was induced by norepinephrine, suggesting a supersensitivity-type effect occurring at the level of the beta-adrenergic receptor. Rats were treated with reserpine (a norepinephrine depleter) and the pineals exposed to ephedrine. Endogenous norepinephrine normally released by the action of ephedrine was thus absent, and under these conditions, levels of melatonin (aMT) and N-acetylserotonin (aHT) were reduced. N-acetyltransferase (NAT) activity was also reduced, but maintained levels pointing to substantial adrenergic activity of ephedrine as well as norepinephrine released by virtue of the drug's action. A subsequent binding study showed a decrease in beta-adrenergic receptor binding with exposure to ephedrine and a further decrease in ephedrine treated pineals from reserpine treated rats, thus ruling out the occurrence of a supersensitivity phenomenon. It is possible that both ephedrine and released norepinephrine have alpha- and beta-receptor activity. Additional pineal metabolites were isolated and measured in the organ culture experiments. A 16-hour time profile of the production of melatonin (aMT) and N-acetylserotonin (aHT) with norepinephrine and ephedrine treatment provided useful information regarding the course of action of the two agents. A pineal cell-culture system was developed and exposed to ephedrine and norepinephrine. N-acetyltransferase (NAT) activity levels measured after exposure to these agents were raised, confirming the adrenergic activity of both in the model. Finally, an HPLC system coupled to a UV detector was used in an attempt to measure melatonin (aMT) extracted from pineal organ culture media. The results showed that melatonin could be measured by this method, however, a more sensitive detection system was recommended for future work
- Full Text:
- Date Issued: 1991
Algal biotechnology and the beneficiation of saline effluent wastes
- Authors: Rose, P D (Peter Dale)
- Date: 1992
- Subjects: Algae -- Biotechnology , Algae culture , Tanneries -- Waste disposal
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4135 , http://hdl.handle.net/10962/d1015967
- Description: Saline deterioration in the South African public water system has been documented and disposal of brine wastes has been identified as part of the problem. The broad aim of this research programme was to undertake an initial technical study to evaluate the feasibility of integrating algal biotechnology into a disposal function for these wastes. A demonstration of utility in the form of products and waste treatment could produce a beneficiation of saline effluents and provide incentives necessary to deal with the disposal issue. The study attempted to demonstrate a synthesis between the two main thrusts in algal biotechnology that have produced large-scale practical applications - stable, predictable algal production in saline media and the cost effective High Rate Oxidation Ponding (HROP) process for incorporating algal production into a waste treatment function. Tannery organic saline effluents and the biotechnology of Dunaliella salina culture producing β- carotene were chosen as paradigms for the study. 1. The alga was shown to grow in certain tannery effluents producing enhanced biomass yields compared to defined inorganic medium cultivation. The potential for amino acid or protein supplementation of defmed culture media was noted. 2. A reduction in organic load simultaneous with the growth of D.salina was recorded in laboratory-scale simulations of the HROP process. Rates similar to the fresh water HROP equivalent were demonstrated. 3. These results suggested the uptake and storage of organic nitrogen by D.salina. The consequent inhibition of β-carotene accumulation by the organism presented a potentially insurmountable obstacle to the feasibility of β-carotene production in this medium. Uptake and release of organic compounds, previously demonstrated in phytoplankton and other micro-algae, was confirmed in this study for D.salina. The evidence acquired indicated the internalization of both glycine and bovine serum albumin. An ultrastructural study demonstrated mechanisms by which this process might occur. 4. The release of substantial quantities of glycerol was shown. A mechanism whereby D. salina may use this to regulate ammonia availability via control of its associated bacterial population was observed. Glycerol release was identified as presenting an application in treating refractory organic wastes, such as secondary sewage sludges, by elevating C:N ratios. This could demonstrate a significant utility for brine waste impoundments. 5. A multistage production process was proposed to deal with the problem of β-carotene inhibition by separation of the growth and metabolite accumulation functions into separate unit operations. It was shown in this study that the stress of nitrogen deficiency combined with high salinity provides for effectiveβ-carotene accumulation under the conditions of low illumination that pertain in dense cultures. Subjected to these conditions effluent-grown cells show delayed but unimpaired {j-carotene accumulation. 6. A role for the plant hormone abscisic acid in mediating the stress response was demonstrated in D.salina. Fluorescence induction studies suggested the presence of a signalling process forming part of a sensitivity control mechanism. Stress induction of β-carotene accumulation could occur through four clearly defined stages. Potential was identified for using this response as a physiological probe for monitoring and regulating the stress induction process. 7. The multistage processing concept requires effective algal cell separation technology. The use of cross-flow ultrafiltration and diafiltration with a polyethersulfone tubular membrane system was demonstrated as an effective process for the recovery and washing of D. salina. Cell concentrates were produced in a viable form. 8. Process designs incorporating the findings of the research programme are presented demonstrating how effluent and organic waste treatment functions may be combined with the production of D.salina and its products. Application of the multi-stage processing concept to β-carotene production in a defined medium process was identified as offering a potential four-fold yield enhancement. This could have a significant impact on a high cost, marginal algal biotechnology process. Aspects of novelty have been claimed in provisional patents applications. A provisional demonstration of the feasibility of D.salina production in tannery effluent indicates that algal biotechnology may provide a utility for, and hence the beneficiation of saline effluent wastes.
- Full Text:
- Date Issued: 1992
- Authors: Rose, P D (Peter Dale)
- Date: 1992
- Subjects: Algae -- Biotechnology , Algae culture , Tanneries -- Waste disposal
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4135 , http://hdl.handle.net/10962/d1015967
- Description: Saline deterioration in the South African public water system has been documented and disposal of brine wastes has been identified as part of the problem. The broad aim of this research programme was to undertake an initial technical study to evaluate the feasibility of integrating algal biotechnology into a disposal function for these wastes. A demonstration of utility in the form of products and waste treatment could produce a beneficiation of saline effluents and provide incentives necessary to deal with the disposal issue. The study attempted to demonstrate a synthesis between the two main thrusts in algal biotechnology that have produced large-scale practical applications - stable, predictable algal production in saline media and the cost effective High Rate Oxidation Ponding (HROP) process for incorporating algal production into a waste treatment function. Tannery organic saline effluents and the biotechnology of Dunaliella salina culture producing β- carotene were chosen as paradigms for the study. 1. The alga was shown to grow in certain tannery effluents producing enhanced biomass yields compared to defined inorganic medium cultivation. The potential for amino acid or protein supplementation of defmed culture media was noted. 2. A reduction in organic load simultaneous with the growth of D.salina was recorded in laboratory-scale simulations of the HROP process. Rates similar to the fresh water HROP equivalent were demonstrated. 3. These results suggested the uptake and storage of organic nitrogen by D.salina. The consequent inhibition of β-carotene accumulation by the organism presented a potentially insurmountable obstacle to the feasibility of β-carotene production in this medium. Uptake and release of organic compounds, previously demonstrated in phytoplankton and other micro-algae, was confirmed in this study for D.salina. The evidence acquired indicated the internalization of both glycine and bovine serum albumin. An ultrastructural study demonstrated mechanisms by which this process might occur. 4. The release of substantial quantities of glycerol was shown. A mechanism whereby D. salina may use this to regulate ammonia availability via control of its associated bacterial population was observed. Glycerol release was identified as presenting an application in treating refractory organic wastes, such as secondary sewage sludges, by elevating C:N ratios. This could demonstrate a significant utility for brine waste impoundments. 5. A multistage production process was proposed to deal with the problem of β-carotene inhibition by separation of the growth and metabolite accumulation functions into separate unit operations. It was shown in this study that the stress of nitrogen deficiency combined with high salinity provides for effectiveβ-carotene accumulation under the conditions of low illumination that pertain in dense cultures. Subjected to these conditions effluent-grown cells show delayed but unimpaired {j-carotene accumulation. 6. A role for the plant hormone abscisic acid in mediating the stress response was demonstrated in D.salina. Fluorescence induction studies suggested the presence of a signalling process forming part of a sensitivity control mechanism. Stress induction of β-carotene accumulation could occur through four clearly defined stages. Potential was identified for using this response as a physiological probe for monitoring and regulating the stress induction process. 7. The multistage processing concept requires effective algal cell separation technology. The use of cross-flow ultrafiltration and diafiltration with a polyethersulfone tubular membrane system was demonstrated as an effective process for the recovery and washing of D. salina. Cell concentrates were produced in a viable form. 8. Process designs incorporating the findings of the research programme are presented demonstrating how effluent and organic waste treatment functions may be combined with the production of D.salina and its products. Application of the multi-stage processing concept to β-carotene production in a defined medium process was identified as offering a potential four-fold yield enhancement. This could have a significant impact on a high cost, marginal algal biotechnology process. Aspects of novelty have been claimed in provisional patents applications. A provisional demonstration of the feasibility of D.salina production in tannery effluent indicates that algal biotechnology may provide a utility for, and hence the beneficiation of saline effluent wastes.
- Full Text:
- Date Issued: 1992
An investigation into the potential immunogenicity of various extracts of the South African bont tick Amblyomma hebraeum
- Authors: Adamson, Deborah Jane
- Date: 1993
- Subjects: Amblyomma -- South Africa , Ticks -- South Africa , Ticks -- Control -- South Africa
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4127 , http://hdl.handle.net/10962/d1015640
- Description: Rabbits and goats were inoculated with crude, membrane-associated and soluble components extracted from unengorged adult females and nymphs of the bont tick Amblyomma hebraeum. Inoculation provided some protection against nymphal infestation, however it had little effect on adult feeding. Histological examination of adults fed on inoculated hosts showed evidence of gut damage. Skin provocation testing with tick extracts elicited a Type I immediate hypersensitivity which was influenced by antihistamine. A delayed skin reaction was also evident. Whether this was attributable to Type III Arthus reaction or Type IV cell-mediated hypersensitivity was not determined. A comparative histological study of sites of tick extract injection, on inoculated and naive hosts, demonstrated the role of eosinophils in the hosts response to tick feeding. Serological examination revealed elevated anti-A hebraeum lgG titres following inoculation. These titres were found to decrease in the ten weeks after inoculation, despite the hosts being repeatedly infested with A hebraeum. Although the IgG titres of naive control hosts increased after each tick infestation, they failed to reach the titres achieved through inoculation. Western blot analysis of serum from inoculated hosts recognized most of the A. hebraeum proteins against which it was screened.
- Full Text:
- Date Issued: 1993
- Authors: Adamson, Deborah Jane
- Date: 1993
- Subjects: Amblyomma -- South Africa , Ticks -- South Africa , Ticks -- Control -- South Africa
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4127 , http://hdl.handle.net/10962/d1015640
- Description: Rabbits and goats were inoculated with crude, membrane-associated and soluble components extracted from unengorged adult females and nymphs of the bont tick Amblyomma hebraeum. Inoculation provided some protection against nymphal infestation, however it had little effect on adult feeding. Histological examination of adults fed on inoculated hosts showed evidence of gut damage. Skin provocation testing with tick extracts elicited a Type I immediate hypersensitivity which was influenced by antihistamine. A delayed skin reaction was also evident. Whether this was attributable to Type III Arthus reaction or Type IV cell-mediated hypersensitivity was not determined. A comparative histological study of sites of tick extract injection, on inoculated and naive hosts, demonstrated the role of eosinophils in the hosts response to tick feeding. Serological examination revealed elevated anti-A hebraeum lgG titres following inoculation. These titres were found to decrease in the ten weeks after inoculation, despite the hosts being repeatedly infested with A hebraeum. Although the IgG titres of naive control hosts increased after each tick infestation, they failed to reach the titres achieved through inoculation. Western blot analysis of serum from inoculated hosts recognized most of the A. hebraeum proteins against which it was screened.
- Full Text:
- Date Issued: 1993
The microbial production of polyphenol oxidase enzyme systems and their application in the treatment of phenolic wastewaters
- Authors: Goetsch, Patricia-Ann
- Date: 1993
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:21060 , http://hdl.handle.net/10962/6190
- Description: Phenolic compounds are a group of organic chemicals present in the wastewaters of many synthetic industrial processes. Due to their extreme toxicity to man and animals, and deleterious impact on the environment, a range of techniques exist for the effective treatment and disposal of these pollutants. Biological degradation using microbial enzymes presents a valuable alternative to conventional wastewater treatment systems. This research was therefore initiated to investigate the polyphenol oxidase enzyme system and the feasibility of its application for effluent treatment and studies in organic solvents. The enzyme system is widely distributed in nature, with Agaricus bisporus (the common mushroom) being the best known producer. Biochemical investigations of the enzyme system were therefore carried out using this extract. A screening programme was initiated to identify microbial polyphenol oxidase producers which could be cultured in liquid media, thereby enabling the production of large quantities of enzyme in fermentation systems. Extensive growth optimization and enzyme induction and optimization studies were carried out on selected cultures. A number of good producers were isolated, namely a bacterial culture designated AECI culture no. 26, Streptomyces antibioticus, Streptomyces glaucescens and a manipulated strain, Streptomyces lividans (pIJ702). Enzyme production by Agaricus bisporus mycelia was optimized in deep-liquid culture; enzyme extracts showed high phenol removal efficiencies. Streptomyces antibioticus, Streptomyces glaucescens, Streptomyces lividans (pIJ702) and AECI culture no. 26 whole cells were also investigated for phenol-removing ability in simulated phenolic effluents. The use of whole cells reduces enzyme inactivation and instability due to the protection of the enzyme system within the cell. All cultures showed improved removal efficiencies in phenolic growth media. These results strongly suggest their use for phenol removal in continuous systems.
- Full Text:
- Date Issued: 1993
- Authors: Goetsch, Patricia-Ann
- Date: 1993
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:21060 , http://hdl.handle.net/10962/6190
- Description: Phenolic compounds are a group of organic chemicals present in the wastewaters of many synthetic industrial processes. Due to their extreme toxicity to man and animals, and deleterious impact on the environment, a range of techniques exist for the effective treatment and disposal of these pollutants. Biological degradation using microbial enzymes presents a valuable alternative to conventional wastewater treatment systems. This research was therefore initiated to investigate the polyphenol oxidase enzyme system and the feasibility of its application for effluent treatment and studies in organic solvents. The enzyme system is widely distributed in nature, with Agaricus bisporus (the common mushroom) being the best known producer. Biochemical investigations of the enzyme system were therefore carried out using this extract. A screening programme was initiated to identify microbial polyphenol oxidase producers which could be cultured in liquid media, thereby enabling the production of large quantities of enzyme in fermentation systems. Extensive growth optimization and enzyme induction and optimization studies were carried out on selected cultures. A number of good producers were isolated, namely a bacterial culture designated AECI culture no. 26, Streptomyces antibioticus, Streptomyces glaucescens and a manipulated strain, Streptomyces lividans (pIJ702). Enzyme production by Agaricus bisporus mycelia was optimized in deep-liquid culture; enzyme extracts showed high phenol removal efficiencies. Streptomyces antibioticus, Streptomyces glaucescens, Streptomyces lividans (pIJ702) and AECI culture no. 26 whole cells were also investigated for phenol-removing ability in simulated phenolic effluents. The use of whole cells reduces enzyme inactivation and instability due to the protection of the enzyme system within the cell. All cultures showed improved removal efficiencies in phenolic growth media. These results strongly suggest their use for phenol removal in continuous systems.
- Full Text:
- Date Issued: 1993
Preliminary investigation of the molecular pathogenicity determinants of Xanthomonas campestris pv. zeae
- Authors: Downing, T G
- Date: 1998
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4066 , http://hdl.handle.net/10962/d1004922
- Description: Xanthomonas campestris pv. zeae was shown to posses a vast range of potential plant cell wall degrading enzymes including at least one protease, a carboxymethylcellulase, pectin lyase, polygalacturonase and a B-endoglucanase. Replicon stability and transfer system efficiencies were determined for a range of oriC, and oriT -tra combinations, and suitable vectors were constructed and identified for insertional inactivation by homologous recombination. Ideal suicide replicons were found to be pACYC184 and p15A, while P, Wand Q group replicons were supported by X campestris pv. zeae. Group P and group N transfer systems were shown to be highly efficient in intra-genus matings between Escherichia coli and X campestris pv. zeae, with the exception of P-tra systems in trans for the delivery of Tn5. Various cloning vectors were tested for stability and mobility. Tn5 was shown to transpose at high frequencies into the genome of the bacterial plant pathogen, and insertion was relatively random. Suitable screening assays were established to allow rapid isolation of mutants with potential virulence or pathogenic deviations, after mutagenesis. Two non-pathogenic mutants were identified, one of which was a putative hrp·, while the other was a leaky virulence. A single mutant showing 40% reduced protease activity was also shown to exhibit reduced virulence indicating a minor role for the protease in pathogenicity. The majority of virulence mutants showed altered growth in different levels of nutritional availability and complexity. Nutritional viability (the ability to acquire and use nutrients at a sufficient rate to grow fast enough to overcome host defences) was shown to be essential for virulence and possibly pathogenicity. Wild-type in-planta behaviour was analysed and growth and spread patterns typical for pathogenic response identified. Chief amongst these was the requirement for a threshold level of cells per leaf area or length, before symptoms could develop. Occlusion of vascular bundles was shown not to be the primary factor in the pathogenicity of X campestris pv. zeae. Threshold levels for lesion development indicate the absence of a diffusable lesion forming element, and possibly the requirement of cell density for induction of certain functions. , KMBT_363
- Full Text:
- Date Issued: 1998
- Authors: Downing, T G
- Date: 1998
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4066 , http://hdl.handle.net/10962/d1004922
- Description: Xanthomonas campestris pv. zeae was shown to posses a vast range of potential plant cell wall degrading enzymes including at least one protease, a carboxymethylcellulase, pectin lyase, polygalacturonase and a B-endoglucanase. Replicon stability and transfer system efficiencies were determined for a range of oriC, and oriT -tra combinations, and suitable vectors were constructed and identified for insertional inactivation by homologous recombination. Ideal suicide replicons were found to be pACYC184 and p15A, while P, Wand Q group replicons were supported by X campestris pv. zeae. Group P and group N transfer systems were shown to be highly efficient in intra-genus matings between Escherichia coli and X campestris pv. zeae, with the exception of P-tra systems in trans for the delivery of Tn5. Various cloning vectors were tested for stability and mobility. Tn5 was shown to transpose at high frequencies into the genome of the bacterial plant pathogen, and insertion was relatively random. Suitable screening assays were established to allow rapid isolation of mutants with potential virulence or pathogenic deviations, after mutagenesis. Two non-pathogenic mutants were identified, one of which was a putative hrp·, while the other was a leaky virulence. A single mutant showing 40% reduced protease activity was also shown to exhibit reduced virulence indicating a minor role for the protease in pathogenicity. The majority of virulence mutants showed altered growth in different levels of nutritional availability and complexity. Nutritional viability (the ability to acquire and use nutrients at a sufficient rate to grow fast enough to overcome host defences) was shown to be essential for virulence and possibly pathogenicity. Wild-type in-planta behaviour was analysed and growth and spread patterns typical for pathogenic response identified. Chief amongst these was the requirement for a threshold level of cells per leaf area or length, before symptoms could develop. Occlusion of vascular bundles was shown not to be the primary factor in the pathogenicity of X campestris pv. zeae. Threshold levels for lesion development indicate the absence of a diffusable lesion forming element, and possibly the requirement of cell density for induction of certain functions. , KMBT_363
- Full Text:
- Date Issued: 1998
Capillary membrane-immobilised polyphenol oxidase and the bioremediation of industrial phenolic effluent
- Authors: Edwards, Wade
- Date: 1999
- Subjects: Membranes (Technology) , Effluent quality , Pollutants , Phenols , Water -- Purification
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4095 , http://hdl.handle.net/10962/d1008458
- Description: Waste-generating industrialisation is intrinsically associated with population and economic proliferation. This places considerable emphasis on South Africa's water shortage due to the integral relationship between population growth rate and infrastructure development. Of the various types of industry-generated effluents, those containing organic pollutants such as phenols are generally difficult to remediate. Much work has been reported in the literature on the use of enzymes for the removal of phenols from these waste-streams but little application of this bioremediation approach has reached practical fruition. This study focuses on integrating and synergistically combining the advantages of enzyme-mediated dephenolisation of synthetic and industrial effluent with that of membrane teclmology. The ability of the enzyme polyphenol oxidase to convert phenol and a number of its derivatives to chemically reactive o-quinones has been reported extensively in the literature. These o-quinones can then physically be removed from solution using various precipitation or adsorption techniques. The enzyme is, however, plagued by a product-induced phenomenon known as suicide inactivation, which renders it inactive and thus limits its application as a bioremediation tool. Integrating membrane technology with the enzyme's catalytic ability by immobilising polyphenol oxidase onto polysulphone and poly(ether sulphone) capillary membranes enabled the physical removal of these inhibitory products from the micro-environment of the immobilised enzyme which therefore increased the phenol conversion capability of the immobilised biocatalyst. Under non-immobilised conditions it was found that when exposed to a mixture of various phenols the substrate preference of the enzyme is a function of the R-group. Under immobilised conditions, however, the substrate preference of the enzyme becomes a function of certain transport constraints imposed by the capillary membrane itself. Furthermore, by integrating a quinone-removal process in the enzyme-immobilised bioreactor configuration, a 21-fold increase in the amount of substrate converted per Unit enzyme was observed when compared to the conversion capacity of the inunobilised enzyme without the product removal step. Comparisons were also made using different membrane bioreactor configurations (orientating the capillaries transverse as opposed to parallel to the module axis) and different immobilisation matrices (poly(ether sulphone) and polysulphone capillary membranes). Conversion efficiencies as high as 77% were maintained for several hours using the combination of transverse-flow modules and novel polysulphone capillary membranes. It was therefore concluded that immobilisation of polyphenol oxidase on capillary membranes does indeed show considerable potential for future development.
- Full Text:
- Date Issued: 1999
- Authors: Edwards, Wade
- Date: 1999
- Subjects: Membranes (Technology) , Effluent quality , Pollutants , Phenols , Water -- Purification
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4095 , http://hdl.handle.net/10962/d1008458
- Description: Waste-generating industrialisation is intrinsically associated with population and economic proliferation. This places considerable emphasis on South Africa's water shortage due to the integral relationship between population growth rate and infrastructure development. Of the various types of industry-generated effluents, those containing organic pollutants such as phenols are generally difficult to remediate. Much work has been reported in the literature on the use of enzymes for the removal of phenols from these waste-streams but little application of this bioremediation approach has reached practical fruition. This study focuses on integrating and synergistically combining the advantages of enzyme-mediated dephenolisation of synthetic and industrial effluent with that of membrane teclmology. The ability of the enzyme polyphenol oxidase to convert phenol and a number of its derivatives to chemically reactive o-quinones has been reported extensively in the literature. These o-quinones can then physically be removed from solution using various precipitation or adsorption techniques. The enzyme is, however, plagued by a product-induced phenomenon known as suicide inactivation, which renders it inactive and thus limits its application as a bioremediation tool. Integrating membrane technology with the enzyme's catalytic ability by immobilising polyphenol oxidase onto polysulphone and poly(ether sulphone) capillary membranes enabled the physical removal of these inhibitory products from the micro-environment of the immobilised enzyme which therefore increased the phenol conversion capability of the immobilised biocatalyst. Under non-immobilised conditions it was found that when exposed to a mixture of various phenols the substrate preference of the enzyme is a function of the R-group. Under immobilised conditions, however, the substrate preference of the enzyme becomes a function of certain transport constraints imposed by the capillary membrane itself. Furthermore, by integrating a quinone-removal process in the enzyme-immobilised bioreactor configuration, a 21-fold increase in the amount of substrate converted per Unit enzyme was observed when compared to the conversion capacity of the inunobilised enzyme without the product removal step. Comparisons were also made using different membrane bioreactor configurations (orientating the capillaries transverse as opposed to parallel to the module axis) and different immobilisation matrices (poly(ether sulphone) and polysulphone capillary membranes). Conversion efficiencies as high as 77% were maintained for several hours using the combination of transverse-flow modules and novel polysulphone capillary membranes. It was therefore concluded that immobilisation of polyphenol oxidase on capillary membranes does indeed show considerable potential for future development.
- Full Text:
- Date Issued: 1999
The development of an immobilised-enzyme bioprobe for the detection of phenolic pollutants in water
- Authors: Russell, Ingrid Margaret
- Date: 1999
- Subjects: Pollutants -- Biodegradation , Pollutants , Chemical reactors , Membrane reactors , Fungi -- Biotechnology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4069 , http://hdl.handle.net/10962/d1006211 , Pollutants -- Biodegradation , Pollutants , Chemical reactors , Membrane reactors , Fungi -- Biotechnology
- Description: The possibility of developing an immobilised-enzyme bioprobe, based on mushroom polyphenol oxidase, for the purely biological detection and quantification of phenolic pollutants in water was investigated. Polyphenol oxidase catalyses the bioconversion of many phenolic compounds into quinone-related coloured products. Thus, in an immobilised form, the enzyme serves as a visible indicator of the presence and concentration of phenolic pollutants in water. The objective of this research was to develop a portable, disposable bioprobe incorporating polyphenol oxidase for this purpose. The intensity of the colour changes produced by the enzyme on reaction with p-cresol, p-chlorophenol and phenol was found to increase proportionally with increasing concentrations of these substrates in solution. Immobilisation of the enzyme on various supports did not appear to significantly affect the catalytic activity of the enzyme. The enzyme was immobilised by adsorption and cross-linking on polyethersulphone, nitrocellulose and nylon membranes with the production of various colour ranges on reaction with the phenolic substrates. The most successful immobilisation of the enzyme, in terms of quantity and distribution of enzyme immobilised and colour production, was obtained with the enzyme immobilised by adsorption on nylon membranes in the presence of 3-methyl-2-benzothiazolinone hydrazone (MBTH). The enzyme, immobilised using this method, produced ranges of maroon colours in phenolic solutions and orange colours in cresylic solutions. The colour intensities produced were found to increase proportionally with increasing substrate concentration after 5 minutes exposure to the substrates. The bioprobe had a broad substrate specificity and was sensitive to substrate concentrations down to 0.05 mg/L. The enzyme activity of the bioprobe was not significantly affected in a pH range from 4 to 10 and in a temperature range from 5-25⁰C. The bioprobe activity was not affected by various concentrations of salt and metal ions and the bioprobe was able to detect and semi-quantify phenolic substrates in industrial effluent samples. These features of the bioprobe indicate that the commercialisation of such a bioprobe is feasible and this technology has been patented (Patent No. SA 97/0227). , KMBT_363 , Adobe Acrobat 9.54 Paper Capture Plug-in
- Full Text:
- Date Issued: 1999
- Authors: Russell, Ingrid Margaret
- Date: 1999
- Subjects: Pollutants -- Biodegradation , Pollutants , Chemical reactors , Membrane reactors , Fungi -- Biotechnology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4069 , http://hdl.handle.net/10962/d1006211 , Pollutants -- Biodegradation , Pollutants , Chemical reactors , Membrane reactors , Fungi -- Biotechnology
- Description: The possibility of developing an immobilised-enzyme bioprobe, based on mushroom polyphenol oxidase, for the purely biological detection and quantification of phenolic pollutants in water was investigated. Polyphenol oxidase catalyses the bioconversion of many phenolic compounds into quinone-related coloured products. Thus, in an immobilised form, the enzyme serves as a visible indicator of the presence and concentration of phenolic pollutants in water. The objective of this research was to develop a portable, disposable bioprobe incorporating polyphenol oxidase for this purpose. The intensity of the colour changes produced by the enzyme on reaction with p-cresol, p-chlorophenol and phenol was found to increase proportionally with increasing concentrations of these substrates in solution. Immobilisation of the enzyme on various supports did not appear to significantly affect the catalytic activity of the enzyme. The enzyme was immobilised by adsorption and cross-linking on polyethersulphone, nitrocellulose and nylon membranes with the production of various colour ranges on reaction with the phenolic substrates. The most successful immobilisation of the enzyme, in terms of quantity and distribution of enzyme immobilised and colour production, was obtained with the enzyme immobilised by adsorption on nylon membranes in the presence of 3-methyl-2-benzothiazolinone hydrazone (MBTH). The enzyme, immobilised using this method, produced ranges of maroon colours in phenolic solutions and orange colours in cresylic solutions. The colour intensities produced were found to increase proportionally with increasing substrate concentration after 5 minutes exposure to the substrates. The bioprobe had a broad substrate specificity and was sensitive to substrate concentrations down to 0.05 mg/L. The enzyme activity of the bioprobe was not significantly affected in a pH range from 4 to 10 and in a temperature range from 5-25⁰C. The bioprobe activity was not affected by various concentrations of salt and metal ions and the bioprobe was able to detect and semi-quantify phenolic substrates in industrial effluent samples. These features of the bioprobe indicate that the commercialisation of such a bioprobe is feasible and this technology has been patented (Patent No. SA 97/0227). , KMBT_363 , Adobe Acrobat 9.54 Paper Capture Plug-in
- Full Text:
- Date Issued: 1999
The Rhodes BioSure process in the treatment of acid mine drainage wastewaters
- Authors: Corbett, Christopher John
- Date: 2001 , 2013-05-03
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4077 , http://hdl.handle.net/10962/d1007405
- Description: While sulphate-enriched wastewaters are generated in a number of industrial processes, such as tanning, paper manufacture and metals processing, the principal contributors to large-scale pollution from this source in South Africa are the gold and coal mining industries. Both biological and physico-chemical processes, set in train by mining operations, give rise to the oxidation of sulphur species, and the resultant generation of AMD. The Vaal River system is most affected and receives large tonnages of mining related salinity as both direct discharges, and in diffuse runoff flows. The long-term burden of this problem, and sustaining ongoing treatment over the time-frames involved will almost certainly resort to the community inhabiting the area, notwithstanding progressive mine closure legislation and comprehensive regulation governing the polluterpays principle. The volume and time-frame of the AMD problem, and the need for a long-term and sustainable response has focused interest in biological treatment approaches. These have concentrated on active and passive treatment systems, both of which rely on microbial activity related to the biological sulphur cycle. Notwithstanding the reactor type, and the particular treatment approach used, widespread application of active AMD treatment has not yet been seen on any large scale. Singular factors constraining process development are bioreactor design, cost of bioreactor construction, and the cost of the carbon source and electron donor for the biological sulphate reduction process. The SRB are able to utilise only a limited range of small organic molecules. The studies reported here were motivated by the need to evaluate low-cost options and the treatment of high volume AMD flows. This has focussed research activity on bioprocess developments using complex organic compounds derived from waste streams as electron donor sources, and the integration of AMD treatment with other waste treatment objectives. The co-disposal of organic wastes with AMD treatment would enable the development of an 'integrated resource management' approach to the problem, including sustainability of treatment operations over the long time-frames involved. Apart from the cost advantages accrued to waste treatment, the recovery of the treated water as a resource to the wider community provides a potentially important value-added function to the combined operation. , KMBT_363 , Adobe Acrobat 9.53 Paper Capture Plug-in
- Full Text:
- Date Issued: 2001
- Authors: Corbett, Christopher John
- Date: 2001 , 2013-05-03
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4077 , http://hdl.handle.net/10962/d1007405
- Description: While sulphate-enriched wastewaters are generated in a number of industrial processes, such as tanning, paper manufacture and metals processing, the principal contributors to large-scale pollution from this source in South Africa are the gold and coal mining industries. Both biological and physico-chemical processes, set in train by mining operations, give rise to the oxidation of sulphur species, and the resultant generation of AMD. The Vaal River system is most affected and receives large tonnages of mining related salinity as both direct discharges, and in diffuse runoff flows. The long-term burden of this problem, and sustaining ongoing treatment over the time-frames involved will almost certainly resort to the community inhabiting the area, notwithstanding progressive mine closure legislation and comprehensive regulation governing the polluterpays principle. The volume and time-frame of the AMD problem, and the need for a long-term and sustainable response has focused interest in biological treatment approaches. These have concentrated on active and passive treatment systems, both of which rely on microbial activity related to the biological sulphur cycle. Notwithstanding the reactor type, and the particular treatment approach used, widespread application of active AMD treatment has not yet been seen on any large scale. Singular factors constraining process development are bioreactor design, cost of bioreactor construction, and the cost of the carbon source and electron donor for the biological sulphate reduction process. The SRB are able to utilise only a limited range of small organic molecules. The studies reported here were motivated by the need to evaluate low-cost options and the treatment of high volume AMD flows. This has focussed research activity on bioprocess developments using complex organic compounds derived from waste streams as electron donor sources, and the integration of AMD treatment with other waste treatment objectives. The co-disposal of organic wastes with AMD treatment would enable the development of an 'integrated resource management' approach to the problem, including sustainability of treatment operations over the long time-frames involved. Apart from the cost advantages accrued to waste treatment, the recovery of the treated water as a resource to the wider community provides a potentially important value-added function to the combined operation. , KMBT_363 , Adobe Acrobat 9.53 Paper Capture Plug-in
- Full Text:
- Date Issued: 2001
Biological generation of reactive alkaline species and their application in a sustainable bioprocess for the remediation of acid and metal contaminated wastewaters
- Authors: Van Hille, Robert Paul
- Date: 2002
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:21049 , http://hdl.handle.net/10962/6129
- Description: This project focused on the development of an integrated biological system for the treatment of acidic and metal-laden effluents, based on the sustainable biological generation of reactive alkaline species. Initial studies concentrated on the binding and accumulation of heavy metals by biomass of the cyanobacteria, Spirulina sp. Metal binding was rapid, with saturation reached in 30 minutes, and followed an affinity series of Pb > Cu > Zn >>Fe. The binding capacity of the Spirulina for each of the metals was relatively low when compared to a range of other biosorbents. The toxicity thresholds of the algae was determined for copper and zinc. These were low (10umoles/g) and as such, the algae were not suitable for application in a treatment system in which they came into direct contact with the toxic metals. The algae were able to increase the pH of the surrounding medium. This occurred as a result of the accumulation of inorganic carbon, from bicarbonate, as a response to low concentrations of carbon dioxide in the medium. The resulting release of a hydroxide ion into solution led to the increase in pH. The increase in pH was shown to be due to a reduction in acidity, rather than an increase in alkalinity. The enzyme carbonic anhydrase was shown to be pivotal in this system. Attempts to determine the enzyme activity directly were unsuccessful, due to the inherent inaccuracy of the assay system. An indirect method of determining enzyme activity, by measuring changes in the carbonate species equilibrium, was developed. Under optimal conditions Spirulina was able to reduce the acidity by an amount equivalent to the addition of 3670umoles NaOH g·' h·'. Predictive modelling showed that this enhanced the potential of the medium to effect metal precipitation. For the algal system to be sustainable, a readily available source of bicarbonate was needed. This was achieved by the oxidation of organic carbon, under sulphidogenic conditions, by a bacterial consortium isolated from the anaerobic component of a facultative pond. The consortium was shown to consist of sulphate reducing (most likely Desulvovibrio and Desulfotomaculum)and acetogenic bacteria. Sulphate removal rates of 500mg 1·' day·' and 135mg 1·' day·' were achieved in a 21 agitated and 281 upflow reactor respectively. The bicarbonate generation rate in the 281 reactor was calculated as 4033umoles 1·' day·', which proved sufficient to act as a feed for the algal system. Sparging the anaerobic digester overflow with air and nitrogen resulted in a reduction in the aqueous sulphide concentration. Using nitrogen, a 70% recovery of sulphide, as H2S gas, was achieved in 60 minutes, while with air, this dropped to 40%, due to the oxidation of the aqueous sulphide. The stripping ofH2S resulted in an increase in pH. The H2S gas was used for the selective precipitation of copper and lead in the integrated system. The dynamics of metal precipitation was investigated. For simple reactions, between individual IV metal and base species, it was possible to generate an accurate predictive model and confirm the precipitating species using wavelength dispersive X-ray spectroscopy (WDS). In more complex systems, where precipitation of the artificial acid mine drainage was examined, the predictive modelling and WDS could not accurately describe the system. The addition of aqueous sulphide to copper and iron resulted in the formation of metastable, amorphous precipitates, which remained in suspension. Ageing of the copper precipitate resulted in the evolution of a stable crystalline structure (covellite) and the aggregation and settling of the precipitate. In the case of iron, the amorphous precipitate underwent oxidation before a stable iron sulphide could evolve and the settled precipitate was an iron oxide or oxyhydroxide. The artificial acid mine drainage was treated with sulphide, hydroxide, anaerobic digester overflow and algal overflow. The best metal removal was achieved with the sulphide and hydroxide, while the algal overflow outperformed the anaerobic digester overflow. The precipitate generated by the addition of sulphide was the most compact, followed by the algal overflow, the anaerobic digester overflow and the hydroxide. Efficient precipitation of all the heavy metals, except manganese, was achieved using the algal overflow at an acidity to alkalinity ratio of 1 :2. This ratio was selected for use in the pilot system. The Spirulina based pilot system was effectively used to treat an effluent from the Black Mountain base metal mine. The necessity to maintain the algae in suspension and avoid biomass washout were practical considerations which counted against this system. The replacement of the Spirulina by Oscillatoria, which adhered to a solid support, overcame these problems. The integrated biological system was able to effectively treat an artificial acid mine drainage for 90 days, reducing the concentration of all metals, except manganese, to below the acceptable environmental risk levels. The treatment of the final effluent in a second anaerobic digester reduced the manganese concentration to 4.5uM and proved that the sulphate reducing bacteria could be cultivated on enriched, partially treated acid mine drainage. The integrated biological treatment system performed well, effectively treating an effluent modelled closely on the quality of the water being discharged from the East Rand Basin. The cost of such a system would be considerably less than a "high tech" physico-chemical system. This, coupled with the potential long term sustainability of a biological system, would make it a potentially attractive option for the treatment of future acid mine drainage discharges.
- Full Text:
- Date Issued: 2002
- Authors: Van Hille, Robert Paul
- Date: 2002
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:21049 , http://hdl.handle.net/10962/6129
- Description: This project focused on the development of an integrated biological system for the treatment of acidic and metal-laden effluents, based on the sustainable biological generation of reactive alkaline species. Initial studies concentrated on the binding and accumulation of heavy metals by biomass of the cyanobacteria, Spirulina sp. Metal binding was rapid, with saturation reached in 30 minutes, and followed an affinity series of Pb > Cu > Zn >>Fe. The binding capacity of the Spirulina for each of the metals was relatively low when compared to a range of other biosorbents. The toxicity thresholds of the algae was determined for copper and zinc. These were low (10umoles/g) and as such, the algae were not suitable for application in a treatment system in which they came into direct contact with the toxic metals. The algae were able to increase the pH of the surrounding medium. This occurred as a result of the accumulation of inorganic carbon, from bicarbonate, as a response to low concentrations of carbon dioxide in the medium. The resulting release of a hydroxide ion into solution led to the increase in pH. The increase in pH was shown to be due to a reduction in acidity, rather than an increase in alkalinity. The enzyme carbonic anhydrase was shown to be pivotal in this system. Attempts to determine the enzyme activity directly were unsuccessful, due to the inherent inaccuracy of the assay system. An indirect method of determining enzyme activity, by measuring changes in the carbonate species equilibrium, was developed. Under optimal conditions Spirulina was able to reduce the acidity by an amount equivalent to the addition of 3670umoles NaOH g·' h·'. Predictive modelling showed that this enhanced the potential of the medium to effect metal precipitation. For the algal system to be sustainable, a readily available source of bicarbonate was needed. This was achieved by the oxidation of organic carbon, under sulphidogenic conditions, by a bacterial consortium isolated from the anaerobic component of a facultative pond. The consortium was shown to consist of sulphate reducing (most likely Desulvovibrio and Desulfotomaculum)and acetogenic bacteria. Sulphate removal rates of 500mg 1·' day·' and 135mg 1·' day·' were achieved in a 21 agitated and 281 upflow reactor respectively. The bicarbonate generation rate in the 281 reactor was calculated as 4033umoles 1·' day·', which proved sufficient to act as a feed for the algal system. Sparging the anaerobic digester overflow with air and nitrogen resulted in a reduction in the aqueous sulphide concentration. Using nitrogen, a 70% recovery of sulphide, as H2S gas, was achieved in 60 minutes, while with air, this dropped to 40%, due to the oxidation of the aqueous sulphide. The stripping ofH2S resulted in an increase in pH. The H2S gas was used for the selective precipitation of copper and lead in the integrated system. The dynamics of metal precipitation was investigated. For simple reactions, between individual IV metal and base species, it was possible to generate an accurate predictive model and confirm the precipitating species using wavelength dispersive X-ray spectroscopy (WDS). In more complex systems, where precipitation of the artificial acid mine drainage was examined, the predictive modelling and WDS could not accurately describe the system. The addition of aqueous sulphide to copper and iron resulted in the formation of metastable, amorphous precipitates, which remained in suspension. Ageing of the copper precipitate resulted in the evolution of a stable crystalline structure (covellite) and the aggregation and settling of the precipitate. In the case of iron, the amorphous precipitate underwent oxidation before a stable iron sulphide could evolve and the settled precipitate was an iron oxide or oxyhydroxide. The artificial acid mine drainage was treated with sulphide, hydroxide, anaerobic digester overflow and algal overflow. The best metal removal was achieved with the sulphide and hydroxide, while the algal overflow outperformed the anaerobic digester overflow. The precipitate generated by the addition of sulphide was the most compact, followed by the algal overflow, the anaerobic digester overflow and the hydroxide. Efficient precipitation of all the heavy metals, except manganese, was achieved using the algal overflow at an acidity to alkalinity ratio of 1 :2. This ratio was selected for use in the pilot system. The Spirulina based pilot system was effectively used to treat an effluent from the Black Mountain base metal mine. The necessity to maintain the algae in suspension and avoid biomass washout were practical considerations which counted against this system. The replacement of the Spirulina by Oscillatoria, which adhered to a solid support, overcame these problems. The integrated biological system was able to effectively treat an artificial acid mine drainage for 90 days, reducing the concentration of all metals, except manganese, to below the acceptable environmental risk levels. The treatment of the final effluent in a second anaerobic digester reduced the manganese concentration to 4.5uM and proved that the sulphate reducing bacteria could be cultivated on enriched, partially treated acid mine drainage. The integrated biological treatment system performed well, effectively treating an effluent modelled closely on the quality of the water being discharged from the East Rand Basin. The cost of such a system would be considerably less than a "high tech" physico-chemical system. This, coupled with the potential long term sustainability of a biological system, would make it a potentially attractive option for the treatment of future acid mine drainage discharges.
- Full Text:
- Date Issued: 2002
The independent high rate algal pond as a unit operation in tertiary wastewater treatment
- Authors: Clark, Stewart James
- Date: 2002
- Subjects: Algae -- Biotechnology , Sewage -- Purification -- Biological treatment
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4092 , http://hdl.handle.net/10962/d1007805
- Description: The development of the High Rate Algal Pond (HRAP) as an independent tertiary treatment unit operation for phosphate and nitrate removal is reported. A novel Integrated Algal Ponding System (lAPS) design is proposed for nutrient removal from the effluents of both a conventional domestic sewage treatment plant and from an Advanced Integrated Wastewater Ponding System (AIWPS). The viability of an independently operated HRAP has been identified and termed the Independent High Rate Algal Pond (l-HRAP). A 500 m² pilot 1- HRAP was operated in such a way as to facilitate the precipitation of calcium phosphate, known to be controlled by pH (greater than 9.4) and resulting in final phosphate levels of less than 1 mg.L⁻¹ as P0₄-P. The incorporation of the I-HRAP into a denitrification process was also investigated. Continuously fed column reactors, utilising algal biomass as a carbon source, showed that the heterotrophic bacterial community dominant in the anaerobic algal sludge were denitrifying the nitrate in the feed. It was demonstrated that as the cultures were stressed (using increased nitrate concentrations, anaerobiosis and light starvation) total polysaccharide (TPS) concentrations increased, with a notable increase 111 the exopolysaccharide (EPS) fraction. These experiments corroborated the hypothesis that harvested microalgal biomass can be manipulated to produce, and release, exopolymeric substances under stress conditions, and which may serve as carbon source for denitrification. In both batch flask studies and in laboratory-scale reactor systems, harvested microalgal biomass from an HRAP was shown to produce exopolymeric substances under stress conditions. Initial high loading-rates of greater than 20 mg.L⁻¹ NO₃-N resulted in double the amount of exopolysaccharide production than in flasks with initial low loading-rates (less than 5 mg.L⁻¹ NO₃-N). Making use of an upflow anaerobic sludge blanket-type degrading-bed reactor, and an anaerobic, flooded trickle filter (ANTRIC) receiving HRAP effluent, the relationship between denitrification and the changes in polysaccharide content was investigated. This phenomenon has considerable beneficial implications in biological wastewater treatment systems where high nitrate concentration in the final effluent is a potential mitigating factor. Identification of the heterotrophic bacteria active in the denitrification process was attempted. This study presents a first report on the development and operation of the I-HRAP and has been followed by a technical-scale pilot plant evaluation of the process in the tertiary treatment of domestic wastewaters.
- Full Text:
- Date Issued: 2002
- Authors: Clark, Stewart James
- Date: 2002
- Subjects: Algae -- Biotechnology , Sewage -- Purification -- Biological treatment
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4092 , http://hdl.handle.net/10962/d1007805
- Description: The development of the High Rate Algal Pond (HRAP) as an independent tertiary treatment unit operation for phosphate and nitrate removal is reported. A novel Integrated Algal Ponding System (lAPS) design is proposed for nutrient removal from the effluents of both a conventional domestic sewage treatment plant and from an Advanced Integrated Wastewater Ponding System (AIWPS). The viability of an independently operated HRAP has been identified and termed the Independent High Rate Algal Pond (l-HRAP). A 500 m² pilot 1- HRAP was operated in such a way as to facilitate the precipitation of calcium phosphate, known to be controlled by pH (greater than 9.4) and resulting in final phosphate levels of less than 1 mg.L⁻¹ as P0₄-P. The incorporation of the I-HRAP into a denitrification process was also investigated. Continuously fed column reactors, utilising algal biomass as a carbon source, showed that the heterotrophic bacterial community dominant in the anaerobic algal sludge were denitrifying the nitrate in the feed. It was demonstrated that as the cultures were stressed (using increased nitrate concentrations, anaerobiosis and light starvation) total polysaccharide (TPS) concentrations increased, with a notable increase 111 the exopolysaccharide (EPS) fraction. These experiments corroborated the hypothesis that harvested microalgal biomass can be manipulated to produce, and release, exopolymeric substances under stress conditions, and which may serve as carbon source for denitrification. In both batch flask studies and in laboratory-scale reactor systems, harvested microalgal biomass from an HRAP was shown to produce exopolymeric substances under stress conditions. Initial high loading-rates of greater than 20 mg.L⁻¹ NO₃-N resulted in double the amount of exopolysaccharide production than in flasks with initial low loading-rates (less than 5 mg.L⁻¹ NO₃-N). Making use of an upflow anaerobic sludge blanket-type degrading-bed reactor, and an anaerobic, flooded trickle filter (ANTRIC) receiving HRAP effluent, the relationship between denitrification and the changes in polysaccharide content was investigated. This phenomenon has considerable beneficial implications in biological wastewater treatment systems where high nitrate concentration in the final effluent is a potential mitigating factor. Identification of the heterotrophic bacteria active in the denitrification process was attempted. This study presents a first report on the development and operation of the I-HRAP and has been followed by a technical-scale pilot plant evaluation of the process in the tertiary treatment of domestic wastewaters.
- Full Text:
- Date Issued: 2002
Thermophilic lignin degrading enzymes from actinomycetes for biotechnological applications
- Authors: Mhlanga, Chido Yvonne Lois
- Date: 2002 , 2013-05-16
- Subjects: Actinomycetales -- Biotechnology , Lignin
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4085 , http://hdl.handle.net/10962/d1007628 , Actinomycetales -- Biotechnology , Lignin
- Description: Phenolic residues which accumulate in the environment as a result of agro-industrial practices has resulted in the need to find and use Eco-Friendly techniques, rather than the traditional methods of burning or burying this kind of waste. Bioremediation and bioconversion are attractive alternatives using whole cell or enzyme-based systems. The aims of this project were to isolate and uses thermophilic Actinomycetes, which produce thermo-tolerant oxidoreductase enzymes, which can be used to bioconvert a model industrial phenolic waste commonly genersated in the wine-making industry of South Africa. Current research in bioconversion and bioremediation focuses on mesophilic microbes in that their enzymes can catalyse reactions at higher temperatures without affecting its activity and lower contamination levels. Three novel Actinomycete isolates were isolated (RU-A0l , RU-A03 and RU-A06) from a compost site and characterized using a combination of conventional identification techniques and 16S rDNA methodology to identity the three isolates. All three isolates belong to the Streptomyces clade. In addition, five known Actinomycetes were selected from an internation culture collection and also screened for oxidoreductase activity in comparision to the three novel isolates. Although the five isolates were selected based on their ability to produce oxidoreductase enzymes, unexpectedly, no activity was detected. Screening assays for peroxidase, polyphenol oxidase and laccase on RU-AO 1, RU-A03 and RU-A06, showed that all three isolated produced peroxidases and peroxidases but no laccase. Substrate specificity studies revealed that the most suitable substrates to determine peroxidase and polyphenol oxidase activity on these isolates were catechol for polyphenol oxidase, 2,4-dichlorophenol for peroxidases and veratryl alcohol for lignin peroxidases. Previous studies have indicated that peroxidases and polyphenol oxidases are produced in Actinomycetes during the primary stage of growth. This was the case with RU-AOI , RU-A03 and RU-A06. Growth rates were higher that other Actinomycetes, with maxImum biomass being reached at 36 hours for the isolates RU-AOI and RU-A06 and 48 hours for isolate RUA03. pH studies showed that the three isolates were adaptable and could grow over a broad pH range. Catabolism studies of phenolic model compounds showed that the three isolates were capable of catabolizing the model phenolic compounds within a period of 24 hours. Further studies were carried out to determine the effect of these microbes and their enzymes in whole cell and enzyme-based systems on a model phenolic waste, graoe waste consisting of compressed grape skins, pips and stalks. Whole cell studies showed that the isolates were capable of bioconverting the waste at a maximum concentration of 30% grape waste (vol:vol). Peroxidase and polyphenol oxidase activity increased indicating induction of these enzymes in the presence of phenolic compounds, with a maximum increase of up to 15.9 fold increase in extracellular lignin peroxidase activity in RU-AO1. HPLC and phenolic determination assays indicated that bioconversion of the phenolic grape waste had occurred in the presence of the three isolates. Attempts were made to isolate and identify a peroxidase or phenol oxidase gene from one the isolates. As bacteria, Actinomycetes are amendable to gene manipulation making them suitable candidates for methods such as site directed evolution in comparison to fungi. Two clones were selected for sequencing based on positive activity results when assayed for peroxidase activity. However the resultant sequences did not identify a functional gene sequence. Southern Blotting was then carried out to determine the nature of the peroxidase gene. Previous studies have been focused on the catalase-peroxidase gene (CalC gene) found Actinomycetes and other bacteria. A probe was developed from the CalC gene. No hybridization occurred with any of the enzyme restricted DNA from the three isolates. The implications of these results are that the peroxidase genets in the three isolates are in fact lignin peroxidase in nature. This project has the potential in the bioconversion of phenolic wastes and is the first description of the use of thermophilic Actinomycetes in the bioconversion of an industrial phenolic waste.
- Full Text:
- Date Issued: 2002
- Authors: Mhlanga, Chido Yvonne Lois
- Date: 2002 , 2013-05-16
- Subjects: Actinomycetales -- Biotechnology , Lignin
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4085 , http://hdl.handle.net/10962/d1007628 , Actinomycetales -- Biotechnology , Lignin
- Description: Phenolic residues which accumulate in the environment as a result of agro-industrial practices has resulted in the need to find and use Eco-Friendly techniques, rather than the traditional methods of burning or burying this kind of waste. Bioremediation and bioconversion are attractive alternatives using whole cell or enzyme-based systems. The aims of this project were to isolate and uses thermophilic Actinomycetes, which produce thermo-tolerant oxidoreductase enzymes, which can be used to bioconvert a model industrial phenolic waste commonly genersated in the wine-making industry of South Africa. Current research in bioconversion and bioremediation focuses on mesophilic microbes in that their enzymes can catalyse reactions at higher temperatures without affecting its activity and lower contamination levels. Three novel Actinomycete isolates were isolated (RU-A0l , RU-A03 and RU-A06) from a compost site and characterized using a combination of conventional identification techniques and 16S rDNA methodology to identity the three isolates. All three isolates belong to the Streptomyces clade. In addition, five known Actinomycetes were selected from an internation culture collection and also screened for oxidoreductase activity in comparision to the three novel isolates. Although the five isolates were selected based on their ability to produce oxidoreductase enzymes, unexpectedly, no activity was detected. Screening assays for peroxidase, polyphenol oxidase and laccase on RU-AO 1, RU-A03 and RU-A06, showed that all three isolated produced peroxidases and peroxidases but no laccase. Substrate specificity studies revealed that the most suitable substrates to determine peroxidase and polyphenol oxidase activity on these isolates were catechol for polyphenol oxidase, 2,4-dichlorophenol for peroxidases and veratryl alcohol for lignin peroxidases. Previous studies have indicated that peroxidases and polyphenol oxidases are produced in Actinomycetes during the primary stage of growth. This was the case with RU-AOI , RU-A03 and RU-A06. Growth rates were higher that other Actinomycetes, with maxImum biomass being reached at 36 hours for the isolates RU-AOI and RU-A06 and 48 hours for isolate RUA03. pH studies showed that the three isolates were adaptable and could grow over a broad pH range. Catabolism studies of phenolic model compounds showed that the three isolates were capable of catabolizing the model phenolic compounds within a period of 24 hours. Further studies were carried out to determine the effect of these microbes and their enzymes in whole cell and enzyme-based systems on a model phenolic waste, graoe waste consisting of compressed grape skins, pips and stalks. Whole cell studies showed that the isolates were capable of bioconverting the waste at a maximum concentration of 30% grape waste (vol:vol). Peroxidase and polyphenol oxidase activity increased indicating induction of these enzymes in the presence of phenolic compounds, with a maximum increase of up to 15.9 fold increase in extracellular lignin peroxidase activity in RU-AO1. HPLC and phenolic determination assays indicated that bioconversion of the phenolic grape waste had occurred in the presence of the three isolates. Attempts were made to isolate and identify a peroxidase or phenol oxidase gene from one the isolates. As bacteria, Actinomycetes are amendable to gene manipulation making them suitable candidates for methods such as site directed evolution in comparison to fungi. Two clones were selected for sequencing based on positive activity results when assayed for peroxidase activity. However the resultant sequences did not identify a functional gene sequence. Southern Blotting was then carried out to determine the nature of the peroxidase gene. Previous studies have been focused on the catalase-peroxidase gene (CalC gene) found Actinomycetes and other bacteria. A probe was developed from the CalC gene. No hybridization occurred with any of the enzyme restricted DNA from the three isolates. The implications of these results are that the peroxidase genets in the three isolates are in fact lignin peroxidase in nature. This project has the potential in the bioconversion of phenolic wastes and is the first description of the use of thermophilic Actinomycetes in the bioconversion of an industrial phenolic waste.
- Full Text:
- Date Issued: 2002