A preliminary investigation into the use of biomarkers and a fish community index to assess estuarine health in selected Eastern Cape estuaries
- Authors: Richardson, Naomi
- Date: 2008
- Subjects: Fish communities -- South Africa -- Eastern Cape Estuarine fishes -- South Africa -- Eastern Cape Estuarine health -- South Africa -- Eastern Cape Estuaries -- South Africa -- Eastern Cape -- Management Coastal zone management -- South Africa -- Eastern Cape Environmental monitoring -- South Africa -- Eastern Cape Estuarine ecology -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:5348 , http://hdl.handle.net/10962/d1007480
- Description: The aims of this study were to determine the potential use of biomarkers at multiple levels of biological organisation together with a fish community bioindicator to assess the estuarine health status of three temporarily open/closed estuaries. The estuaries investigated were the East Kleinemonde (EK), Old Woman's (OW) and Mtana (MTN), all of which are situated in the Eastern Cape Province. Three biomarkers, the acetylcholinesterase (AChE) assay, lipid peroxidation (LPx) assay, liver histopathology and a condition factor were used to determine sub-organism health and one bioindicator, the Estuarine Fish Community Index (EFCI), was used as a bioindicator of community health. The estuarine-dependent marine species Rhabdosargus holubi was selected as an indicator species for the sub-organism level analyses. The results from the community analyses indicated that the EK and OW estuaries were in 'good' condition, while the MTN was found to be in 'moderate' condition. Histological analyses revealed that R. holubi from all three estuaries showed signs of pathological changes to the liver, with the fish from the MTN eliciting the highest occurrence of these changes. The LPx assay found that R. holubi from both the OW and MTN showed signs of oxidative damage in the liver tissue, but those from the EK did not appear to be affected. The AChE assay showed that only the fish from the OW had been affected by anticholinesterase compounds. A laboratory study was undertaken using R. holubi as a positive control for the AChE and LPx assay. The fish were exposed to 3 μg/L chlorpyrifos, a known cholinesterase inhibitor, for six hours and their tissues were examined for changes to LPx levels and AChE activities. AChE activity was significantly inhibited (Mann Whitney U test, Z = 3.65, n = 38, P < 0.001) by the exposure, but LPx levels were not significantly affected. A composite index incorporating the biomarkers at different biological levels of organisation was developed. The index was designed to assist managers and scientists to determine whether the ichthyofauna of a system was being affected by environmental stressors and what management interventions could be undertaken to ameliorate the water quality in an estuary. The index was applied to the three estuaries investigated during the present study and both the OW and MTN were assessed to be in need of immediate management intervention. The fish in the OW were found to be stressed at all the sub-organism levels measured and the reason for this was hypothesised to be as a result of golf course activities in this adjacent estuary. A number of management actions are proposed to reduce the sub-organism stress observed in the fish from the OW. The livers of fish from the MTN were shown to be under stress; however the causative agent of this stress was unknown because there is no formal development in the MTN catchment. However, a possible contaminant source is proposed and management interventions to alleviate the stress on the biota of the MTN are suggested. The EK does not require immediate management intervention, however, continuous routine monitoring is recommended to ensure that conditions do not deteriorate. Shortcomings of the index were outlined and a number of suggestions were made in terms of other measures of biological health which could be incorporated into the index.
- Full Text:
- Date Issued: 2008
- Authors: Richardson, Naomi
- Date: 2008
- Subjects: Fish communities -- South Africa -- Eastern Cape Estuarine fishes -- South Africa -- Eastern Cape Estuarine health -- South Africa -- Eastern Cape Estuaries -- South Africa -- Eastern Cape -- Management Coastal zone management -- South Africa -- Eastern Cape Environmental monitoring -- South Africa -- Eastern Cape Estuarine ecology -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:5348 , http://hdl.handle.net/10962/d1007480
- Description: The aims of this study were to determine the potential use of biomarkers at multiple levels of biological organisation together with a fish community bioindicator to assess the estuarine health status of three temporarily open/closed estuaries. The estuaries investigated were the East Kleinemonde (EK), Old Woman's (OW) and Mtana (MTN), all of which are situated in the Eastern Cape Province. Three biomarkers, the acetylcholinesterase (AChE) assay, lipid peroxidation (LPx) assay, liver histopathology and a condition factor were used to determine sub-organism health and one bioindicator, the Estuarine Fish Community Index (EFCI), was used as a bioindicator of community health. The estuarine-dependent marine species Rhabdosargus holubi was selected as an indicator species for the sub-organism level analyses. The results from the community analyses indicated that the EK and OW estuaries were in 'good' condition, while the MTN was found to be in 'moderate' condition. Histological analyses revealed that R. holubi from all three estuaries showed signs of pathological changes to the liver, with the fish from the MTN eliciting the highest occurrence of these changes. The LPx assay found that R. holubi from both the OW and MTN showed signs of oxidative damage in the liver tissue, but those from the EK did not appear to be affected. The AChE assay showed that only the fish from the OW had been affected by anticholinesterase compounds. A laboratory study was undertaken using R. holubi as a positive control for the AChE and LPx assay. The fish were exposed to 3 μg/L chlorpyrifos, a known cholinesterase inhibitor, for six hours and their tissues were examined for changes to LPx levels and AChE activities. AChE activity was significantly inhibited (Mann Whitney U test, Z = 3.65, n = 38, P < 0.001) by the exposure, but LPx levels were not significantly affected. A composite index incorporating the biomarkers at different biological levels of organisation was developed. The index was designed to assist managers and scientists to determine whether the ichthyofauna of a system was being affected by environmental stressors and what management interventions could be undertaken to ameliorate the water quality in an estuary. The index was applied to the three estuaries investigated during the present study and both the OW and MTN were assessed to be in need of immediate management intervention. The fish in the OW were found to be stressed at all the sub-organism levels measured and the reason for this was hypothesised to be as a result of golf course activities in this adjacent estuary. A number of management actions are proposed to reduce the sub-organism stress observed in the fish from the OW. The livers of fish from the MTN were shown to be under stress; however the causative agent of this stress was unknown because there is no formal development in the MTN catchment. However, a possible contaminant source is proposed and management interventions to alleviate the stress on the biota of the MTN are suggested. The EK does not require immediate management intervention, however, continuous routine monitoring is recommended to ensure that conditions do not deteriorate. Shortcomings of the index were outlined and a number of suggestions were made in terms of other measures of biological health which could be incorporated into the index.
- Full Text:
- Date Issued: 2008
Isolation, expression and purification of the hydantoin hydrolysing enzymes of agrobacterium tumefaciens
- Authors: Clark, Sally-Ann
- Date: 2003
- Subjects: Agrobacterium tumefaciens , Amino acids Hydantoin Enzymes
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4140 , http://hdl.handle.net/10962/d1016233
- Description: The production of enantiomerically pure amino acids is of industrial importance as they are used in the synthesis of a number of pharmaceuticals, insecticides and herbicides and biologically active peptides and hormones. A number of microorganisms have been identified which possess hydantoin hydrolysing enzymes that stereoselectively convert racemic hydantoins into anantiomerically pure amino acids. Consequently these microorganisms and their enzymes are sought after as biocatalysts for the production of amino acids. The isolation of novel hydantoin hydrolising enzymes with unique or improved biocatalytic characteristics is of importance for the development of potential biocatalysts to be used in the production of enantiomerically pure amino acids. The genes encoding an N-carbamoyl-amino acid amidohydrolase, an enzyme involved in the hydrolysis of hydantoin, was isolated by screening a genomic DNA library of Agrobacterium tumefacience RU-AE01. Nucleotide sequence analysis of the region upstream of this gene revealed a fragment of a gene encoding the hydantoinase enzyme. I this study, a DNA probe consisting of the gene encoding the N-carbamoyl amino acid amidohydrolase, on a large enough fragment of the genomic DNA library which would allow for the simultaneous isolation the hydantoinase gene located upstream. Recombinant expression of the genes encoding hydantoin hydrolysing enzymes has been used to facilitate the production and purification of these enzymes for their use as biocatalysts. Two genes (ncaR1 and ncaR2) encoding different N-carbamoyl-amino acid amidohydrolases with distinct nucleotide and deduced amino acid sequences were isolated from the genome of A, tumefaciens RU-OR. In this study, the heterologous expression of ncaR1 and ncaR2 was explored. Investigation into the optimisation of the heterologous expression of ncaR1 showed that reducing the growth temperature of the recombinant E. coli producing NcaR1 resulted in a two-fold increase in N-carbamoyl-amino acid amidohydrolase activity and solubility. Furthermore, NcaR1 was produced with a C-terminal 6xHis tag, but NcaR1-6xHis did not possess N-carbamoyl amino acid amidohydrolase activity. Furthermore, purification of NcaR-6xHis under native conditions using affinity chromatography performed, and used for the production of antibodies.
- Full Text:
- Date Issued: 2003
- Authors: Clark, Sally-Ann
- Date: 2003
- Subjects: Agrobacterium tumefaciens , Amino acids Hydantoin Enzymes
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4140 , http://hdl.handle.net/10962/d1016233
- Description: The production of enantiomerically pure amino acids is of industrial importance as they are used in the synthesis of a number of pharmaceuticals, insecticides and herbicides and biologically active peptides and hormones. A number of microorganisms have been identified which possess hydantoin hydrolysing enzymes that stereoselectively convert racemic hydantoins into anantiomerically pure amino acids. Consequently these microorganisms and their enzymes are sought after as biocatalysts for the production of amino acids. The isolation of novel hydantoin hydrolising enzymes with unique or improved biocatalytic characteristics is of importance for the development of potential biocatalysts to be used in the production of enantiomerically pure amino acids. The genes encoding an N-carbamoyl-amino acid amidohydrolase, an enzyme involved in the hydrolysis of hydantoin, was isolated by screening a genomic DNA library of Agrobacterium tumefacience RU-AE01. Nucleotide sequence analysis of the region upstream of this gene revealed a fragment of a gene encoding the hydantoinase enzyme. I this study, a DNA probe consisting of the gene encoding the N-carbamoyl amino acid amidohydrolase, on a large enough fragment of the genomic DNA library which would allow for the simultaneous isolation the hydantoinase gene located upstream. Recombinant expression of the genes encoding hydantoin hydrolysing enzymes has been used to facilitate the production and purification of these enzymes for their use as biocatalysts. Two genes (ncaR1 and ncaR2) encoding different N-carbamoyl-amino acid amidohydrolases with distinct nucleotide and deduced amino acid sequences were isolated from the genome of A, tumefaciens RU-OR. In this study, the heterologous expression of ncaR1 and ncaR2 was explored. Investigation into the optimisation of the heterologous expression of ncaR1 showed that reducing the growth temperature of the recombinant E. coli producing NcaR1 resulted in a two-fold increase in N-carbamoyl-amino acid amidohydrolase activity and solubility. Furthermore, NcaR1 was produced with a C-terminal 6xHis tag, but NcaR1-6xHis did not possess N-carbamoyl amino acid amidohydrolase activity. Furthermore, purification of NcaR-6xHis under native conditions using affinity chromatography performed, and used for the production of antibodies.
- Full Text:
- Date Issued: 2003
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