Assessment of phytochemical constituents, antioxidant properties and cytotoxicity potentials of Leonutis leonurus and Boophone disticha extracts used in the management of Alzheimer’s disease in Hogsback, Eastern Cape, South Africa
- Authors: Tonisi, Sipho
- Date: 2019
- Subjects: Medicinal plants Alzheimer's disease
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: http://hdl.handle.net/10353/14726 , vital:40067
- Description: In this present study, the phytochemical compounds, antioxidant activities, and cytotoxicity potentials of solvent extracts (polar solvents) of Leonotis leonurus (L. leonurus) and Boophone disitcha (B. disticha) commonly used in the management of Alzheimer’s diseases in Hogsback, in the Eastern Cape, South Africa were investigated. The phytochemical compounds were determined by standard protocols of Folin–Ciocalteu method, aluminium chloride colorimetric method as well as Mayer’s and Wagner’s method. The antioxidant capabilities were assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydrogen peroxide, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and nitric oxide radical scavenging assays. The majority of extracts from both plant species exhibited free radical scavenging potential for oxidants they were tested against viz; DPPH, hydrogen peroxide, ABTS and nitric oxide. They further displayed good reducing potential against ferric iron. The results have shown a concentration-dependent dose of inhibition/scavenging activity (in DPPH, hydrogen peroxide, ABTS and nitric oxide assays) and reducing ability (in ferric reducing assay). However, leaf extracts of L. leonurus displayed ineffective scavenging activity against nitric oxide whereas extracts from L. leonurus flower and B. disticha bulb had higher scavenging potential against the production of nitric oxide radical. The GC-MS profiles of the plants extracts revealed the presence of numerous bioactive compounds that are known to possess excellent anti-inflammatory and most importantly antioxidant properties. Another interesting observation of GC-MS analysis data was the presence of compounds reported to inhibit Acetycholinesterase (AChE), which is a key enzyme targeted in the treatment of Alzheimers’ Disease (AD). The cytotoxicy tests for these extracts were carried out in order to assess the potentential detrimental effects on human health using Alamar Blue assay. It was noted that an increase in the extract concentration of both plant species resulted in a noticeable decline in cell population. These extracts were then declared xix to be quite toxic against HeLa (human cervix adenocarcinoma cells) cells as the cell viability in some extracts was quite low. Hence, toxicity testing of the studied plants’ extracts against normal cells is highly imperative in the future study. The results from this present study scientifically validated the use of the studied plants in folkoric medicine for the management of Alzheimer’s diseases.
- Full Text:
- Date Issued: 2019
- Authors: Tonisi, Sipho
- Date: 2019
- Subjects: Medicinal plants Alzheimer's disease
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: http://hdl.handle.net/10353/14726 , vital:40067
- Description: In this present study, the phytochemical compounds, antioxidant activities, and cytotoxicity potentials of solvent extracts (polar solvents) of Leonotis leonurus (L. leonurus) and Boophone disitcha (B. disticha) commonly used in the management of Alzheimer’s diseases in Hogsback, in the Eastern Cape, South Africa were investigated. The phytochemical compounds were determined by standard protocols of Folin–Ciocalteu method, aluminium chloride colorimetric method as well as Mayer’s and Wagner’s method. The antioxidant capabilities were assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), hydrogen peroxide, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and nitric oxide radical scavenging assays. The majority of extracts from both plant species exhibited free radical scavenging potential for oxidants they were tested against viz; DPPH, hydrogen peroxide, ABTS and nitric oxide. They further displayed good reducing potential against ferric iron. The results have shown a concentration-dependent dose of inhibition/scavenging activity (in DPPH, hydrogen peroxide, ABTS and nitric oxide assays) and reducing ability (in ferric reducing assay). However, leaf extracts of L. leonurus displayed ineffective scavenging activity against nitric oxide whereas extracts from L. leonurus flower and B. disticha bulb had higher scavenging potential against the production of nitric oxide radical. The GC-MS profiles of the plants extracts revealed the presence of numerous bioactive compounds that are known to possess excellent anti-inflammatory and most importantly antioxidant properties. Another interesting observation of GC-MS analysis data was the presence of compounds reported to inhibit Acetycholinesterase (AChE), which is a key enzyme targeted in the treatment of Alzheimers’ Disease (AD). The cytotoxicy tests for these extracts were carried out in order to assess the potentential detrimental effects on human health using Alamar Blue assay. It was noted that an increase in the extract concentration of both plant species resulted in a noticeable decline in cell population. These extracts were then declared xix to be quite toxic against HeLa (human cervix adenocarcinoma cells) cells as the cell viability in some extracts was quite low. Hence, toxicity testing of the studied plants’ extracts against normal cells is highly imperative in the future study. The results from this present study scientifically validated the use of the studied plants in folkoric medicine for the management of Alzheimer’s diseases.
- Full Text:
- Date Issued: 2019
Isolation and characterisation of lignocellulose degrading bacteria from Tyume River in the Eastern Cape Province, South Africa
- Authors: Tembisa, Papiyana Ayavuya
- Date: 2015
- Subjects: Lignocellulose -- Biodegradation -- South Africa -- Eastern Cape , Bacillus (Bacteria) -- South Africa -- Eastern Cape , Waterborne infection -- South Africa -- Eastern Cape , Bacteriophages -- South Africa -- Eastern Cape , Sediments (Geology) -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11299 , http://hdl.handle.net/10353/d1021293 , Lignocellulose -- Biodegradation -- South Africa -- Eastern Cape , Bacillus (Bacteria) -- South Africa -- Eastern Cape , Waterborne infection -- South Africa -- Eastern Cape , Bacteriophages -- South Africa -- Eastern Cape , Sediments (Geology) -- South Africa -- Eastern Cape
- Description: This study focuses on the isolation and characterization of bacteria from lignocellulosic biomass obtained from the sediments of the Tyume River in Alice, Eastern Cape and to determine those bacterial isolates with good potential for modification and decomposition of lignocellulosic biomass for industrial application. Several bacterial isolates were recovered and screened for ability to degrade various lignocellulosic materials. Nine of the isolates were positive for lignocellulolytic activity. Four isolates were cellulase positive and six were xylanase positive. Moreover, one isolate (SB1) was positive for both xylanase and cellulase activities and showed the best hydrolysis zone on solid media. This isolate was then chosen as the best and identified molecularly. The 16S rDNA sequence analysis indicated that SB1 was a Bacillus cereus species. Factors affecting the cellulose and xylanase enzyme production by the organisms were studied. The organisms produced the enzymes maximally at earlier hours of incubation (12-30 hr) and optimally at acidic pH (3-5) and at moderate temperatures (35-45ºC). SB1 appears to hold promise in the decomposition of lignocellulosic wastes.
- Full Text:
- Date Issued: 2015
- Authors: Tembisa, Papiyana Ayavuya
- Date: 2015
- Subjects: Lignocellulose -- Biodegradation -- South Africa -- Eastern Cape , Bacillus (Bacteria) -- South Africa -- Eastern Cape , Waterborne infection -- South Africa -- Eastern Cape , Bacteriophages -- South Africa -- Eastern Cape , Sediments (Geology) -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11299 , http://hdl.handle.net/10353/d1021293 , Lignocellulose -- Biodegradation -- South Africa -- Eastern Cape , Bacillus (Bacteria) -- South Africa -- Eastern Cape , Waterborne infection -- South Africa -- Eastern Cape , Bacteriophages -- South Africa -- Eastern Cape , Sediments (Geology) -- South Africa -- Eastern Cape
- Description: This study focuses on the isolation and characterization of bacteria from lignocellulosic biomass obtained from the sediments of the Tyume River in Alice, Eastern Cape and to determine those bacterial isolates with good potential for modification and decomposition of lignocellulosic biomass for industrial application. Several bacterial isolates were recovered and screened for ability to degrade various lignocellulosic materials. Nine of the isolates were positive for lignocellulolytic activity. Four isolates were cellulase positive and six were xylanase positive. Moreover, one isolate (SB1) was positive for both xylanase and cellulase activities and showed the best hydrolysis zone on solid media. This isolate was then chosen as the best and identified molecularly. The 16S rDNA sequence analysis indicated that SB1 was a Bacillus cereus species. Factors affecting the cellulose and xylanase enzyme production by the organisms were studied. The organisms produced the enzymes maximally at earlier hours of incubation (12-30 hr) and optimally at acidic pH (3-5) and at moderate temperatures (35-45ºC). SB1 appears to hold promise in the decomposition of lignocellulosic wastes.
- Full Text:
- Date Issued: 2015
Assessment of antibacterial potentials of Garcinia Kola seed extracts and their interactions with antibiotics
- Authors: Sibanda, Thulani
- Date: 2007
- Subjects: Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11242 , http://hdl.handle.net/10353/71 , Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants
- Description: The antibacterial potency of the extracts of the seed of Garcinia kola (bitter kola) was investigated in this study against a panel of referenced, environmental and clinical bacterial strains. The killing rates of the active extract as well as their potential for combination antibacterial therapy with standard antibiotics were also elucidated using standard procedures. The aqueous and acetone extracts of the seed were screened for activity against 27 bacterial isolates. The aqueous extract exhibited activity mainly against Gram positive organisms with Minimum inhibitory concentration (MIC) values ranging from 5 mgml-1 – 20 mgml-1, while the acetone extract showed activity against both Gram negative and Gram positive organisms with MIC values ranging from 10 mgml-1 - 0.156 mgml-1. The acetone extract also showed rapid bactericidal activity against Staphylococcus aureus ATCC 6538 with a 3.097 Log10 reduction in counts within 4 hours at 0.3125 mgml-1 and a 1.582 Log10 reduction against Proteus vulgaris CSIR 0030 at 5 mgml-1 after 1 hour. In addition, the aqueous, methanol and acetone extracts of the seeds also exhibited activity against four clinical strains of Staphylococcus isolated from wound sepsis specimens. The MIC values for the aqueous extract were 10 mgml-1 for all the isolates while the acetone and methanol extracts had lower values ranging from 0.3125 - 0.625 mgml-1. The acetone extract was strongly bactericidal against Staphylococcus aureus OKOH3 resulting in a 2.70 Log10 reduction in counts at 1.25 mgml-1 within 4 hours of exposure and a complete elimination of the organism after 8 hours. The bactericidal vi activity of the same extract against Staphylococcus aureus OKOH1 was weak, achieving only a 2.92 Log10 reduction in counts at 1.25 mgml-1 (4× MIC) in 24 hours. In the test for interactions between the acetone extract of the seeds and antibiotics, synergistic interactions were observed largely against Gram positive organisms using the FIC indices, (indices of 0.52 - 0.875) with combinations against Gram negatives yielding largely antagonistic interactions (indices of 2.0 to 5.0). Synergy (≥ 1000 times or ≥ 3 Log10 potentiation of the bactericidal activity) against both Gram negative and Gram positive organisms was detected by time kill assays mainly involving the antibiotics tetracycline, chloramphenicol, amoxycillin and penicillin G. Combinations involving erythromycin and ciprofloxacin consistently gave antagonistic or indifferent interactions. We conclude that the acetone extract of Garcinia kola seeds possess strong bactericidal activities against both Gram positive and Gram negative organisms and can be therapeutically useful in the treatment of bacterial infections including the problematic staphylococcal wound infections. In addition, the acetone extract can be a potential source of broad spectrum resistance modifying compounds that can potentially improve the performance of antibiotics in the treatment of drug resistant infections.
- Full Text:
- Date Issued: 2007
- Authors: Sibanda, Thulani
- Date: 2007
- Subjects: Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11242 , http://hdl.handle.net/10353/71 , Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants
- Description: The antibacterial potency of the extracts of the seed of Garcinia kola (bitter kola) was investigated in this study against a panel of referenced, environmental and clinical bacterial strains. The killing rates of the active extract as well as their potential for combination antibacterial therapy with standard antibiotics were also elucidated using standard procedures. The aqueous and acetone extracts of the seed were screened for activity against 27 bacterial isolates. The aqueous extract exhibited activity mainly against Gram positive organisms with Minimum inhibitory concentration (MIC) values ranging from 5 mgml-1 – 20 mgml-1, while the acetone extract showed activity against both Gram negative and Gram positive organisms with MIC values ranging from 10 mgml-1 - 0.156 mgml-1. The acetone extract also showed rapid bactericidal activity against Staphylococcus aureus ATCC 6538 with a 3.097 Log10 reduction in counts within 4 hours at 0.3125 mgml-1 and a 1.582 Log10 reduction against Proteus vulgaris CSIR 0030 at 5 mgml-1 after 1 hour. In addition, the aqueous, methanol and acetone extracts of the seeds also exhibited activity against four clinical strains of Staphylococcus isolated from wound sepsis specimens. The MIC values for the aqueous extract were 10 mgml-1 for all the isolates while the acetone and methanol extracts had lower values ranging from 0.3125 - 0.625 mgml-1. The acetone extract was strongly bactericidal against Staphylococcus aureus OKOH3 resulting in a 2.70 Log10 reduction in counts at 1.25 mgml-1 within 4 hours of exposure and a complete elimination of the organism after 8 hours. The bactericidal vi activity of the same extract against Staphylococcus aureus OKOH1 was weak, achieving only a 2.92 Log10 reduction in counts at 1.25 mgml-1 (4× MIC) in 24 hours. In the test for interactions between the acetone extract of the seeds and antibiotics, synergistic interactions were observed largely against Gram positive organisms using the FIC indices, (indices of 0.52 - 0.875) with combinations against Gram negatives yielding largely antagonistic interactions (indices of 2.0 to 5.0). Synergy (≥ 1000 times or ≥ 3 Log10 potentiation of the bactericidal activity) against both Gram negative and Gram positive organisms was detected by time kill assays mainly involving the antibiotics tetracycline, chloramphenicol, amoxycillin and penicillin G. Combinations involving erythromycin and ciprofloxacin consistently gave antagonistic or indifferent interactions. We conclude that the acetone extract of Garcinia kola seeds possess strong bactericidal activities against both Gram positive and Gram negative organisms and can be therapeutically useful in the treatment of bacterial infections including the problematic staphylococcal wound infections. In addition, the acetone extract can be a potential source of broad spectrum resistance modifying compounds that can potentially improve the performance of antibiotics in the treatment of drug resistant infections.
- Full Text:
- Date Issued: 2007
Prevalence of pathogenic Escherichia coli strains in the final effluents of four wastewater treatment plants in the Eastern Cape Province of South Africa
- Authors: Seti, Nozuko Zukiswa
- Date: 2014
- Subjects: Escherichia coli -- South Africa -- Eastern Cape , Bacterial diseases -- Pathogenesis , Sewage -- Purification -- South Africa -- Eastern Cape , Sewage disposal plants -- South Africa -- Eastern Cape , Whole effluent toxicity testing -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11286 , http://hdl.handle.net/10353/d1019808 , Escherichia coli -- South Africa -- Eastern Cape , Bacterial diseases -- Pathogenesis , Sewage -- Purification -- South Africa -- Eastern Cape , Sewage disposal plants -- South Africa -- Eastern Cape , Whole effluent toxicity testing -- South Africa -- Eastern Cape
- Description: Water is an essential need that stimulates health and well being. Increase in population size and urbanization negatively affect water resources due to high demands of effluent outputs. Wastewater is an important reservoir for Escherichia coli and can present significant acute toxicity if released into receiving water sources without being adequately treated. E. coli is used as indicator organism for the detection of faecal contamination. These strains have been considered to be one of the primary causes of diarrhoeal infections worldwide. The present study was conducted between September 2012 and June 2013 to assess the prevalence of pathogenic E. coli strains in the final effluents of four wastewater treatment plants in Chris Hani and Buffalo City Municipalities in the Eastern Cape Province of South Africa. Standard membrane filtration technique was used for bacteriological analysis and molecular based technique was used for identification of E. coli pathotypes. The results were recorded in colony forming units/100 ml. Faecal coliforms ranged between 0-9.6×10³ CFU/100 ml for the wwtp-Q and E. coli densities ranged between 0-8.4×10³ CFU/100ml. Faecal coliforms ranged between 4×10²-9.7×10³ CFU/100 ml for wwtp-M and E. coli densities ranged between 1.2×10¹-8.4×10³ CFU/100 ml. The wwtp-E showed to have bacterial counts of faecal coliforms ranging between 4.0×10³-8.2×10³ CFU/100 ml and E. coli densities ranging between 3.5×10¹-7.1×10³ CFU/100 ml. The WWTP-K in this study was only assessed for the presence of E. coli. Faecal coliforms were assessed by the other members of the group. This plant showed to have E. coli densities ranging between 0-7.5×10²CFU/100 ml. A total of 200 presumptive E. coli isolates were subjected to screening by conventional PCR in which (29%) of the wwtp-M isolates were positively identified as E. coli, (16%) of the wwtp-K, (22%) of the wwtp-Q and (34%) of the wwtp-E isolates were positively confirmed as E. coli. A total of 100 randomly selected E. coli isolates were characterised into different pathotypes. (16%) of positive isolates were detected as EPEC and 11% were detected as UPEC strains. There was no detection for the ETEC strains. Antibiotic susceptibility patterns of E. coli strains showed high levels of resistance to Penicillin G, Erythromycin, Tetracycline and Sulfamethoxazole. High levels of Susceptibility were observed in antibiotics such as Chloramphenicol, Amoxicillin and Tetracycline. The results of this study reveal that the plants were above the recommended Standard limit of zero CFU/100 ml for effluents meant to be discharge into receiving water sources. This study reveals inadequacy of the plants studied to produce effluents of acceptable quality.
- Full Text:
- Date Issued: 2014
- Authors: Seti, Nozuko Zukiswa
- Date: 2014
- Subjects: Escherichia coli -- South Africa -- Eastern Cape , Bacterial diseases -- Pathogenesis , Sewage -- Purification -- South Africa -- Eastern Cape , Sewage disposal plants -- South Africa -- Eastern Cape , Whole effluent toxicity testing -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11286 , http://hdl.handle.net/10353/d1019808 , Escherichia coli -- South Africa -- Eastern Cape , Bacterial diseases -- Pathogenesis , Sewage -- Purification -- South Africa -- Eastern Cape , Sewage disposal plants -- South Africa -- Eastern Cape , Whole effluent toxicity testing -- South Africa -- Eastern Cape
- Description: Water is an essential need that stimulates health and well being. Increase in population size and urbanization negatively affect water resources due to high demands of effluent outputs. Wastewater is an important reservoir for Escherichia coli and can present significant acute toxicity if released into receiving water sources without being adequately treated. E. coli is used as indicator organism for the detection of faecal contamination. These strains have been considered to be one of the primary causes of diarrhoeal infections worldwide. The present study was conducted between September 2012 and June 2013 to assess the prevalence of pathogenic E. coli strains in the final effluents of four wastewater treatment plants in Chris Hani and Buffalo City Municipalities in the Eastern Cape Province of South Africa. Standard membrane filtration technique was used for bacteriological analysis and molecular based technique was used for identification of E. coli pathotypes. The results were recorded in colony forming units/100 ml. Faecal coliforms ranged between 0-9.6×10³ CFU/100 ml for the wwtp-Q and E. coli densities ranged between 0-8.4×10³ CFU/100ml. Faecal coliforms ranged between 4×10²-9.7×10³ CFU/100 ml for wwtp-M and E. coli densities ranged between 1.2×10¹-8.4×10³ CFU/100 ml. The wwtp-E showed to have bacterial counts of faecal coliforms ranging between 4.0×10³-8.2×10³ CFU/100 ml and E. coli densities ranging between 3.5×10¹-7.1×10³ CFU/100 ml. The WWTP-K in this study was only assessed for the presence of E. coli. Faecal coliforms were assessed by the other members of the group. This plant showed to have E. coli densities ranging between 0-7.5×10²CFU/100 ml. A total of 200 presumptive E. coli isolates were subjected to screening by conventional PCR in which (29%) of the wwtp-M isolates were positively identified as E. coli, (16%) of the wwtp-K, (22%) of the wwtp-Q and (34%) of the wwtp-E isolates were positively confirmed as E. coli. A total of 100 randomly selected E. coli isolates were characterised into different pathotypes. (16%) of positive isolates were detected as EPEC and 11% were detected as UPEC strains. There was no detection for the ETEC strains. Antibiotic susceptibility patterns of E. coli strains showed high levels of resistance to Penicillin G, Erythromycin, Tetracycline and Sulfamethoxazole. High levels of Susceptibility were observed in antibiotics such as Chloramphenicol, Amoxicillin and Tetracycline. The results of this study reveal that the plants were above the recommended Standard limit of zero CFU/100 ml for effluents meant to be discharge into receiving water sources. This study reveals inadequacy of the plants studied to produce effluents of acceptable quality.
- Full Text:
- Date Issued: 2014
Phytochemical analysis and bioactivity of Garcinia Kola (Heckel) seeds on selected bacterial pathogens
- Seanego, Christinah Tshephisho
- Authors: Seanego, Christinah Tshephisho
- Date: 2012
- Subjects: Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants , Microbial sensitivity tests , Streptococcal infections , Streptococcus , Staphylococcus aureus infections , Salmonella typhimurium , Traditional medicine
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11259 , http://hdl.handle.net/10353/420 , Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants , Microbial sensitivity tests , Streptococcal infections , Streptococcus , Staphylococcus aureus infections , Salmonella typhimurium , Traditional medicine
- Description: Garcinia kola is one of the plants used in folklore remedies for the treatment of microbial infections. Bacterial resistance to commonly used antibiotics has necessitated the search for newer and alternative compounds for the treatment of drug resistant microbial infections. This study focuses on the bioactivity of G. kola seeds on Streptococcus pyogenes (ATCC 49399), Staphylococcus aureus (NCTC 6571), Plesiomonas Shigelloides (ATCC 51903) and Salmonella typhimurium (ATCC 13311), organisms which can cause illnesses from mild to severe with potentially fatal outcomes. The crude ethyl acetate, ethanol, methanol, acetone and aqueous extracts were screened by agar-well diffusion method and the activities of the extract were further determined by Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) assays. The inhibition zones ranged from 0 - 24 mm, while MIC and MBC of the extract ranged between 0.04 - 1.25 mg/mL and 0.081 - 2.5 mg/mL respectively. Chloroform/ Ethyl Acetate/ Formic acid (CEF) solvent system separated more active compounds followed by Ethyl Acetate/ Methanol/ Water (EMW) and Benzene/ Ethanol/ Ammonium Hydroxide (BEA). The extracts were fractionated by Thin Layer Chromatography (TLC). Bioautography was used to assess the activity of the possible classes of compounds present in the more active extracts. Column chromatography was used to purify the active compounds from the mixture while Gas Chromatography-Mass Spectrometry (GC-MS) was used to identify the phyto components of the fractions. The MIC of the fractions ranged between 0.0006 - 2.5 mg/mL. CEF 3 (F3), CEF 11 (F11) and CEF 12 (F12) revealed the presence of high levels fatty acids Linoleic acid, 1, 2-Benzenedicarboxylic acid and 2, 3-Dihydro-3, 5-dihydroxy-6-methyl, respectively. The results obtained from this study justify the use of this plant in traditional medicine and provide leads which could be further exploited for the development of new and potent antimicrobials.
- Full Text:
- Date Issued: 2012
- Authors: Seanego, Christinah Tshephisho
- Date: 2012
- Subjects: Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants , Microbial sensitivity tests , Streptococcal infections , Streptococcus , Staphylococcus aureus infections , Salmonella typhimurium , Traditional medicine
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11259 , http://hdl.handle.net/10353/420 , Drug resistance in microorganisms , Garcinia , Antibiotics , Medicinal plants , Microbial sensitivity tests , Streptococcal infections , Streptococcus , Staphylococcus aureus infections , Salmonella typhimurium , Traditional medicine
- Description: Garcinia kola is one of the plants used in folklore remedies for the treatment of microbial infections. Bacterial resistance to commonly used antibiotics has necessitated the search for newer and alternative compounds for the treatment of drug resistant microbial infections. This study focuses on the bioactivity of G. kola seeds on Streptococcus pyogenes (ATCC 49399), Staphylococcus aureus (NCTC 6571), Plesiomonas Shigelloides (ATCC 51903) and Salmonella typhimurium (ATCC 13311), organisms which can cause illnesses from mild to severe with potentially fatal outcomes. The crude ethyl acetate, ethanol, methanol, acetone and aqueous extracts were screened by agar-well diffusion method and the activities of the extract were further determined by Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) assays. The inhibition zones ranged from 0 - 24 mm, while MIC and MBC of the extract ranged between 0.04 - 1.25 mg/mL and 0.081 - 2.5 mg/mL respectively. Chloroform/ Ethyl Acetate/ Formic acid (CEF) solvent system separated more active compounds followed by Ethyl Acetate/ Methanol/ Water (EMW) and Benzene/ Ethanol/ Ammonium Hydroxide (BEA). The extracts were fractionated by Thin Layer Chromatography (TLC). Bioautography was used to assess the activity of the possible classes of compounds present in the more active extracts. Column chromatography was used to purify the active compounds from the mixture while Gas Chromatography-Mass Spectrometry (GC-MS) was used to identify the phyto components of the fractions. The MIC of the fractions ranged between 0.0006 - 2.5 mg/mL. CEF 3 (F3), CEF 11 (F11) and CEF 12 (F12) revealed the presence of high levels fatty acids Linoleic acid, 1, 2-Benzenedicarboxylic acid and 2, 3-Dihydro-3, 5-dihydroxy-6-methyl, respectively. The results obtained from this study justify the use of this plant in traditional medicine and provide leads which could be further exploited for the development of new and potent antimicrobials.
- Full Text:
- Date Issued: 2012
Effects of genetically modified maize (MON810) and its residues on the functional diversity of microorganisms in two South African soils
- Authors: Puta, Usanda
- Date: 2011
- Subjects: Genetically modified foods -- South Africa , Transgenic plants -- South Africa , Crops -- Genetic engineering -- South Africa , Soil microbiology -- South Africa , Microorganisms , Microbial ecology , Rhizosphere -- Microbiology , Vesicular-arbuscular mycorrhizas , Corn -- South Africa
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11250 , http://hdl.handle.net/10353/419 , Genetically modified foods -- South Africa , Transgenic plants -- South Africa , Crops -- Genetic engineering -- South Africa , Soil microbiology -- South Africa , Microorganisms , Microbial ecology , Rhizosphere -- Microbiology , Vesicular-arbuscular mycorrhizas , Corn -- South Africa
- Description: Genetically modified (GM) crops are commercially cultivated worldwide but there are concerns on their possible negative impacts on soil biodiversity. A glasshouse study was conducted to determine effects of Bt maize residues on soil microbial diversity. Residues of Bt maize (PAN 6Q-308B) and non-Bt maize (PAN 6Q-121) were incorporated into the soil and corresponding maize seeds planted. The treatments were replicated three times. Fertilizer and water application were similar for both treatments. Rhizosphere and bulk soil was destructively sampled from each treatment and analyzed for microbial community level physiological profiles using Biolog plates with 31 different carbon substrates. Absorbance in the Biolog plates was recorded after 72 h of incubation at 20oC. Arbuscular mycorrhizal fungi spore counts were also determined. Field studies were conducted at the University of Free State and University of Fort Hare Research Farms to determine the effects of growing Bt maize on soil microbial diversity. One Bt maize cultivar (PAN6Q-308B) and non-Bt maize (PAN6Q-121) were grown in a paired experiment at University of Free State farm, while two Bt maize (DKC61-25B and PAN6Q-321B) and their near-isogenic non-Bt maize lines (DKC61-24 and PAN6777) were grown in a randomized complete block design with three replicates. Fertilization, weed control and water application, were similar for both Bt maize cultivars and their non-Bt maize counterparts. Rhizosphere soil samples were collected by uprooting whole plants and collecting the soil attached to the roots. The samples were analysed for microbial diversity and for arbuscular mycorrhizae fungal spore counts. Principal component analysis showed that soil microbial diversity was affected more by sampling time whereas genetic modification had minimal effects. Presence of residues also increased the diversity of microorganisms. Mycorrhizal fungal spores were not affected by the presence of Bt maize residues. Growing Bt maize had no effect on the soil microbial diversity in the rhizosphere.
- Full Text:
- Date Issued: 2011
- Authors: Puta, Usanda
- Date: 2011
- Subjects: Genetically modified foods -- South Africa , Transgenic plants -- South Africa , Crops -- Genetic engineering -- South Africa , Soil microbiology -- South Africa , Microorganisms , Microbial ecology , Rhizosphere -- Microbiology , Vesicular-arbuscular mycorrhizas , Corn -- South Africa
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11250 , http://hdl.handle.net/10353/419 , Genetically modified foods -- South Africa , Transgenic plants -- South Africa , Crops -- Genetic engineering -- South Africa , Soil microbiology -- South Africa , Microorganisms , Microbial ecology , Rhizosphere -- Microbiology , Vesicular-arbuscular mycorrhizas , Corn -- South Africa
- Description: Genetically modified (GM) crops are commercially cultivated worldwide but there are concerns on their possible negative impacts on soil biodiversity. A glasshouse study was conducted to determine effects of Bt maize residues on soil microbial diversity. Residues of Bt maize (PAN 6Q-308B) and non-Bt maize (PAN 6Q-121) were incorporated into the soil and corresponding maize seeds planted. The treatments were replicated three times. Fertilizer and water application were similar for both treatments. Rhizosphere and bulk soil was destructively sampled from each treatment and analyzed for microbial community level physiological profiles using Biolog plates with 31 different carbon substrates. Absorbance in the Biolog plates was recorded after 72 h of incubation at 20oC. Arbuscular mycorrhizal fungi spore counts were also determined. Field studies were conducted at the University of Free State and University of Fort Hare Research Farms to determine the effects of growing Bt maize on soil microbial diversity. One Bt maize cultivar (PAN6Q-308B) and non-Bt maize (PAN6Q-121) were grown in a paired experiment at University of Free State farm, while two Bt maize (DKC61-25B and PAN6Q-321B) and their near-isogenic non-Bt maize lines (DKC61-24 and PAN6777) were grown in a randomized complete block design with three replicates. Fertilization, weed control and water application, were similar for both Bt maize cultivars and their non-Bt maize counterparts. Rhizosphere soil samples were collected by uprooting whole plants and collecting the soil attached to the roots. The samples were analysed for microbial diversity and for arbuscular mycorrhizae fungal spore counts. Principal component analysis showed that soil microbial diversity was affected more by sampling time whereas genetic modification had minimal effects. Presence of residues also increased the diversity of microorganisms. Mycorrhizal fungal spores were not affected by the presence of Bt maize residues. Growing Bt maize had no effect on the soil microbial diversity in the rhizosphere.
- Full Text:
- Date Issued: 2011
In-vitro anti-vibrio activities of crude extracts of Garcinia Kola seeds
- Authors: Penduka, Dambudzo
- Date: 2011
- Subjects: Microbial sensitivity tests , Drug resistance in microorganisms , Antibiotics , Garcinia , Medicinal plants
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11256 , http://hdl.handle.net/10353/405 , Microbial sensitivity tests , Drug resistance in microorganisms , Antibiotics , Garcinia , Medicinal plants
- Description: The n-Hexane, dichloromethane, methanol and aqueous crude extracts of Garcinia kola (Heckel) seeds were screened for their anti-Vibrio activities against 50 Vibrio bacteria isolated from wastewater final effluents. The 50 isolates consisted of different Vibrio species namely V. fluvialis (14), V. vulnificus (12), V. parahaemolyticus (12), V. metschnikovii (3) and 9 others unidentified to the specie level. The n-Hexane, dichloromethane and methanol extracts had activities against 16 (32 percent) of the Vibrio isolates, while the aqueous extracts had activities against 12 (24 percent) all at a screening concentration of 10 mg/ml. The minimum inhibitory concentrations (MICs) were 0.313-0.625 mg/ml, 0.313-0.625 mg/ml, 0.313-2.5 mg/ml and 10 mg/ml for n-Hexane, dichloromethane, methanol and aqueous extracts respectively. Rate of kill studies were carried out against three different Vibrio species namely V. vulnificus (AL042), V. parahaemolyticus (AL049) and V. fluvialis ( AL040) using the n-Hexane, dichloromethane and methanol extracts at 1× to 4 × MICs and 2 hour exposure. About 96.3 percent, 82.2 percent, and 78.1 percent (V. fluvialis AL040); 92.6 percent, 87.8 percent and 68.9 percent (V. parahaemolyticus AL049); and 91.6 percent, 64.4 percent, 60 percent (V. vulnificus AL042) of the bacteria were killed by the crude n-Hexane, dichloromethane and methanol extracts respectively after 2 hour exposure time at 4× MIC. The patterns of activity were bacteriostatic, with the n-Hexane extracts being most effective in activity. We conclude that the Garcinia kola seeds have promise in the treatment and management of infections caused by Vibrio species.
- Full Text:
- Date Issued: 2011
- Authors: Penduka, Dambudzo
- Date: 2011
- Subjects: Microbial sensitivity tests , Drug resistance in microorganisms , Antibiotics , Garcinia , Medicinal plants
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11256 , http://hdl.handle.net/10353/405 , Microbial sensitivity tests , Drug resistance in microorganisms , Antibiotics , Garcinia , Medicinal plants
- Description: The n-Hexane, dichloromethane, methanol and aqueous crude extracts of Garcinia kola (Heckel) seeds were screened for their anti-Vibrio activities against 50 Vibrio bacteria isolated from wastewater final effluents. The 50 isolates consisted of different Vibrio species namely V. fluvialis (14), V. vulnificus (12), V. parahaemolyticus (12), V. metschnikovii (3) and 9 others unidentified to the specie level. The n-Hexane, dichloromethane and methanol extracts had activities against 16 (32 percent) of the Vibrio isolates, while the aqueous extracts had activities against 12 (24 percent) all at a screening concentration of 10 mg/ml. The minimum inhibitory concentrations (MICs) were 0.313-0.625 mg/ml, 0.313-0.625 mg/ml, 0.313-2.5 mg/ml and 10 mg/ml for n-Hexane, dichloromethane, methanol and aqueous extracts respectively. Rate of kill studies were carried out against three different Vibrio species namely V. vulnificus (AL042), V. parahaemolyticus (AL049) and V. fluvialis ( AL040) using the n-Hexane, dichloromethane and methanol extracts at 1× to 4 × MICs and 2 hour exposure. About 96.3 percent, 82.2 percent, and 78.1 percent (V. fluvialis AL040); 92.6 percent, 87.8 percent and 68.9 percent (V. parahaemolyticus AL049); and 91.6 percent, 64.4 percent, 60 percent (V. vulnificus AL042) of the bacteria were killed by the crude n-Hexane, dichloromethane and methanol extracts respectively after 2 hour exposure time at 4× MIC. The patterns of activity were bacteriostatic, with the n-Hexane extracts being most effective in activity. We conclude that the Garcinia kola seeds have promise in the treatment and management of infections caused by Vibrio species.
- Full Text:
- Date Issued: 2011
Characterization of some virulence and antibiotic resistance genes of Staphylococcus aureus isolated from cases of Bovine Mastitis in Nkonkobe Municipality, Eastern Cape Province, RSA
- Authors: Pekana, Abongile
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11293 , http://hdl.handle.net/10353/d1021133
- Description: Staphylococcus aureus is one of the predominant causative agents of mastitis disease in dairy herds. Mastitis disease has a negative impact in the economic losses in the dairy sector across the globe. The aim of this study is to detect some of the virulence genes in the S. aureus isolated from 400 milk samples of subclinical and clinical mastitis dairy cows in Fort Hare dairy farm and Middle Drift dairy farm in Alice in the Eastern Cape province of South Africa. In addition antibiotic resistance pattern and antibiotic resistance genes were investigated. Gram-staining, oxidase test, catalase test and API Staph kit were preliminary biochemical tests used for the identification of S. aureus isolates. The MALDI-TOF-MS was also used for further identification. Polymerase chain reaction was performed of genes encoding antibiotic resistance as well as clumping (clfA), coagulase (coa) gene, toxic shock syndrome (tsst), exfoliative toxin A and B (eta and etb), and the gene segment encoding the immunoglobulin G binding region and X region of protein gene spa. A total of 20 (5%) S. aureus strains obtained from 400 milk samples from the two farms were subjected to 16 antibiotics for antibiotic susceptibility testing. In Middle Drift dairy farm 11 (5.5%) isolates were obtained from 200 samples and 9 (4.5%) isolates were obtained in Fort Hare dairy farm from 200 samples. A large percent of the isolates were resistant to penicillin G (60%), followed by trimethoprim (60%) and tetracycline (60%), trimethoprim-sulfamethaxazole (55%), telithroprim (55%) and doxycycline (45%). Most of the isolates were sensitive to several (50-85%) antibiotics. Of the twenty isolates tested 12 samples contained the penicillin antibiotic resistance gene (blaZ gene), 8 samples contained at least one aminoglycoside-modifying enzyme gene (AME gene); the (aac(6’)/aph(2’’) gene and no amplification occurred for aph(3’)-IIIa and ant(4’)-Ia) genes. In the case of the tetracycline antibiotic resistance gene (tetK and tetM), 2 samples contained tetM and a single sample contained tetK gene. No amplification was observed for the erythromycin antibiotic resistance genes (ermA, ermB, ermC, Mef and msrA). All the samples tested were negative for the expression of toxic syndrome gene (tsst), etb, and Immunoglobulin G binding region. However, amplification of the clumping factor was observed in 7 (35%) isolates of S. aureus, exfoliative toxin (eta) expressed 4(20%) isolates; coagulase gene (coa) yielded six DNA bands of six differences sizes from 16 (80%) isolates. A total of four different bands size were expressed for the spa X region from 12 (60%) isolates. The data obtained in this study suggests that poor hygienic practices and inadequate management practices are responsible for the increase in Staphylococcus aureus isolation. The high resistance of S. aureus to antibiotics and the distribution of virulence genes contribute in bovine mastitis in these farms may cause health problems in the community consuming raw milk purchased from these farms.
- Full Text:
- Date Issued: 2015
- Authors: Pekana, Abongile
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11293 , http://hdl.handle.net/10353/d1021133
- Description: Staphylococcus aureus is one of the predominant causative agents of mastitis disease in dairy herds. Mastitis disease has a negative impact in the economic losses in the dairy sector across the globe. The aim of this study is to detect some of the virulence genes in the S. aureus isolated from 400 milk samples of subclinical and clinical mastitis dairy cows in Fort Hare dairy farm and Middle Drift dairy farm in Alice in the Eastern Cape province of South Africa. In addition antibiotic resistance pattern and antibiotic resistance genes were investigated. Gram-staining, oxidase test, catalase test and API Staph kit were preliminary biochemical tests used for the identification of S. aureus isolates. The MALDI-TOF-MS was also used for further identification. Polymerase chain reaction was performed of genes encoding antibiotic resistance as well as clumping (clfA), coagulase (coa) gene, toxic shock syndrome (tsst), exfoliative toxin A and B (eta and etb), and the gene segment encoding the immunoglobulin G binding region and X region of protein gene spa. A total of 20 (5%) S. aureus strains obtained from 400 milk samples from the two farms were subjected to 16 antibiotics for antibiotic susceptibility testing. In Middle Drift dairy farm 11 (5.5%) isolates were obtained from 200 samples and 9 (4.5%) isolates were obtained in Fort Hare dairy farm from 200 samples. A large percent of the isolates were resistant to penicillin G (60%), followed by trimethoprim (60%) and tetracycline (60%), trimethoprim-sulfamethaxazole (55%), telithroprim (55%) and doxycycline (45%). Most of the isolates were sensitive to several (50-85%) antibiotics. Of the twenty isolates tested 12 samples contained the penicillin antibiotic resistance gene (blaZ gene), 8 samples contained at least one aminoglycoside-modifying enzyme gene (AME gene); the (aac(6’)/aph(2’’) gene and no amplification occurred for aph(3’)-IIIa and ant(4’)-Ia) genes. In the case of the tetracycline antibiotic resistance gene (tetK and tetM), 2 samples contained tetM and a single sample contained tetK gene. No amplification was observed for the erythromycin antibiotic resistance genes (ermA, ermB, ermC, Mef and msrA). All the samples tested were negative for the expression of toxic syndrome gene (tsst), etb, and Immunoglobulin G binding region. However, amplification of the clumping factor was observed in 7 (35%) isolates of S. aureus, exfoliative toxin (eta) expressed 4(20%) isolates; coagulase gene (coa) yielded six DNA bands of six differences sizes from 16 (80%) isolates. A total of four different bands size were expressed for the spa X region from 12 (60%) isolates. The data obtained in this study suggests that poor hygienic practices and inadequate management practices are responsible for the increase in Staphylococcus aureus isolation. The high resistance of S. aureus to antibiotics and the distribution of virulence genes contribute in bovine mastitis in these farms may cause health problems in the community consuming raw milk purchased from these farms.
- Full Text:
- Date Issued: 2015
Studies on the antimicrobial, antioxidant and antiproliferative potential of the ethyl acetate extract and compounds of Peltophorum africanum
- Authors: Okeleye, Benjamin Ifeoluwa
- Date: 2014
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11282 , http://hdl.handle.net/10353/d1016165
- Description: Cells are constantly exposed to a variety of oxidizing agents, some of which are necessary for life. Oxidants produced in excess can cause an imbalance, leading to oxidative stress, especially in chronic bacterial, viral, and parasitic infections. This can result to damage of biomolecules such as lipids, proteins, and DNA, hence, an increased risk for cancer. Plants have a long history of use in the treatment of cancer. Plant secondary metabolites have proved to be an excellent reservoir of new medical compounds. Fruits, vegetables, and whole grains contain a wide variety of antioxidant phytochemicals, such as phenolics and carotenoids, and may help protect cellular systems from oxidative damage and also may lower the risk of chronic diseases. Peltophorum africanum, a member of the family Fabaceae (Sond) is also known as the African weeping wattle and is used in traditional medicine in South Africa. This study investigated the antimicrobial, antioxidant and antiproliferative potential of the ethyl acetate extract and compounds of Peltophorum africanum in order to validate its pharmacological use. The study assessed the in vitro antimicrobial activity of ethyl acetate extract (EAE) of Peltophorum africanum stem bark and its fractions by the agar well and macrodilution methods. The toxicity on a normal human liver cell (Chang liver cell) and antiproliferation of human breast (MCF-7), colon (HT-29) and cervical (HeLa) cancer cell lines were determined using the CellTiter-Blue cell viability assay and the mechanism of action delineated using the Nucleic Acid and Protein Purification Nucleospin® Tissue Kit, Scanning Electron Microscopy (SEM), Propidium iodide (PI) and Acridine orange (AO) double-staining techniques, the Cleaved Caspase 3 (Asp 175) Alexa Fluor® 488 Antibody and the Coulter® DNA PrepTM Reagents Kit. Purification and identification of the compounds from EAE and fractions as well as the morphological alteration of bacteria, yeast and cancer cells were determined using thin layer chromatography, infrared spectra fingerprint and GC-MS analysis, micro-dilution and scanning electron microscopy with energy-dispersive X-ray analysis. In vitro antioxidant activity of EAE was determined by means of radical scavenging and ferric reducing power analysis using 2, 2-diphenyl-1-picrylhydrazyl (DPPH), 2, 2`-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) kit, hydrogen peroxide (H2O2), iron (iii) chloride (Fe3+) and nitric oxide (NO). To assess the likely effects of secondary metabolites on the activities observed; total proanthocyanidins, phenolics, flavonols, and flavonoids were determined using standard phytochemical methods. Data were analyzed by one way analysis of variance (ANOVA; SPSS Version 17.0, 2011), regression analysis (MINITAB, version 12 for windows), probit analysis test (software NCSS, 2007) and GraphPad Prism4 software package. The p-values < 0.05 were considered significant. Marked activity of the extract was observed against Plesiomonas shigelloides ATCC 51903, with MIC and MLC values of 0.15625 and 0.3125mg/mL, respectively. The extract was both bactericidal (MICindex ≤ 2) and bacteriostatic/fungistatic (MICindex > 2) in activity. Lethal dose at 50 (LD50) showed 82.64 ± 1.40 degree of toxicity at 24 hrs, and 95 percentile of cell death dose activity ranged from log 3.12 ± 0.01 to 4.59 ± 0.03. The activity of the eight fractions tested ranged from 1.0 ± 0.5 to 3.7 ± 1.6 mg/mL (IC50) and from 2.1 ± 0.8 to 6.25 ± 0 mg/mL (IC90) (Chapter 3). Due to the effect of compounds present in the crude extract and fractions, the P. aeruginosa treated with EAE had a reduction of sodium from 5.55 % (untreated) - 1.50 %. For C. albicans, pottasium was reduced from 4.16 % (untreated) - 0.76 % (T1). Remarkable morphological alterations were observed including deformation of the germ tubes and perforation of the cell wall (Chapter 4). Extract scavenging activity of 88.73± 6.69 % (25 μg mL-1), 53.93±1.09 % (25 μg mL-1) were recorded for H2O2 and NO respectively with proanthocyanidins (92.18±4.68 mg/g) occurring more (p < 0.05) in the extract compared to all other phenolics compounds (Chapter 5). Significant reduction in cell viability of the cells was noted as the MCF-7 cells were reduced from 100 - 54.33±1.84 % after 72 hrs of treatment with 5 μg/mL of EAE (P. value < 0.05). TEt10 was cytotoxic against human normal cells (chang liver cell) at EC50 of 37 μg/mL and 74 μg/mL after 24 and 48 h of treatment respectively. Marked antiproliferative activity of 13.2 μg/mL (EC50) was observed when HeLa cells were treated for 48 h. Internucleosomal DNA of MCF-7, HT-29 and HeLa cells randomly fragmented into an uninterrupted spectrum of sizes, complemented by the intercalation of nucleic acid-specific fluorochromes by PI and AO spotting two phases of apoptosis; early (EA) and late (LA) apoptosis. Distinctive ultramorphological changes observed include; cell shrinkage, membrane blebbing, and typical cell induced death. The study also recorded 705.102 ± 28.56 % TEt10 caspase-3 activity compared to curcumin 592.857 ± 165.76 % (positive control) and untreated (negative control; 100 ± 15.81 %) cells. Percentage HeLa cell with Sub-G1 DNA phase increased from 0.13 ± 0.06 % (negative control) to 13.8 ± 3.04 % compared to curcumin (8.17 ± 2.20 %) after treatment with TEt10. The compounds identified in the fractions including Colchicine, N-(trifluoroacetyl)methyl-N-deacetyl-, Lupeol and .gamma.-Sitosterol may be responsible for the induction of apoptosis observed and could be further studied in vivo as a potential template for new anticancer treatment (Chapter 6 & 7).
- Full Text:
- Date Issued: 2014
- Authors: Okeleye, Benjamin Ifeoluwa
- Date: 2014
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11282 , http://hdl.handle.net/10353/d1016165
- Description: Cells are constantly exposed to a variety of oxidizing agents, some of which are necessary for life. Oxidants produced in excess can cause an imbalance, leading to oxidative stress, especially in chronic bacterial, viral, and parasitic infections. This can result to damage of biomolecules such as lipids, proteins, and DNA, hence, an increased risk for cancer. Plants have a long history of use in the treatment of cancer. Plant secondary metabolites have proved to be an excellent reservoir of new medical compounds. Fruits, vegetables, and whole grains contain a wide variety of antioxidant phytochemicals, such as phenolics and carotenoids, and may help protect cellular systems from oxidative damage and also may lower the risk of chronic diseases. Peltophorum africanum, a member of the family Fabaceae (Sond) is also known as the African weeping wattle and is used in traditional medicine in South Africa. This study investigated the antimicrobial, antioxidant and antiproliferative potential of the ethyl acetate extract and compounds of Peltophorum africanum in order to validate its pharmacological use. The study assessed the in vitro antimicrobial activity of ethyl acetate extract (EAE) of Peltophorum africanum stem bark and its fractions by the agar well and macrodilution methods. The toxicity on a normal human liver cell (Chang liver cell) and antiproliferation of human breast (MCF-7), colon (HT-29) and cervical (HeLa) cancer cell lines were determined using the CellTiter-Blue cell viability assay and the mechanism of action delineated using the Nucleic Acid and Protein Purification Nucleospin® Tissue Kit, Scanning Electron Microscopy (SEM), Propidium iodide (PI) and Acridine orange (AO) double-staining techniques, the Cleaved Caspase 3 (Asp 175) Alexa Fluor® 488 Antibody and the Coulter® DNA PrepTM Reagents Kit. Purification and identification of the compounds from EAE and fractions as well as the morphological alteration of bacteria, yeast and cancer cells were determined using thin layer chromatography, infrared spectra fingerprint and GC-MS analysis, micro-dilution and scanning electron microscopy with energy-dispersive X-ray analysis. In vitro antioxidant activity of EAE was determined by means of radical scavenging and ferric reducing power analysis using 2, 2-diphenyl-1-picrylhydrazyl (DPPH), 2, 2`-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) kit, hydrogen peroxide (H2O2), iron (iii) chloride (Fe3+) and nitric oxide (NO). To assess the likely effects of secondary metabolites on the activities observed; total proanthocyanidins, phenolics, flavonols, and flavonoids were determined using standard phytochemical methods. Data were analyzed by one way analysis of variance (ANOVA; SPSS Version 17.0, 2011), regression analysis (MINITAB, version 12 for windows), probit analysis test (software NCSS, 2007) and GraphPad Prism4 software package. The p-values < 0.05 were considered significant. Marked activity of the extract was observed against Plesiomonas shigelloides ATCC 51903, with MIC and MLC values of 0.15625 and 0.3125mg/mL, respectively. The extract was both bactericidal (MICindex ≤ 2) and bacteriostatic/fungistatic (MICindex > 2) in activity. Lethal dose at 50 (LD50) showed 82.64 ± 1.40 degree of toxicity at 24 hrs, and 95 percentile of cell death dose activity ranged from log 3.12 ± 0.01 to 4.59 ± 0.03. The activity of the eight fractions tested ranged from 1.0 ± 0.5 to 3.7 ± 1.6 mg/mL (IC50) and from 2.1 ± 0.8 to 6.25 ± 0 mg/mL (IC90) (Chapter 3). Due to the effect of compounds present in the crude extract and fractions, the P. aeruginosa treated with EAE had a reduction of sodium from 5.55 % (untreated) - 1.50 %. For C. albicans, pottasium was reduced from 4.16 % (untreated) - 0.76 % (T1). Remarkable morphological alterations were observed including deformation of the germ tubes and perforation of the cell wall (Chapter 4). Extract scavenging activity of 88.73± 6.69 % (25 μg mL-1), 53.93±1.09 % (25 μg mL-1) were recorded for H2O2 and NO respectively with proanthocyanidins (92.18±4.68 mg/g) occurring more (p < 0.05) in the extract compared to all other phenolics compounds (Chapter 5). Significant reduction in cell viability of the cells was noted as the MCF-7 cells were reduced from 100 - 54.33±1.84 % after 72 hrs of treatment with 5 μg/mL of EAE (P. value < 0.05). TEt10 was cytotoxic against human normal cells (chang liver cell) at EC50 of 37 μg/mL and 74 μg/mL after 24 and 48 h of treatment respectively. Marked antiproliferative activity of 13.2 μg/mL (EC50) was observed when HeLa cells were treated for 48 h. Internucleosomal DNA of MCF-7, HT-29 and HeLa cells randomly fragmented into an uninterrupted spectrum of sizes, complemented by the intercalation of nucleic acid-specific fluorochromes by PI and AO spotting two phases of apoptosis; early (EA) and late (LA) apoptosis. Distinctive ultramorphological changes observed include; cell shrinkage, membrane blebbing, and typical cell induced death. The study also recorded 705.102 ± 28.56 % TEt10 caspase-3 activity compared to curcumin 592.857 ± 165.76 % (positive control) and untreated (negative control; 100 ± 15.81 %) cells. Percentage HeLa cell with Sub-G1 DNA phase increased from 0.13 ± 0.06 % (negative control) to 13.8 ± 3.04 % compared to curcumin (8.17 ± 2.20 %) after treatment with TEt10. The compounds identified in the fractions including Colchicine, N-(trifluoroacetyl)methyl-N-deacetyl-, Lupeol and .gamma.-Sitosterol may be responsible for the induction of apoptosis observed and could be further studied in vivo as a potential template for new anticancer treatment (Chapter 6 & 7).
- Full Text:
- Date Issued: 2014
In vitro activity of bioactive compounds of selected South African medicinal plants on clinical isolates of Helicobacter pylori
- Authors: Okeleye, Benjamin Ifeoluwa
- Date: 2011
- Subjects: Helicobacter pylori , Microbial sensitivity tests , Traditional medicine -- South Africa , Gram-negative bacterial infections
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11255 , http://hdl.handle.net/10353/310 , Helicobacter pylori , Microbial sensitivity tests , Traditional medicine -- South Africa , Gram-negative bacterial infections
- Description: The stem bark of Peltophorum africanum and Bridelia micrantha are used in South Africa traditional medicine for treatment of intestinal parasites, relieve problems and human immunodeficiency virus/ acquired immune deficiency syndrome (HIV/AIDS). The growing problem of antibiotic resistance by Helicobacter pylori the major etiological agent in gastritis, gastric cancer, peptic and gastric ulcer demands the search for novel compounds from plant based sources. This study was aimed to determine the antimicrobial activity of five solvent (ethylacetate, acetone, ethanol, methanol and water) extracts of the stem bark of P. africanum and B. micrantha on clinical strains of H. pylori in a bid to identify potential sources of cheap starting materials for the synthesis of new drugs. H. pylori strains were isolated from patients presenting with gastric related morbidities at the Livingstone Hospital, Port Elizabeth for endoscopy and confirmed following standard microbiology procedures. The plant extracts including clarithromycin were tested against 31 clinical strains of H. pylori by the agar well diffusion method. The most potent extract was evaluated by the microdilution method to determine the Minimum Inhibitory Concentration (MIC50&90), followed by the rate of kill. Preliminary phytochemical analysis was carried out. The one way ANOVA test was used to statistically analyse the results. All the extracts demonstrated anti-H. pylori activity with zone diameters of inhibition that ranged from 0 to 23 mm for the extracts and 0 to 35 mm for clarithromycin. Marked susceptibility (100%) was recorded for the ethyl acetate extract of P. africanum (P. afr. EA) and the acetone extract of B. micrantha (B. mic. A), which were statistically significant (P < 0.05) compared to all other extracts and clarithromycin. For B. micrantha ethyl acetate extract, 93.5 percent susceptibility was observed while for the control iv antibiotic, clarithromycin it was 58.1 percent. The MIC50 ranged from 0.0048 to 0.313 mg/mL for P. afr. EA, and from 0.0048 to 0.156 mg/mL for B. mic. EA; MIC90 ranged from 0.156 mg/mL to 0.625 mg/mL and 0.0048 to 2.5 mg/mL for P. afr. EA and B. mic. EA respectively. There was a significant statistical difference observed in potency of both P. afr. EA and B. mic. A compared to the two antibiotics (P < 0.05). One hundred percent killing by P. afr EA was observed at 0.05 mg/mL (½ x MIC) and 0.2 mg/mL (2 x MIC) in 66 h for strain PE466C and PE252C respectively. For B. mic. EA, 100 percent killing effect of both strains (PE430C and PE369C) was observed at 0.1 mg/mL (2 x MIC) in 66 h. Qualitative phytochemical analysis confirmed the presence of alkaloids, flavonoids, steroids, tannins and saponins in the ethyl acetate extracts of both plants, which could be a potential template of lead molecule for the design of new anti- Helicobacter pylori therapies.
- Full Text:
- Date Issued: 2011
- Authors: Okeleye, Benjamin Ifeoluwa
- Date: 2011
- Subjects: Helicobacter pylori , Microbial sensitivity tests , Traditional medicine -- South Africa , Gram-negative bacterial infections
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11255 , http://hdl.handle.net/10353/310 , Helicobacter pylori , Microbial sensitivity tests , Traditional medicine -- South Africa , Gram-negative bacterial infections
- Description: The stem bark of Peltophorum africanum and Bridelia micrantha are used in South Africa traditional medicine for treatment of intestinal parasites, relieve problems and human immunodeficiency virus/ acquired immune deficiency syndrome (HIV/AIDS). The growing problem of antibiotic resistance by Helicobacter pylori the major etiological agent in gastritis, gastric cancer, peptic and gastric ulcer demands the search for novel compounds from plant based sources. This study was aimed to determine the antimicrobial activity of five solvent (ethylacetate, acetone, ethanol, methanol and water) extracts of the stem bark of P. africanum and B. micrantha on clinical strains of H. pylori in a bid to identify potential sources of cheap starting materials for the synthesis of new drugs. H. pylori strains were isolated from patients presenting with gastric related morbidities at the Livingstone Hospital, Port Elizabeth for endoscopy and confirmed following standard microbiology procedures. The plant extracts including clarithromycin were tested against 31 clinical strains of H. pylori by the agar well diffusion method. The most potent extract was evaluated by the microdilution method to determine the Minimum Inhibitory Concentration (MIC50&90), followed by the rate of kill. Preliminary phytochemical analysis was carried out. The one way ANOVA test was used to statistically analyse the results. All the extracts demonstrated anti-H. pylori activity with zone diameters of inhibition that ranged from 0 to 23 mm for the extracts and 0 to 35 mm for clarithromycin. Marked susceptibility (100%) was recorded for the ethyl acetate extract of P. africanum (P. afr. EA) and the acetone extract of B. micrantha (B. mic. A), which were statistically significant (P < 0.05) compared to all other extracts and clarithromycin. For B. micrantha ethyl acetate extract, 93.5 percent susceptibility was observed while for the control iv antibiotic, clarithromycin it was 58.1 percent. The MIC50 ranged from 0.0048 to 0.313 mg/mL for P. afr. EA, and from 0.0048 to 0.156 mg/mL for B. mic. EA; MIC90 ranged from 0.156 mg/mL to 0.625 mg/mL and 0.0048 to 2.5 mg/mL for P. afr. EA and B. mic. EA respectively. There was a significant statistical difference observed in potency of both P. afr. EA and B. mic. A compared to the two antibiotics (P < 0.05). One hundred percent killing by P. afr EA was observed at 0.05 mg/mL (½ x MIC) and 0.2 mg/mL (2 x MIC) in 66 h for strain PE466C and PE252C respectively. For B. mic. EA, 100 percent killing effect of both strains (PE430C and PE369C) was observed at 0.1 mg/mL (2 x MIC) in 66 h. Qualitative phytochemical analysis confirmed the presence of alkaloids, flavonoids, steroids, tannins and saponins in the ethyl acetate extracts of both plants, which could be a potential template of lead molecule for the design of new anti- Helicobacter pylori therapies.
- Full Text:
- Date Issued: 2011
Assessment of antibiotic production by some marine Streptomyces isolated from the Nahoon Beach
- Ogunmwonyi, Isoken Nekpen Henrietta
- Authors: Ogunmwonyi, Isoken Nekpen Henrietta
- Date: 2010
- Subjects: Streptomyces , Actinobacteria , Gram-positive bacteria , Antibiotics , Antibiotics -- Testing , Drug resistance in microorganisms
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11243 , http://hdl.handle.net/10353/264 , Streptomyces , Actinobacteria , Gram-positive bacteria , Antibiotics , Antibiotics -- Testing , Drug resistance in microorganisms
- Description: Rapidly emerging strains of bacteria resistant to most advanced antibiotics have become issues of very important public health concern. Research currently directed towards marine actinomycetes presents a vast potential for new compounds that could be able to safely and effectively target resistant species. In this regard, ten putative Streptomyces strains isolated from the Nahoon beach were selected and assessed for antibiotic production and activity against a wide range of bacteria including reference strains, environmental strain and clinical isolates. The ethyl acetate extracts of the putative Streptomyces isolates showed activities against at least 6 and up to 26 of the 32 test bacteria. Inhibition zones were found to range between 9-32 mm diameters at a concentration of 10 mg/ml. The minimum inhibitory concentrations (MICs) of the crude extracts ranged from 0.039 - 10 mg/ml and the least minimum bactericidal concentration (MBC) demonstrated was 0.625 mg/ml against a reference strain Staphylococcus aureus ATCC 6538. Time kill kinetics of all extracts revealed bacteristatic and bactericidal activities. Average Log reductions in viable cell counts for all the extracts ranged from 0.86 Log10 and 3.99 Log10 cfu/ml after 3 h interaction and 0.01 Log10 and 4.86 Log10 after 6 h interaction at MIC, 2 × MIC, 3 × MIC and 4 × MIC concentrations. Most of the extracts were speedily bactericidal at 3 × MIC and 4 × MIC resulting in over 50 % elimination of most of the test bacteria within 3 h and 6 h interaction. The partial characterization of the crude extracts by IR spectral analysis revealed possibility of terpenoid, long chain fatty acids and secondary amine derivatives compounds in the extracts. It is therefore recommended that further investigation should address the relationship between the structure of the active component of the extracts and the broad spectrum activity, as well as a rapid method for large scale production and purification and whether this group of antibiotics has any application in managing human infectious disease.
- Full Text:
- Date Issued: 2010
- Authors: Ogunmwonyi, Isoken Nekpen Henrietta
- Date: 2010
- Subjects: Streptomyces , Actinobacteria , Gram-positive bacteria , Antibiotics , Antibiotics -- Testing , Drug resistance in microorganisms
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11243 , http://hdl.handle.net/10353/264 , Streptomyces , Actinobacteria , Gram-positive bacteria , Antibiotics , Antibiotics -- Testing , Drug resistance in microorganisms
- Description: Rapidly emerging strains of bacteria resistant to most advanced antibiotics have become issues of very important public health concern. Research currently directed towards marine actinomycetes presents a vast potential for new compounds that could be able to safely and effectively target resistant species. In this regard, ten putative Streptomyces strains isolated from the Nahoon beach were selected and assessed for antibiotic production and activity against a wide range of bacteria including reference strains, environmental strain and clinical isolates. The ethyl acetate extracts of the putative Streptomyces isolates showed activities against at least 6 and up to 26 of the 32 test bacteria. Inhibition zones were found to range between 9-32 mm diameters at a concentration of 10 mg/ml. The minimum inhibitory concentrations (MICs) of the crude extracts ranged from 0.039 - 10 mg/ml and the least minimum bactericidal concentration (MBC) demonstrated was 0.625 mg/ml against a reference strain Staphylococcus aureus ATCC 6538. Time kill kinetics of all extracts revealed bacteristatic and bactericidal activities. Average Log reductions in viable cell counts for all the extracts ranged from 0.86 Log10 and 3.99 Log10 cfu/ml after 3 h interaction and 0.01 Log10 and 4.86 Log10 after 6 h interaction at MIC, 2 × MIC, 3 × MIC and 4 × MIC concentrations. Most of the extracts were speedily bactericidal at 3 × MIC and 4 × MIC resulting in over 50 % elimination of most of the test bacteria within 3 h and 6 h interaction. The partial characterization of the crude extracts by IR spectral analysis revealed possibility of terpenoid, long chain fatty acids and secondary amine derivatives compounds in the extracts. It is therefore recommended that further investigation should address the relationship between the structure of the active component of the extracts and the broad spectrum activity, as well as a rapid method for large scale production and purification and whether this group of antibiotics has any application in managing human infectious disease.
- Full Text:
- Date Issued: 2010
Phytochemical analysis and bioactivity of the stem bark of Combretum Molle on some selected bacterial pathogens
- Authors: Nyenje, Mirriam, E
- Date: 2011
- Subjects: Drug resistance in microorganisms , Materia medica, Vegetable , Antibiotics , Microbial sensitivity tests , Gram-negative bacterial infections
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11261 , http://hdl.handle.net/10353/391 , Drug resistance in microorganisms , Materia medica, Vegetable , Antibiotics , Microbial sensitivity tests , Gram-negative bacterial infections
- Description: Antimicrobial resistance is a worldwide problem that has deleterious long-term effects as the development of drug resistance outpaces the development of new drugs. Plants have been used for many generations for healing purposes, and screening of extracts of these plants has often yielded positive outcomes. This study was aimed at isolating and characterizing the major active antimicrobial compounds present in the stem bark of C. molle, in a bid to identify potential sources of cheap starting materials for the synthesis of new drugs. Various solvents (hexane, ethyl acetate, dichloromethane, acetone, ethanol and methanol) were used for extraction. The agar well diffusion technique was used to screen for antimicrobial activity of C. molle extracts against Streptococcus pyogenes ATCC 49399, Plesiomonas shigelloides ATCC 51903, Pseudomonas aeruginosa ATCC 15442, Helicobacter pylori ATCC 43526 and Helicobacter pylori 252C (clinical isolate); minimum inhibition concentration (MIC) of the most active extracts was determined by the broth dilution method. Fractionation of acetone extract was done by thin layer chromatography (TLC) and bioautography to determine the compounds present and their antimicrobial activity respectively. The acetone extract was purified by column chromatography and their MIC determined. The most potent fraction (EA4) was subjected to Gas chromatography- Mass spectrometry (GC-MS) and High performance liquid chromatography (HPLC) for identification of the active compounds. Results were analyzed by the Fisher‟s exact test. All the extracts tested demonstrated antimicrobial activity with zone diameters of inhibition ranging from 0–32 mm. Acetone was the most potent extract with its MIC ranging from 0.078–5.0 mg/mL. Seventeen fractions were collected from column chromatography and the most active fraction against all the organisms was EA 4 (eluted with 100 percent ethyl acetate), with its MIC ranging from 0.078 - 2.5mg/mL. There was no statistically significant difference (P>0.05) in the potency of the xii four extracts (acetone, methanol, ethanol and ethyl acetate) and antibiotic (ciprofloxacin) on the different bacterial strains tested, likewise the crude extract and the fractions. No compound was detected by GC-MS whereas numerous peaks were identified by HPLC implying that the active compounds in this plant are non volatile. We could not identify the compounds thereby proposing further studies using Nuclear magnetic resonance to identify the compounds. The study revealed that the acetone extract of C. molle was the most active against all the test organisms and therefore justifies the use of this plant in traditional medicine.
- Full Text:
- Date Issued: 2011
- Authors: Nyenje, Mirriam, E
- Date: 2011
- Subjects: Drug resistance in microorganisms , Materia medica, Vegetable , Antibiotics , Microbial sensitivity tests , Gram-negative bacterial infections
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11261 , http://hdl.handle.net/10353/391 , Drug resistance in microorganisms , Materia medica, Vegetable , Antibiotics , Microbial sensitivity tests , Gram-negative bacterial infections
- Description: Antimicrobial resistance is a worldwide problem that has deleterious long-term effects as the development of drug resistance outpaces the development of new drugs. Plants have been used for many generations for healing purposes, and screening of extracts of these plants has often yielded positive outcomes. This study was aimed at isolating and characterizing the major active antimicrobial compounds present in the stem bark of C. molle, in a bid to identify potential sources of cheap starting materials for the synthesis of new drugs. Various solvents (hexane, ethyl acetate, dichloromethane, acetone, ethanol and methanol) were used for extraction. The agar well diffusion technique was used to screen for antimicrobial activity of C. molle extracts against Streptococcus pyogenes ATCC 49399, Plesiomonas shigelloides ATCC 51903, Pseudomonas aeruginosa ATCC 15442, Helicobacter pylori ATCC 43526 and Helicobacter pylori 252C (clinical isolate); minimum inhibition concentration (MIC) of the most active extracts was determined by the broth dilution method. Fractionation of acetone extract was done by thin layer chromatography (TLC) and bioautography to determine the compounds present and their antimicrobial activity respectively. The acetone extract was purified by column chromatography and their MIC determined. The most potent fraction (EA4) was subjected to Gas chromatography- Mass spectrometry (GC-MS) and High performance liquid chromatography (HPLC) for identification of the active compounds. Results were analyzed by the Fisher‟s exact test. All the extracts tested demonstrated antimicrobial activity with zone diameters of inhibition ranging from 0–32 mm. Acetone was the most potent extract with its MIC ranging from 0.078–5.0 mg/mL. Seventeen fractions were collected from column chromatography and the most active fraction against all the organisms was EA 4 (eluted with 100 percent ethyl acetate), with its MIC ranging from 0.078 - 2.5mg/mL. There was no statistically significant difference (P>0.05) in the potency of the xii four extracts (acetone, methanol, ethanol and ethyl acetate) and antibiotic (ciprofloxacin) on the different bacterial strains tested, likewise the crude extract and the fractions. No compound was detected by GC-MS whereas numerous peaks were identified by HPLC implying that the active compounds in this plant are non volatile. We could not identify the compounds thereby proposing further studies using Nuclear magnetic resonance to identify the compounds. The study revealed that the acetone extract of C. molle was the most active against all the test organisms and therefore justifies the use of this plant in traditional medicine.
- Full Text:
- Date Issued: 2011
Genetic and phenotypic characterisation of foodborne bacteria isolated from ready-to-eat foods in Alice, South Africa
- Authors: Nyenje, Mirriam E
- Date: 2014
- Subjects: Foodborne diseases -- Microbiology , Pathogenic bacteria
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11279 , http://hdl.handle.net/10353/d1016109 , Foodborne diseases -- Microbiology , Pathogenic bacteria
- Description: Foodborne illnesses following the ingestion of contaminated food are a major public health problem worldwide. They include a broad group of illnesses ranging from mild to chronic or life-threatening; caused by either toxins released from the disease-causing microbes, or by the microbes themselves. Antimicrobial susceptibility data shows an alarming increase in the frequency of antimicrobial resistance of foodborne pathogens, a situation which is worrisome as it decreases the effectiveness of drugs employed to reduce the morbidity and mortality associated with serious and life-threatening infections and thus, compromising human health. This study was therefore designed to assess the occurrence and characterization of bacterial foodborne pathogens in various foods sold in Alice, Eastern Cape Province of South Africa in an effort to throw more light on the inherent risk associated with such foods. The study was conducted during the period of 2011 - 2013. Two university restaurants and eight ready-to-eat food vending sites in Alice Town were selected based on their prominence to the students, workers and rest of the community. Microbiological analysis was conducted on 252 samples which included vegetables, potatoes, rice, pies, beef and chicken stew. The isolates were identified using biochemical tests and confirmation of the two most prevalent organisms (Listeria ivanovii and Enterobacter cloacae) was done using PCR techniques. The antimicrobial susceptibility profile of Listeria ivanovii and Enterobacter cloacae strains were identified using the disc diffusion technique; minimum inhibitory concentration was determined by the broth dilution method and M.I.C. Evaluator test strips. The microtiter plate adherence assay was employed to ascertain the ability of these isolates to adhere to a surface whereas the role of cell surface properties in biofilm formation was assessed using the coaggregation and autoaggregation assays. The architecture of the formed biofilms was examined under the scanning electron microscope. The virulence and resistant genes were also detected and characterised by sequencing the PCR products. Bacterial growth was present in all the food types tested; organisms isolated included: Listeria spp. (22%), Enterobacter spp. (18%), Aeromonas hydrophila (12%), Klebsiella oxytoca (8%), Proteus mirabilis (6.3%), Staphylococcus aureus (3.2%) and Pseudomonas luteola (2.4%). PCR confirmed 30 (97%) isolates as E. cloacae complex while 44% (22/50) tested positive for L. ivanovii. All the strains of E. cloacae (100%) and 96% of L. ivanovii isolates (based on phenotypic identification) were resistant to at least four or more of the antibiotics. In this study, bla-TEM was also detected from 48 (96%) of L. ivanovii and 30 (100%) of E. cloacae strains; further analysis of the bla-TEM demonstrated the occurrence of bla-TEM-1. Of the 56 bla-TEM-1 positive isolates sequenced, 7% (4/56) had mutation of either insertion or substitution of a nucleotide. Two virulence genes (ucaA and hlyA) were detected in E. cloacae isolates and none in L. ivanovii using PCR. Sequence analysis of the hsp60 gene reported the presence of two sub-species for E. cloacae; E. cloacae cluster III (75%) and E. cloacae cluster IV (25%); while analysis of the iap60 gene demonstrated that 55.8% (19/34) were L. ivanovii, 44% (15/34) L. seeligeri and 14.7% (5/34) L. welshemeri. A total of 90% L. ivanovii and 88% E. cloacae strains demonstrated the ability to form biofilms; the coaggregation index ranged from 12 to 77% while the autoaggregation index varied from 11 to 55% for L. ivanovii and 27% to 98% for E. cloacae. The findings of this study indicate that most of the ready-to-eat food samples examined did not meet bacteriological quality standards, thus posing potential risks to consumers. This should draw the attention of the relevant authorities to certify that hygienic standards are improved to curtain foodborne infections. Furthermore, the presence of multi-resistant strains is of major concern as these foods could serve as important vehicles transmitting multi-resistant bacteria and genes to humans. In addition the ability of these pathogens to form biofilms may lead to adherence of these organisms to kitchen utensils and other environments leading to cross-contamination of food processed in these areas and increase resistance of organisms to antimicrobial agents.
- Full Text:
- Date Issued: 2014
- Authors: Nyenje, Mirriam E
- Date: 2014
- Subjects: Foodborne diseases -- Microbiology , Pathogenic bacteria
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11279 , http://hdl.handle.net/10353/d1016109 , Foodborne diseases -- Microbiology , Pathogenic bacteria
- Description: Foodborne illnesses following the ingestion of contaminated food are a major public health problem worldwide. They include a broad group of illnesses ranging from mild to chronic or life-threatening; caused by either toxins released from the disease-causing microbes, or by the microbes themselves. Antimicrobial susceptibility data shows an alarming increase in the frequency of antimicrobial resistance of foodborne pathogens, a situation which is worrisome as it decreases the effectiveness of drugs employed to reduce the morbidity and mortality associated with serious and life-threatening infections and thus, compromising human health. This study was therefore designed to assess the occurrence and characterization of bacterial foodborne pathogens in various foods sold in Alice, Eastern Cape Province of South Africa in an effort to throw more light on the inherent risk associated with such foods. The study was conducted during the period of 2011 - 2013. Two university restaurants and eight ready-to-eat food vending sites in Alice Town were selected based on their prominence to the students, workers and rest of the community. Microbiological analysis was conducted on 252 samples which included vegetables, potatoes, rice, pies, beef and chicken stew. The isolates were identified using biochemical tests and confirmation of the two most prevalent organisms (Listeria ivanovii and Enterobacter cloacae) was done using PCR techniques. The antimicrobial susceptibility profile of Listeria ivanovii and Enterobacter cloacae strains were identified using the disc diffusion technique; minimum inhibitory concentration was determined by the broth dilution method and M.I.C. Evaluator test strips. The microtiter plate adherence assay was employed to ascertain the ability of these isolates to adhere to a surface whereas the role of cell surface properties in biofilm formation was assessed using the coaggregation and autoaggregation assays. The architecture of the formed biofilms was examined under the scanning electron microscope. The virulence and resistant genes were also detected and characterised by sequencing the PCR products. Bacterial growth was present in all the food types tested; organisms isolated included: Listeria spp. (22%), Enterobacter spp. (18%), Aeromonas hydrophila (12%), Klebsiella oxytoca (8%), Proteus mirabilis (6.3%), Staphylococcus aureus (3.2%) and Pseudomonas luteola (2.4%). PCR confirmed 30 (97%) isolates as E. cloacae complex while 44% (22/50) tested positive for L. ivanovii. All the strains of E. cloacae (100%) and 96% of L. ivanovii isolates (based on phenotypic identification) were resistant to at least four or more of the antibiotics. In this study, bla-TEM was also detected from 48 (96%) of L. ivanovii and 30 (100%) of E. cloacae strains; further analysis of the bla-TEM demonstrated the occurrence of bla-TEM-1. Of the 56 bla-TEM-1 positive isolates sequenced, 7% (4/56) had mutation of either insertion or substitution of a nucleotide. Two virulence genes (ucaA and hlyA) were detected in E. cloacae isolates and none in L. ivanovii using PCR. Sequence analysis of the hsp60 gene reported the presence of two sub-species for E. cloacae; E. cloacae cluster III (75%) and E. cloacae cluster IV (25%); while analysis of the iap60 gene demonstrated that 55.8% (19/34) were L. ivanovii, 44% (15/34) L. seeligeri and 14.7% (5/34) L. welshemeri. A total of 90% L. ivanovii and 88% E. cloacae strains demonstrated the ability to form biofilms; the coaggregation index ranged from 12 to 77% while the autoaggregation index varied from 11 to 55% for L. ivanovii and 27% to 98% for E. cloacae. The findings of this study indicate that most of the ready-to-eat food samples examined did not meet bacteriological quality standards, thus posing potential risks to consumers. This should draw the attention of the relevant authorities to certify that hygienic standards are improved to curtain foodborne infections. Furthermore, the presence of multi-resistant strains is of major concern as these foods could serve as important vehicles transmitting multi-resistant bacteria and genes to humans. In addition the ability of these pathogens to form biofilms may lead to adherence of these organisms to kitchen utensils and other environments leading to cross-contamination of food processed in these areas and increase resistance of organisms to antimicrobial agents.
- Full Text:
- Date Issued: 2014
Studies on bioflocculant production by a consortium of two bacterial species belonging to the Methylobacterium and Actinobacterium genera
- Authors: Ntsaluba, Luvuyo
- Date: 2012
- Subjects: Flocculation , Actinobacteria , Methylobacterium , Water -- Purification
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11266 , http://hdl.handle.net/10353/482 , Flocculation , Actinobacteria , Methylobacterium , Water -- Purification
- Description: Bioflocculants produced by two identified bacteria: Actinobacterium sp. Mayor and Methylobacterium sp. Obi were investigated with regard to their physicochemical and flocculating characteristics. The two strains were later combined to form a consortium for further studies. The optimum culture conditions for the bioflocculant production were similar for all strains except in the case of Actinobacterium sp. Mayor and the consortium, where glucose was replaced by sodium carbonate as a carbon source. Multi-nitrogen source was the best nitrogen source compare to individual sources for both strains. The divalent cation, Ca2+ proved to be a better flocculating activity stimulus for all produced bioflocculants in this study. The optimum flocculating activities obtained for both individual strains and the consortium were all at alkaline pH. The yield of purified bioflocculant produced by the consortium was 8.203 g/l, while 4.190 g/l and 4.610 g/l were recovered for single strains of Actinobacterium sp. Mayor and Methylobacterium sp. Obi respectively. Further characterization of pure bioflocculants revealed that a bioflocculant dosage of 0.3 mg/ml resulted in the highest flocculating activity for both individual strains while 1.0 mg/ml of the bioflocculant produced by the consortium was required to enhance maximum flocculating efficiency. These bioflocculants proved to be all thermo stable at a temperature range of 20 to 900°C with a heating rate of 10oC/min under a constant flow of nitrogen gas. The presence of functional groups normally required for bioflocculation such as hydroxyl, carboxyl and amino was also detected. The findings of this study suggest that the producedbioflocculants can be utilized as excellent substitutes for harmful synthetic flocculants in both water and wastewater treatments as well as in other industrial applications.
- Full Text:
- Date Issued: 2012
- Authors: Ntsaluba, Luvuyo
- Date: 2012
- Subjects: Flocculation , Actinobacteria , Methylobacterium , Water -- Purification
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11266 , http://hdl.handle.net/10353/482 , Flocculation , Actinobacteria , Methylobacterium , Water -- Purification
- Description: Bioflocculants produced by two identified bacteria: Actinobacterium sp. Mayor and Methylobacterium sp. Obi were investigated with regard to their physicochemical and flocculating characteristics. The two strains were later combined to form a consortium for further studies. The optimum culture conditions for the bioflocculant production were similar for all strains except in the case of Actinobacterium sp. Mayor and the consortium, where glucose was replaced by sodium carbonate as a carbon source. Multi-nitrogen source was the best nitrogen source compare to individual sources for both strains. The divalent cation, Ca2+ proved to be a better flocculating activity stimulus for all produced bioflocculants in this study. The optimum flocculating activities obtained for both individual strains and the consortium were all at alkaline pH. The yield of purified bioflocculant produced by the consortium was 8.203 g/l, while 4.190 g/l and 4.610 g/l were recovered for single strains of Actinobacterium sp. Mayor and Methylobacterium sp. Obi respectively. Further characterization of pure bioflocculants revealed that a bioflocculant dosage of 0.3 mg/ml resulted in the highest flocculating activity for both individual strains while 1.0 mg/ml of the bioflocculant produced by the consortium was required to enhance maximum flocculating efficiency. These bioflocculants proved to be all thermo stable at a temperature range of 20 to 900°C with a heating rate of 10oC/min under a constant flow of nitrogen gas. The presence of functional groups normally required for bioflocculation such as hydroxyl, carboxyl and amino was also detected. The findings of this study suggest that the producedbioflocculants can be utilized as excellent substitutes for harmful synthetic flocculants in both water and wastewater treatments as well as in other industrial applications.
- Full Text:
- Date Issued: 2012
Assessment of bioflocculant production by two marine bacteria isolated from the bottom sediment of marine Algoa Bay
- Authors: Ntozonke, Ncedo
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11298 , http://hdl.handle.net/10353/d1021290
- Description: Bioflocculants are polymers, mostly, of microbial origin which floc out suspended particles from liquid medium. The ability of these biopolymers to remove suspended particles from solutions is termed bioflocculation, and the efficiency of flocculation activities depends on the characteristics of the flocculants. In comparison with conventionally used flocculants, bioflocculants have the advantage of being safe (no toxic effects known), biodegradable and harmlessness to the environment. The study assessed production of bioflocculant by two marine bacteria from the bottom sediment of marine environment. The 16S rDNA was used for identification, and the two bacteria species were identified as Enterococcus hirae and Bacillus thuringiensis. Factors affecting the production and activity of the bioflocculants produced by these two organisms were studied. The bacteria optimally produced bioflocculant with fructose (91.7%) and urea (91%) as sole carbon and nitrogen sources respectively. Mg2+ (87%) and Ca2+ (86%), likewise, served as best cation sources on the production of the bioflocculant at pH 5(93%). Additionally, the flocculating activity of the bioflocculant increased with the addition of Mg2+ (81%) and Na+ (81%), and the highest flocculating activity was at pH 5 of the kaolin clay. The Fourier transform infrared spectroscopy (FTIR) shows that the bioflocculant is a glycoprotein. The second bacterium (Bacillus thuringiensis) produced bioflocculant optimally when the media had mixed nitrogen sources (Urea, ammonium chloride and tryptone (67%)) and glucose (85.65%) as a sole carbon source, also Ca2+ (74.6%) was the best cation that induced the production of bioflocculant. After purification, the bioflocculant flocculated optimally in alkaline pH 12 (81%) in the presence of Mn2+ (73%) and Ca2+ (72.8%). Chemical analysis of the bioflocculant revealed it to be a polysaccharide. Both bioflocculants flocculate efficiently and can be used to replace synthetic flocculants in water treatment, wastewater, in downstream processing, and processing of food and chemicals and other industrial uses of flocculants. Challenges though (i) are to develop conditions for large scale production of the bioflocculant, (ii) to do further characterization of the both bioflocculants (iii) to assess the bioflocculants for treatments of water/wastewater, and to apply it in various industrial processes.
- Full Text:
- Date Issued: 2015
- Authors: Ntozonke, Ncedo
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11298 , http://hdl.handle.net/10353/d1021290
- Description: Bioflocculants are polymers, mostly, of microbial origin which floc out suspended particles from liquid medium. The ability of these biopolymers to remove suspended particles from solutions is termed bioflocculation, and the efficiency of flocculation activities depends on the characteristics of the flocculants. In comparison with conventionally used flocculants, bioflocculants have the advantage of being safe (no toxic effects known), biodegradable and harmlessness to the environment. The study assessed production of bioflocculant by two marine bacteria from the bottom sediment of marine environment. The 16S rDNA was used for identification, and the two bacteria species were identified as Enterococcus hirae and Bacillus thuringiensis. Factors affecting the production and activity of the bioflocculants produced by these two organisms were studied. The bacteria optimally produced bioflocculant with fructose (91.7%) and urea (91%) as sole carbon and nitrogen sources respectively. Mg2+ (87%) and Ca2+ (86%), likewise, served as best cation sources on the production of the bioflocculant at pH 5(93%). Additionally, the flocculating activity of the bioflocculant increased with the addition of Mg2+ (81%) and Na+ (81%), and the highest flocculating activity was at pH 5 of the kaolin clay. The Fourier transform infrared spectroscopy (FTIR) shows that the bioflocculant is a glycoprotein. The second bacterium (Bacillus thuringiensis) produced bioflocculant optimally when the media had mixed nitrogen sources (Urea, ammonium chloride and tryptone (67%)) and glucose (85.65%) as a sole carbon source, also Ca2+ (74.6%) was the best cation that induced the production of bioflocculant. After purification, the bioflocculant flocculated optimally in alkaline pH 12 (81%) in the presence of Mn2+ (73%) and Ca2+ (72.8%). Chemical analysis of the bioflocculant revealed it to be a polysaccharide. Both bioflocculants flocculate efficiently and can be used to replace synthetic flocculants in water treatment, wastewater, in downstream processing, and processing of food and chemicals and other industrial uses of flocculants. Challenges though (i) are to develop conditions for large scale production of the bioflocculant, (ii) to do further characterization of the both bioflocculants (iii) to assess the bioflocculants for treatments of water/wastewater, and to apply it in various industrial processes.
- Full Text:
- Date Issued: 2015
Enterococcus pathotypes as reservoirs of antibiotic resistance determinants in the Kat River and Fort Beaufort abstraction waters
- Authors: Ntloko, Phindiwe
- Date: 2014
- Subjects: Enterococcus , Drug resistance
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11290 , http://hdl.handle.net/10353/d1019821 , Enterococcus , Drug resistance
- Description: In this study, 400 presumptive Enterococcus isolates previously recovered from Kat River and Fort Beaufort Abstraction water dam were subjected to molecular confirmation and pathotyping. Two hundred and seventy-four (68%) of these isolates were confirmed to be enterococci species. Confirmations studies were polymerase chain reaction (PCR) based, using enterococci specific primers targeting the tuf gene. The confirmed enterococci isolates were further differentiated into their pathotypes, the targets of which were: E. faecalis, E. avium, E. hirae, E. casseliflavarus and E. gallinarum using well documented species specific primer sequences. E. faecalis accounted for 20% of the isolates, followed by E. avium (16%), E. hirae (13%), E. casseliflavarus (5%) and E. gallinarum (3%). Furthermore, all the confirmed isolates were analysed for antibiotic susceptibilities using a panel of nine different antibiotics, namely vancomycin, linezolid, ciprofloxacin, ampicillin, gentamicin, chloramphenicol, tetracycline, erythromycin, penicillin, and those that were resistant were assayed for the presence of relevant antibiotic resistance genes. All the 274 isolates were found to harbour vanA resistance gene confirming their phenotypic resistance to the vancomycin. Similarly, 60% (109/180) of the isolates showed phenotypic resistance to erythromycin which was further confirmed by the presence of ermA genes in these isolates. The presence of antibiotic resistant bacteria in surface waters poses a risk to public health.
- Full Text:
- Date Issued: 2014
- Authors: Ntloko, Phindiwe
- Date: 2014
- Subjects: Enterococcus , Drug resistance
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11290 , http://hdl.handle.net/10353/d1019821 , Enterococcus , Drug resistance
- Description: In this study, 400 presumptive Enterococcus isolates previously recovered from Kat River and Fort Beaufort Abstraction water dam were subjected to molecular confirmation and pathotyping. Two hundred and seventy-four (68%) of these isolates were confirmed to be enterococci species. Confirmations studies were polymerase chain reaction (PCR) based, using enterococci specific primers targeting the tuf gene. The confirmed enterococci isolates were further differentiated into their pathotypes, the targets of which were: E. faecalis, E. avium, E. hirae, E. casseliflavarus and E. gallinarum using well documented species specific primer sequences. E. faecalis accounted for 20% of the isolates, followed by E. avium (16%), E. hirae (13%), E. casseliflavarus (5%) and E. gallinarum (3%). Furthermore, all the confirmed isolates were analysed for antibiotic susceptibilities using a panel of nine different antibiotics, namely vancomycin, linezolid, ciprofloxacin, ampicillin, gentamicin, chloramphenicol, tetracycline, erythromycin, penicillin, and those that were resistant were assayed for the presence of relevant antibiotic resistance genes. All the 274 isolates were found to harbour vanA resistance gene confirming their phenotypic resistance to the vancomycin. Similarly, 60% (109/180) of the isolates showed phenotypic resistance to erythromycin which was further confirmed by the presence of ermA genes in these isolates. The presence of antibiotic resistant bacteria in surface waters poses a risk to public health.
- Full Text:
- Date Issued: 2014
Evaluation of incidence of Mycobacterium tuberculosis complex associated with soil, hayfeed and water in three Agricultural facilities in Amathole District Municipality in the Eastern Cape Province, South Africa
- Authors: Ntloko, Athini
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: http://hdl.handle.net/10353/756 , vital:26494
- Description: Mycobacterium bovis and other species of Mycobacterium tuberculosis complex (MTBC) can result to a zoonotic infection known as Bovine tuberculosis (bTB). MTBC has members that may contaminate an extensive range of hosts, including wildlife. Diverse wild species are known to cause disease in domestic livestock and are acknowledged as TB reservoirs. It has been a main study worldwide to deliberate on bTB risk factors as a result some studies focused on particular parts of risk factors such as wildlife and herd management. The objectives of this study were to design questionnaires from commercial farms and smallholding farms; isolate and identify MTBC from collected samples using culture and PCR assays recovered from Fort Hare, Middledrift and Seven star dairy farms; and assessing genotypic drug resistance through detection of mutations conferring resistance to INH and RMP associated with first line treatment for MTBC infection. Questionnaires were administered to thirty (30) smallholding farm owners in the two villages (kwaMasele and Qungqwala) and three (3) three commercial farms ( Fort Hare dairy farm, Middledrift dairy farm and Seven star dairy farm). Detection of M. tuberculosis complex was achieved by Polymerase Chain Reaction using primers for IS6110; whereas a genotypic drug resistance mutation was detected using Genotype MTBDRplus assays. Nine percent (9%) of respondents had more than 40 cows in their herd, while 60% reported between 10 and 20 cows in their herd. Relationship between farm size and vaccination for TB differed from forty one percent (41%) being the highest to the least five percent (5%). The highest number of respondents who knew about relationship between TB cases and cattle location was ninety one percent (91%). Approximately fifty one percent (51%) of respondents had knowledge about wild life access to the farms. Relationship between import of cattle and farm size ranged from nine percent (9%) to thirty five percent (35%). Cattle sickness in relation to farm size differed from forty three (43%) being the highest to the least three percent (3%); while thirty three percent (33%) of respondents had knowledge about health management. Respondents with knowledge about the occurrence of TB infections in farms were forty eight percent (48%). The frequency of DNA isolation from samples ranged from the highest forty five percent (45%) from water to the least twenty two percent (22%) from soil. Fort Hare dairy farm had the highest number of positive samples forty four percent (44%) from water samples; whereas Middledrift dairy farm had the lowest positive from water, seventeen percent (17%). Twelve (22%) out of 55 isolates showed resistance to INH and RMP that is, multi-drug resistance (MDR) and nine percent (9%) were sensitive to either INH or RMP. The mutations at rpoB gene differed from 58% being the highest to the least (23%). Fifty seven percent (57%) of samples showed a S315T1 mutation while only 14% possessed a S531L in the katG gene. The highest inhA mutations were detected in T8A (80%) eighty percent and the least was observed in A16G (17%). The results of this study reveals that risk factors for bTB in cattle and dairy farm workers is a serious issue abound in the Eastern Cape of South Africa; with the possibility of widespread dissemination of multidrug resistant determinants in MTBC from the environment.
- Full Text:
- Date Issued: 2015
- Authors: Ntloko, Athini
- Date: 2015
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: http://hdl.handle.net/10353/756 , vital:26494
- Description: Mycobacterium bovis and other species of Mycobacterium tuberculosis complex (MTBC) can result to a zoonotic infection known as Bovine tuberculosis (bTB). MTBC has members that may contaminate an extensive range of hosts, including wildlife. Diverse wild species are known to cause disease in domestic livestock and are acknowledged as TB reservoirs. It has been a main study worldwide to deliberate on bTB risk factors as a result some studies focused on particular parts of risk factors such as wildlife and herd management. The objectives of this study were to design questionnaires from commercial farms and smallholding farms; isolate and identify MTBC from collected samples using culture and PCR assays recovered from Fort Hare, Middledrift and Seven star dairy farms; and assessing genotypic drug resistance through detection of mutations conferring resistance to INH and RMP associated with first line treatment for MTBC infection. Questionnaires were administered to thirty (30) smallholding farm owners in the two villages (kwaMasele and Qungqwala) and three (3) three commercial farms ( Fort Hare dairy farm, Middledrift dairy farm and Seven star dairy farm). Detection of M. tuberculosis complex was achieved by Polymerase Chain Reaction using primers for IS6110; whereas a genotypic drug resistance mutation was detected using Genotype MTBDRplus assays. Nine percent (9%) of respondents had more than 40 cows in their herd, while 60% reported between 10 and 20 cows in their herd. Relationship between farm size and vaccination for TB differed from forty one percent (41%) being the highest to the least five percent (5%). The highest number of respondents who knew about relationship between TB cases and cattle location was ninety one percent (91%). Approximately fifty one percent (51%) of respondents had knowledge about wild life access to the farms. Relationship between import of cattle and farm size ranged from nine percent (9%) to thirty five percent (35%). Cattle sickness in relation to farm size differed from forty three (43%) being the highest to the least three percent (3%); while thirty three percent (33%) of respondents had knowledge about health management. Respondents with knowledge about the occurrence of TB infections in farms were forty eight percent (48%). The frequency of DNA isolation from samples ranged from the highest forty five percent (45%) from water to the least twenty two percent (22%) from soil. Fort Hare dairy farm had the highest number of positive samples forty four percent (44%) from water samples; whereas Middledrift dairy farm had the lowest positive from water, seventeen percent (17%). Twelve (22%) out of 55 isolates showed resistance to INH and RMP that is, multi-drug resistance (MDR) and nine percent (9%) were sensitive to either INH or RMP. The mutations at rpoB gene differed from 58% being the highest to the least (23%). Fifty seven percent (57%) of samples showed a S315T1 mutation while only 14% possessed a S531L in the katG gene. The highest inhA mutations were detected in T8A (80%) eighty percent and the least was observed in A16G (17%). The results of this study reveals that risk factors for bTB in cattle and dairy farm workers is a serious issue abound in the Eastern Cape of South Africa; with the possibility of widespread dissemination of multidrug resistant determinants in MTBC from the environment.
- Full Text:
- Date Issued: 2015
Major spoligotype families of Mycobacterium tuberculosis strains isolated from tuberculosis patients in Port Elizabeth, Eastern Cape, South Africa
- Authors: Nqini, Babalwa J
- Date: 2012
- Subjects: Mycobacterium tuberculosis -- South Africa -- Eastern Cape , Tuberculosis -- Patients -- South Africa -- Eastern Cape , Drug resistance , Multidrug-resistant tuberculosis -- South Africa -- Eastern Cape , HIV infections -- South Africa -- Eastern Cape , AIDS (Disease) -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11271 , http://hdl.handle.net/10353/d1006877 , Mycobacterium tuberculosis -- South Africa -- Eastern Cape , Tuberculosis -- Patients -- South Africa -- Eastern Cape , Drug resistance , Multidrug-resistant tuberculosis -- South Africa -- Eastern Cape , HIV infections -- South Africa -- Eastern Cape , AIDS (Disease) -- South Africa -- Eastern Cape
- Description: South Africa is burdened with tuberculosis (TB) which is aggravated by the concurrent epidemic of HIV as well as the emergence of drug resistance. In most developed countries molecular techniques have been used to look at the dynamics of the TB epidemic however, despite the prevalence that is high in sub-Saharan Africa, there is little data on strain types that are available in Port Elizabeth. This study aims to find the major clades of M. tuberculosis that are circulating in Port Elizabeth. Two hundred MDR-TB DNA samples were obtained from the National Health Laboratory Services TB laboratory in Port Elizabeth. Spoligotyping and MIRU-VNTR were used to genotype the strains. Two hundred strains were sent to the University of Stellenbosch for spoligotyping and 179 of those were typed. Spoligotype defined families were further typed by MIRU-VNTR typing, so as to further differentiate and assess clonal diversity within the spoligotype families. The Beijing family was the dominant family and the MANU family being the least dominant, with percentages of 71 percent and 0.5 percent respectively. A comparison of spoligotyping results with the international spoligotyping database (SITVIT2) showed a total of 15 shared international types. Forty four percent (44 percent) of the isolates that were typed by MIRU-VNTR showed similarities, suggesting epidemiological relatedness. Thirty eight percent of isolates from spoligotyping were from the same family, the Beijing family, with the same shared international type STI1, but when typed by 12 MIRU-VNTR they showed no epidemiological relatedness and 18 percent of the isolates showed no relatedness when typed by 12 MIRU-VNTR but spoligotyping showed that they were from the LAM family. Results from our study illustrate the effectiveness of MIRU-VNTR typing together with spoligotyping in epidemiological studies in the region of Port Elizabeth.
- Full Text:
- Date Issued: 2012
- Authors: Nqini, Babalwa J
- Date: 2012
- Subjects: Mycobacterium tuberculosis -- South Africa -- Eastern Cape , Tuberculosis -- Patients -- South Africa -- Eastern Cape , Drug resistance , Multidrug-resistant tuberculosis -- South Africa -- Eastern Cape , HIV infections -- South Africa -- Eastern Cape , AIDS (Disease) -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11271 , http://hdl.handle.net/10353/d1006877 , Mycobacterium tuberculosis -- South Africa -- Eastern Cape , Tuberculosis -- Patients -- South Africa -- Eastern Cape , Drug resistance , Multidrug-resistant tuberculosis -- South Africa -- Eastern Cape , HIV infections -- South Africa -- Eastern Cape , AIDS (Disease) -- South Africa -- Eastern Cape
- Description: South Africa is burdened with tuberculosis (TB) which is aggravated by the concurrent epidemic of HIV as well as the emergence of drug resistance. In most developed countries molecular techniques have been used to look at the dynamics of the TB epidemic however, despite the prevalence that is high in sub-Saharan Africa, there is little data on strain types that are available in Port Elizabeth. This study aims to find the major clades of M. tuberculosis that are circulating in Port Elizabeth. Two hundred MDR-TB DNA samples were obtained from the National Health Laboratory Services TB laboratory in Port Elizabeth. Spoligotyping and MIRU-VNTR were used to genotype the strains. Two hundred strains were sent to the University of Stellenbosch for spoligotyping and 179 of those were typed. Spoligotype defined families were further typed by MIRU-VNTR typing, so as to further differentiate and assess clonal diversity within the spoligotype families. The Beijing family was the dominant family and the MANU family being the least dominant, with percentages of 71 percent and 0.5 percent respectively. A comparison of spoligotyping results with the international spoligotyping database (SITVIT2) showed a total of 15 shared international types. Forty four percent (44 percent) of the isolates that were typed by MIRU-VNTR showed similarities, suggesting epidemiological relatedness. Thirty eight percent of isolates from spoligotyping were from the same family, the Beijing family, with the same shared international type STI1, but when typed by 12 MIRU-VNTR they showed no epidemiological relatedness and 18 percent of the isolates showed no relatedness when typed by 12 MIRU-VNTR but spoligotyping showed that they were from the LAM family. Results from our study illustrate the effectiveness of MIRU-VNTR typing together with spoligotyping in epidemiological studies in the region of Port Elizabeth.
- Full Text:
- Date Issued: 2012
Antibiogram profiling of Escherichia coli pathotypes isolated from Kat River and Fort Beaufort abstraction water
- Authors: Nontongana, Nolonwabo
- Date: 2014
- Subjects: Escherichia coli infections -- South Africa -- Eastern Cape , Water -- Pollution -- Environmental aspects -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11289 , http://hdl.handle.net/10353/d1019820 , Escherichia coli infections -- South Africa -- Eastern Cape , Water -- Pollution -- Environmental aspects -- South Africa -- Eastern Cape
- Description: Escherichia coli (E. coli) is a widespread species that includes a broad variety of strains, ranging from highly pathogenic strains causing worldwide outbreaks of severe disease to virulent isolates which are part of the normal intestinal flora or which are well characterized and safe laboratory strains. The pathogenicity of a given E. coli strain is mainly determined by specific virulence factors which include adhesins, invasins, toxins and capsule. The aim of this study was to evaluate the prevalence and antibiogram profiles of E. coli pathotypes previously isolated from Kat River and Fort Beaufort abstraction water. A total of 171 E. coli isolates showed at least one pathogenic determinant among the isolated 278 E. coli. The other 107 isolates were negative for the tested virulence genes. All 278 presumptive isolates tested positive for the UidA gene, and were therefore classified as non-categorized pathogenic E. coli. The 171 pathogenic isolates had at least one characteristic gene of pathogenic E. coli and were identified and classified as enteropathogenic E. coli (6%), enterotoxigenicE. coli (131), uropathogenic E. coli (6), neonatal meningitis E. coli (14), diffusely adherent E. coli (1) and enterohaemrrhagic E. coli (1). Interestingly, no virulence genes were detected for the enteroinvasive E. coli and the enteroaggregative E. coli. The antibiotic resistance profiles for all isolates that were identified as E. coli showed 100% resistance to penicillin G, 98% resistance to ampicillin, 38% resistance to trimethoprim-sulphamethoxazole and 8% resistance to streptomycin. Multiple antibacterial resistance (MAR) was also observed, where 44% of the isolates were resistant to three antibiotics and 8% resistant to four antibiotics. The results of this study showed the Kat River and Fort Beaufort abstraction water are reservoirs of pathogenic strains of E. coli which harbour antibiotic resistance determinants that can cause serious health risks to the people in the surrounding communities.
- Full Text:
- Date Issued: 2014
- Authors: Nontongana, Nolonwabo
- Date: 2014
- Subjects: Escherichia coli infections -- South Africa -- Eastern Cape , Water -- Pollution -- Environmental aspects -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11289 , http://hdl.handle.net/10353/d1019820 , Escherichia coli infections -- South Africa -- Eastern Cape , Water -- Pollution -- Environmental aspects -- South Africa -- Eastern Cape
- Description: Escherichia coli (E. coli) is a widespread species that includes a broad variety of strains, ranging from highly pathogenic strains causing worldwide outbreaks of severe disease to virulent isolates which are part of the normal intestinal flora or which are well characterized and safe laboratory strains. The pathogenicity of a given E. coli strain is mainly determined by specific virulence factors which include adhesins, invasins, toxins and capsule. The aim of this study was to evaluate the prevalence and antibiogram profiles of E. coli pathotypes previously isolated from Kat River and Fort Beaufort abstraction water. A total of 171 E. coli isolates showed at least one pathogenic determinant among the isolated 278 E. coli. The other 107 isolates were negative for the tested virulence genes. All 278 presumptive isolates tested positive for the UidA gene, and were therefore classified as non-categorized pathogenic E. coli. The 171 pathogenic isolates had at least one characteristic gene of pathogenic E. coli and were identified and classified as enteropathogenic E. coli (6%), enterotoxigenicE. coli (131), uropathogenic E. coli (6), neonatal meningitis E. coli (14), diffusely adherent E. coli (1) and enterohaemrrhagic E. coli (1). Interestingly, no virulence genes were detected for the enteroinvasive E. coli and the enteroaggregative E. coli. The antibiotic resistance profiles for all isolates that were identified as E. coli showed 100% resistance to penicillin G, 98% resistance to ampicillin, 38% resistance to trimethoprim-sulphamethoxazole and 8% resistance to streptomycin. Multiple antibacterial resistance (MAR) was also observed, where 44% of the isolates were resistant to three antibiotics and 8% resistant to four antibiotics. The results of this study showed the Kat River and Fort Beaufort abstraction water are reservoirs of pathogenic strains of E. coli which harbour antibiotic resistance determinants that can cause serious health risks to the people in the surrounding communities.
- Full Text:
- Date Issued: 2014
Evaluation of the final effluents of some wastewater treatment plants in Amathole and Chris Hani District Municipality of the Eastern Cape Province as sources of vibrio pathogens in the aquatic environment
- Authors: Nongogo, Vuyokazi
- Date: 2014
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11287 , http://hdl.handle.net/10353/d1019813
- Description: Certain areas in the world still depend on the receiving water bodies as sources of domestic water and for recreational purposes. The discharge of poor quality effluents from wastewater treatment plants can impact negatively on these water bodies, as they can act as vehicles for pathogens to the environment, posing a threat to humans if such water is used without precaution. Vibrio species are amongst those pathogens that can survive wastewater treatment processes, ending up in the environment, hence the aim of this study was to evaluate the final effluents of some wastewater treatment plants as sources of vibrio pathogens. Five wastewater treatment plants (WWTP) located in Amathole and Chris Hani District Municipalities in the Eastern Cape were used in this study. Samples were collected monthly from September 2012 – August 2013 and analysed using the standard membrane filtration technique. Yellow and green colonies on TCBS agar were enumerated as presumptive Vibrio species and expressed as CFU/100ml for each plant. Colonies were later picked based on their phenotypic characteristics, sub-cultured on fresh TCBS agar to ascertain purity. These presumptive isolates were then subjected to Gram staining and Oxidase test. Gram negative and Oxidase positive isolates were selected for further confirmation using Polymerised Chain Reaction (PCR). PCR was also employed for characterisation of Vibrio into three species viz V. parahaemolyticus, V. fluvialis and V. vulnificus. Antibiogram profile of the characterised species was then determined together with the presence of relevant antibiotic resistance genes Vibrio densities for the twelve month period ranged between 0 - 1.48×104 CFU/100ml with two of the plants located in East bank and Queenstown characterized by extremely high counts and one plant( Reeston) with very low counts.Three hundred presumptive Vibrio isolates were screened for identity confirmation. Of these, the dominating species found was V. fluvialis (28.6%) followed by V. vulnificus (28%) and the least was found to be V. parahaemolyticus (11.6%). The remaining unidentified 31.6% were suspected to belong to other Vibrio species not covered within the scope of this study. All the confirmed isolates i.e., V. parahaemolyticus, V. vulnificus and V. fluvialis were susceptible to imipenem, gentamicin and meropenem and resistant to only tetracycline. Between 60-100% of the V. parahaemolyticus isolates, 7.1% to 100 % V. vulnificus isolates and 2.5 to 100 % V. fluvialis showed resistances to polymixin B, sulfamethazole, erythromycin, penicillin G, chloramphenicol, trimethroprim and trimethroprim & sulfamethazole. Antibiotic Resistance Genes that were assessed included dfRA, SXT, floR and Sul2 varying in proportion with each species showing diversity in the Vibrio community. The dfR A gene was detected in all the V. parahaemolyticus isolates while floR gene was not detected in any of the isolates belonging to the three species. The distribution of sul2 gene cut across the species being 1% (1) in V. fluvialis, 3% (1) in V. parahaemolyticus and 4% (3) in V. vulnificus. The SXT gene was only determined in V. parahaemolyticus. It is clear that the final effluents of the selected plants are reservoirs for Vibrio pathogens as well as antibiotic resistance genes in the environment. The isolation of Vibrio from WWTP shows that this pathogen is in circulation in some pockets of the population. Therefore, wastewater treatment plants need to be properly monitored to ensure that they comply with set guidelines.
- Full Text:
- Date Issued: 2014
- Authors: Nongogo, Vuyokazi
- Date: 2014
- Language: English
- Type: Thesis , Masters , MSc (Microbiology)
- Identifier: vital:11287 , http://hdl.handle.net/10353/d1019813
- Description: Certain areas in the world still depend on the receiving water bodies as sources of domestic water and for recreational purposes. The discharge of poor quality effluents from wastewater treatment plants can impact negatively on these water bodies, as they can act as vehicles for pathogens to the environment, posing a threat to humans if such water is used without precaution. Vibrio species are amongst those pathogens that can survive wastewater treatment processes, ending up in the environment, hence the aim of this study was to evaluate the final effluents of some wastewater treatment plants as sources of vibrio pathogens. Five wastewater treatment plants (WWTP) located in Amathole and Chris Hani District Municipalities in the Eastern Cape were used in this study. Samples were collected monthly from September 2012 – August 2013 and analysed using the standard membrane filtration technique. Yellow and green colonies on TCBS agar were enumerated as presumptive Vibrio species and expressed as CFU/100ml for each plant. Colonies were later picked based on their phenotypic characteristics, sub-cultured on fresh TCBS agar to ascertain purity. These presumptive isolates were then subjected to Gram staining and Oxidase test. Gram negative and Oxidase positive isolates were selected for further confirmation using Polymerised Chain Reaction (PCR). PCR was also employed for characterisation of Vibrio into three species viz V. parahaemolyticus, V. fluvialis and V. vulnificus. Antibiogram profile of the characterised species was then determined together with the presence of relevant antibiotic resistance genes Vibrio densities for the twelve month period ranged between 0 - 1.48×104 CFU/100ml with two of the plants located in East bank and Queenstown characterized by extremely high counts and one plant( Reeston) with very low counts.Three hundred presumptive Vibrio isolates were screened for identity confirmation. Of these, the dominating species found was V. fluvialis (28.6%) followed by V. vulnificus (28%) and the least was found to be V. parahaemolyticus (11.6%). The remaining unidentified 31.6% were suspected to belong to other Vibrio species not covered within the scope of this study. All the confirmed isolates i.e., V. parahaemolyticus, V. vulnificus and V. fluvialis were susceptible to imipenem, gentamicin and meropenem and resistant to only tetracycline. Between 60-100% of the V. parahaemolyticus isolates, 7.1% to 100 % V. vulnificus isolates and 2.5 to 100 % V. fluvialis showed resistances to polymixin B, sulfamethazole, erythromycin, penicillin G, chloramphenicol, trimethroprim and trimethroprim & sulfamethazole. Antibiotic Resistance Genes that were assessed included dfRA, SXT, floR and Sul2 varying in proportion with each species showing diversity in the Vibrio community. The dfR A gene was detected in all the V. parahaemolyticus isolates while floR gene was not detected in any of the isolates belonging to the three species. The distribution of sul2 gene cut across the species being 1% (1) in V. fluvialis, 3% (1) in V. parahaemolyticus and 4% (3) in V. vulnificus. The SXT gene was only determined in V. parahaemolyticus. It is clear that the final effluents of the selected plants are reservoirs for Vibrio pathogens as well as antibiotic resistance genes in the environment. The isolation of Vibrio from WWTP shows that this pathogen is in circulation in some pockets of the population. Therefore, wastewater treatment plants need to be properly monitored to ensure that they comply with set guidelines.
- Full Text:
- Date Issued: 2014