Orientador: Priscila Gava Mazzola / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T03:28:14Z (GMT). No. of bitstreams: 1 Amaral_LilianFerreiraBarbosa_D.pdf: 2465793 bytes, checksum: e61ffae5669fa378935586624f7137b2 (MD5) Previous issue date: 2014 / Resumo: As indústrias cosméticas e farmacêuticas têm um crescente interesse na substituição dos antimicrobianos sintéticos nos produtos dermatológicos, devido à resistência dos microrganismos aos antimicrobianos convencionais. O pequi (Caryocar brasiliense Camb) é uma frutífera nativa do Cerrado brasileiro utilizada na medicina popular, na indústria cosmética e na alimentação, com atividades leishmanicida e antimicrobiana descritas na literatura. O objetivo principal deste trabalho foi avaliar a segurança e a eficácia do extrato de Caryocar brasilense obtido por CO2 supercrítico visando sua aplicação cosmética. A concentração inibitória mínima (CIM) frente às bactérias Escherichia coli, Pseudomonas aeruginosa e Staphylococcus aureus foi determinada pelo método clássico de microdiluição em placas. O potencial antioxidante do extrato foi determinado por um método baseado na oxidação do 2,2-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). Para avaliação da citotoxicidade e fototoxicidade in vitro, foram utilizados métodos colorimétricos baseados na conversão do corante tetrazólio (XTT) e o método do vermelho neutro (3T3 NRU), respectivamente. Na avaliação do potencial de irritação ocular empregou-se o teste na membrana corioalantóide do ovo de galinha (HET-CAM). O Perfil fitoquímico do extrato foi analisado quanto à presença de alcalóides, saponinas, antraquinonas, esteróides, taninos, flavonóides e compostos fenólicos, de acordo com métodos colorimétricos padronizados. Os resultados obtidos indicam que o extrato de Caryocar brasilense obtido por CO2 supercrítico demonstra atividade antimicrobiana frente às bactérias testadas, além de potencial antioxidante comparado ao padrão testado. Adicionalmente, o extrato de Caryocar brasilense obtido por CO2 supercrítico não apresenta efeitos tóxicos, mostrando-se um extrato seguro. Estes resultados fornecem perspectivas de desenvolvimento de produtos para o cuidado pessoal, principalmente aqueles com atividade antisséptica e os que minimizam os danos causados pelos radicais livres / Abstract: The cosmetic and pharmaceutical industries have an increasing interest in replacing synthetic antimicrobials in dermatological products due to increased microbial resistance to conventional antimicrobial agents. Caryocar brasiliense Camb (pequi) is a typical Brazilian Cerrado fruit tree. Its fruit is used as a vitamin source for culinary purposes and as a source of oil for the manufacture of cosmetics. Leishmanicidal and antimicrobial activities have been reported previously. This study was designed to evaluate the safety and efficacy of C. brasiliense extract obtained by supercritical CO2 extraction. The minimum inhibitory concentrations (MICs) against Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus were determined by the classical microdilution method. Antiseptic activity against these organisms was evaluated by the plate diffusion method. The antioxidant potential of the extract was evaluated using a method based on the oxidation of 2,2-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). In vitro cytotoxicity and phototoxicity of C. brasiliense supercritical CO2 extracts were assessed using a tetrazolium-based colorimetric assay (XTT) and Neutral Red methods; eye irritation potential was assessed using the Hen's Egg Test ¿ Chorioallantoic Membrane (HET-CAM) Test Method. The extract¿s chemical profile was analyzed for the presence of alkaloids, saponins, anthraquinones, steroids, tannins, flavonoids, and phenolic compounds according to standard colorimetric methods. The C. brasiliense supercritical CO2 extract exhibits antimicrobial activity against all bacteria tested. It also possesses antioxidant activity, when compared to a vitamin E standard. We also found that the C. brasiliense (pequi) extract obtained by supercritical CO2 extraction did not present cytotoxic and phototoxic hazards This finding suggests that C. brasiliense supercritical may be useful for the development of cosmetic and/or pharmaceutical products, primarily for antiseptic skin products that inactivate, reduce, prevent, or arrest the growth of microorganisms with the inherent intent to mitigate or prevent disease as well as products that minimize damage caused by free radicals / Doutorado / Ciencias Biomedicas / Doutora em Ciências Médicas

Wong, Kai Yi. / Thesis (M.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 101-110). / Abstracts in English and Chinese. / Abstract --- p.4 / 摘要 --- p.7 / Acknowledgements --- p.9 / List of Tables --- p.10 / List of Figures --- p.11 / List of Abbreviations and Symbols --- p.12 / Chapter Chapter 1 --- Introduction --- p.14 / Chapter 1.1 --- Staphylococcus aureus --- p.14 / Chapter 1.2 --- Methicillin-resistant Staphylococcus aureus (MRSA) --- p.14 / Chapter 1.2.1 --- Methicillin resistance of MRSA --- p.15 / Chapter 1.2.2 --- Staphylococcal Chromosomal Cassette mec (SCCmec) --- p.16 / Chapter 1.2.3 --- Hospital-associated MRSA (HA-MRSA) and Community-associated MRSA (CA-MRSA) --- p.22 / Chapter 1.2.3.1 --- Hospital-associated MRSA (HA-MRSA) --- p.23 / Chapter 1.2.3.2 --- Community-associated MRSA (CA-MRSA) --- p.23 / Chapter 1.2.4 --- Pathogenesis of MRSA infection --- p.27 / Chapter 1.2.4.1 --- Possible virulence genes contributing to necrotizing pneumonia --- p.29 / Chapter 1.2.4.1.1 --- Panton-Valentine Leukocidin (PVL) --- p.29 / Chapter 1.2.4.1.2 --- Phenol-soluble modulins (PSMs) --- p.36 / Chapter 1.3 --- Evolution of MRSA --- p.36 / Chapter 1.4 --- Epidemiology of MRSA --- p.38 / Chapter 1.4.1 --- Epidemiology of MRSA worldwide --- p.38 / Chapter 1.4.1.1 --- Epidemiology of HA-MRSA worldwide --- p.38 / Chapter 1.4.1.2 --- Epidemiology of CA-MRSA worldwide --- p.39 / Chapter 1.4.2 --- Epidemiology of MRSA in Hong Kong --- p.40 / Chapter 1.5 --- Clinical significance of MRSA --- p.41 / Chapter 1.6 --- Antibiotics --- p.43 / Chapter 1.6.1 --- Beta-lactams --- p.43 / Chapter 1.6.2 --- Fluoroquinolone --- p.44 / Chapter 1.6.3 --- Linezolid --- p.45 / Chapter 1.6.4 --- Glycopeptides --- p.45 / Chapter 1.6.5 --- Aminoglycosides --- p.46 / Chapter 1.6.6 --- Fusidic acid --- p.46 / Chapter 1.6.7 --- Clindamycin --- p.47 / Chapter 1.7 --- Hypothesis --- p.47 / Chapter Chapter 2 --- Methods and Materials --- p.50 / Chapter 2.1 --- Bacterial isolate --- p.50 / Chapter 2.2 --- Effect of subinhibitory antibiotics on the expression of mRNA in MRSA in vitro --- p.53 / Chapter 2.2.1 --- Collection of bacterial fraction --- p.53 / Chapter 2.2.2 --- RNA extraction and DNA digestion --- p.53 / Chapter 2.2.3 --- Reverse transcription for cDNA synthesis --- p.54 / Chapter 2.2.4 --- Quantitative real-time PCR (qPCR) analysis (pvl and psma\-A expression) --- p.55 / Chapter 2.2.5 --- Preparation of standard controls for quantification of DNA copy number in qPCR reactions... --- p.58 / Chapter 2.3 --- Effect of subinhibitory concentration of antibiotics on MRSA pneumonia in a murine model --- p.60 / Chapter 2.4 --- Statistical Analysis --- p.62 / Chapter Chapter 3 --- Results --- p.63 / Chapter 3.1 --- Effect of subinhibitory antibiotics on the expression ofmRNA in MRSA in vitro --- p.63 / Chapter 3.2 --- Effect of subinhibitory concentration of antibiotics on MRSA pneumonia in a murine model --- p.74 / Chapter Chapter 4 --- Discussion --- p.80 / Chapter 4.1 --- Effect of subinhibitory antibiotics on the expression of mRNA in MRSA in vitro --- p.81 / Chapter 4.2 --- Effect of subinhibitory concentration of antibiotics on MRSA pneumonia in a murine model --- p.87 / Chapter 4.3 --- Correlation of effects of subinhibitory antibiotics on the expression ofmRNA in MRSA in vitro and on MRSA pneumonia in a murine model --- p.91 / Chapter 4.4 --- Limitations of Study --- p.95 / Chapter 4.5 --- Future Work --- p.95 / Chapter Chapter 5 --- Conclusions --- p.97 / References --- p.99 / Chapter Appendix I- --- Materials and Reagents --- p.109 / Chapter Appendix II- --- Average and standard deviation of the copy number ratio (pvl or psmal-4 copy number/JdiS1 copy number) --- p.111 / Chapter Appendix III- --- In-vivo experimental data for infected control group and seven antibiotic groups --- p.116

Thesis (M.S.)--Medical College of Ohio, 1996. / Major advisor: William J. Davis. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Thesis (M.S.)--Medical College of Ohio, 1996. / Major advisor: William J. Davis. Includes bibliographical references.

Listeria monocytogenes is known to cause epidemic and sporadic cases of listeriosis. The present study investigated its occurrence, antibiotic sensitivity and serotyping of the organism in foods in various retail outlets in Gaborone, Botswana. Food samples were obtained randomly from selected supermarkets and street vendors from 5 geographical areas in Gaborone from May to September 2007. Listeria monocytogenes was isolated and positively identified by using morphological and biochemical tests. Furthermore, the organism was identified using multiplex PCR. From a total of 1324 food samples tested 57(4.3 %) were positive for Listeria monocytogenes. Out of the 57 isolates, 7 (12.3%), 3 (5.3%), 0 (0.0%), 27 (47.4%) and 20 (35.1%) were isolated from cheese, raw milk, meat (biltong), frozen cabbage and salad (coleslaw). From the 5 geographical areas selected for sampling in this study, Gaborone south recorded the most number 19 (33.3%) of L. monocytogenes isolates while Gaborone west recorded the least, 7 (12.3%). Most of the isolates (49%) belonged to serogroups 4a, 4b and 4c. These isolates were found mostly in cabbage. This was followed by serogroups 4b, 4d and 4e which comprised 30% of the isolates. This is in contrast to most studies that have found serotypes 1/2a and 1/2b to be the most common serotypes in food. That serotype 4b was detected in this study was a significant finding, because this is the number one serotype associated with human listeriosis. REP-PCR was used as a typing tool to characterize the L. monocytogenes strains. The method showed great promise as all of the L. monocytogenes strains were typable using this method, with good correlation between the REP-PCR profiles and the antibiotic resistant profiles. The findings reveal the presence of multi-drug resistant and virulent L. monocytogenes serotype 4b in ready to eat food in Gaborone, Botswana and highlight the need for education and training in food safety programmes. / Life and Consumer Sciences / M. Sc. (Microbiology (Life Sciences))

Thesis (MSc)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: The ability of biofilms to resist antimicrobial treatment, when planktonic microbes cannot, is of not only fundamental scientific interest, but also a concern in industrial and medical fields. The inability to control biofouling of water distribution networks and products, as well as recurrent infections of implanted medical devices, is not only costly, but also potentially lethal. Several mechanisms whereby biofilms are able to evade antibiotic and biocidal agents have been proposed and investigated, but no universally relevant characteristic has been identified. . Initial investigation, involving BacLightTh ! LIVEIDEAD viability probes, epifluorescence microscopy and image analysis into the ability of natural biofilm and planktonic populations, .cultured in situ in a cooling tower, to survive treatment with a commercial biocide was not conclusive. Subsequent laboratory experimentation with a bacterial isolate from the cooling tower water revealed that the ability of attached biofilms to resist antimicrobial treatment exceeded that of planktonic cells shed from the biofilm. The reduced ability of suspended cells to survive antimicrobial treatment was not statistically significant, compared to that of the biofilm (P = 0.05). This is in contrast to the wealth of literature published on the subject of biofilm antimicrobial resistance The dilution rate in the flowcells in which biofilms were cultivated was more than 100 times higher than the maximum specific growth rate of the test organism. Nevertheless, there was typically more than I x 108 cells/ml in the effluent, suggesting that a metabolically active, rapidly dividing layer of cells existed at the biofilm bulk-liquid interface, from where daughter cells continuously detached. Treatment with an antimicrobial agent resulted in a significant reduction in the viability and number of cells detached from the biofilm, suggesting that this metabolically active layer of the biofilm was more sensitive to antimicrobial treatment, possibly due to a higher specific growth rate. Antimicrobial resistance was shown to be affected by the growth rate for planktonic bacterial populations, with an increased ability to survive, correlated with a decrease in specific growth rate. This supports the contention that growth rate plays a role in the susceptibility of the active layer. The bacterial cells in the layers closest to the attachment surface of the biofilm has frequently been shown to be slow growing, due to nutrient and oxygen limitation, while the outer biofilm layer is more susceptible to unfavourable environmental conditions. It is possible that such differentiation, which results in a responsive outer biofilm layer, provides a mechanism for the protection of the cells in the deeper layers, and thus survival over time. The results presented here support several hypotheses put forth in literature to account for the increased resistance of biofilms towards antimicrobial agents. Future work will include an investigation into changes in the patterns of gene expression when a bacteria becomes attached to a surface, upon subsequent release from the biofilm, and the influence this has on the ability to resist antimicrobial treatment. / AFRIKAANSE OPSOMMING: Die vermoë van aangehegte mikrobes, in teenstelling met vrydrywende mikroorganismes, om behandeling met antimikrobiese middels te oorleef, is nie net van belang vanuit 'n fundamenteel wetenskaplike oogpunt nie, maar ook betekenisvol vir die industriële en mediese velde. Die beheer van bio-bevuiling van waterverspreidingsnetwerke en produkte, sowel as herhaalde infeksies van mediese inplantings, is nie net van kostebelang nie, maar ook potensieël lewensgevaarlik. Verskeie meganismes wat biofilms in staat stelom antimikrobiese behandeling te oorleef, IS voorgestel en ondersoek, maar geen alomteenwoordige eienskap is tot dusver geïdentifiseer nie. Aanvanklike ondersoeke na die vermoë van natuurlike biofilms en planktoniese 'gemeenskappe, om biosiedbehandeling in situ in 'n lugversorgingskoeltoring se water te oorleef, was onbeslis. Die eksperimentele metodes het gebruik gemaak van BacLight™ LIVE/DEAD lewensvatbaarheidkleurstof, epifluoressensie-mikroskopie en beeldanalise. Daaropvolgende ondersoeke met 'n bakteriese isolaat vanuit die koeltoring het daarop gedui dat biofilms beter in staat is om antimikrobiese behandeling te oorleef as selle wat vrygelaat word vanuit die biofilm. Die afname in the lewensvatbaarheid van vrydrywende selle, na afloop van biosiedbehandeling, was nie statisties beduidend in vergelyking met die van die biofilm nie (P = 0.05). Die bevinding is in teenstelling met wat algemeen aanvaar word in die literatuur. Die verdunningstempo waaronder die biofilms in die vloeiselle gekweek is, was meer as 100- voudig hoër as die maksimum spesifieke groeitempo van die toetsorganisme. Ten spyte hiervan was daar tipies meer as 1 x 108 selle/ml in die uitvloeisel teenwoordig. Dit dui op 'n metabolies aktiewe, vinnig verdelende laag selle in die boonste laag van die biofilm, naaste aan die vloeistof fase, waarvandaan dogterselle voortdurend vrygestel word. Behandeling met die antimikrobiese agent het 'n beduidende afname in die lewensvatbaarheid en aantal dogterselle tot gevolg gehad, wat lei tot die gevolgtrekking dat die metabolies aktiewe laag van die biofilm meer sensitief is vir antimikrobiese behandeling, moontlik weens 'n hoër spesifieke groeitempo. Daar is verder bewys dat die vermoë om die werking van die antimikrobiese middel teen te staan, afhanklik is van die spesifieke groeitempo van planktoniese populasies. 'n Afname in groeitempo word geassosieer met 'n toename in oorlewing na antimikrobiese behandeling, wat die voorstel dat die groeitempo van die aktiewe laag 'n rol speel in die vatbaarheid daarvan, ondersteun. Dit is bekend dat die metaboliese aktiwiteit van bakteriese selle nader aan die aanhegtingsoppervlak van die biofilm verlaag is, weens 'n afname in diffusie van suurstof en nutriente in daardie deel van die biofilm. Dit is moontlik dat hierdie differensiasie, wat lei tot die vatbaarheid van die buitenste laag van die biofilm vir ongunstige omgewingstoestande, 'n oorlewingsmeganisme daarstel wat die onderliggende selle beskerm. Die resultate wat hier voorgelê word, ondersteun verskeie hipoteses wat die verhoogde weerstandbiedendheid van biofilms teen antimikrobiese middels beskryf. Toekomstige werk sluit ondersoeke in na veranderende patrone van geenuitdrukking wat plaasvind wanneer 'n bakterie in aanraking kom met 'n oppervlak, vasheg en ook weer vrygestel word, asook die invloed hiervan op die vermoë om antimikrobiese behandeling te oorleef.

Thesis (MScEng (Process Engineering))--University of Stellenbosch, 2009. / Surfactants are compounds that reduce interfacial surface tension, resulting in detergency, emulsifying, foaming and dispersing properties. Surfactants produced via biochemical processes (biosurfactants) form a niche market with their low toxicity, biodegradability and high specificity attributes. Biosurfactants have recently received considerable attention owing to their potential as biomedical molecules. In this study a knowledge base was established for the development of a process which produces biosurfactants for use as antimicrobial agents. Specifically, rhamnolipid biosurfactants were produced from Pseudomonas aeruginosa and tested for antimicrobial activity against target organisms. Accurate and reproducible analyses for the quantification of rhamnolipids and antimicrobial activity were developed. The amount of rhamnolipid was determined indirectly by measuring the rhamnose concentration. A novel HPLC method as well as an orcinol colorimetric method were developed for rhamnose measurement. In order to obtain accuracy with the orcinol method it was found that samples must be extracted at least three times prior to the analysis. An examination of literature on rhamnolipid production showed that many studies used colorimetric methods without extraction. Antibacterial activity was quantified by zone clearing around wells of supernatant in soft agar containing the target organism Mycobacterium aurum. This target organism is especially important in a South African context, since it is used to indicate possible susceptibility of tuberculosis to antibiotics. This method was developed for antibacterial testing, after a standard disk diffusion method proved to be ineffective. Antifungal activity of rhamnolipids was evaluated against the fungus Botrytis cinerea, by growing a lawn of fungus on a plate and adding rhamnolipid. The factors influencing rhamnolipid production were studied by growing different Pseudomonas aeruginosa strains from the ATCC culture collection, namely ATCC 9027 and ATCC 27853 as well as a locally isolated strain under different media conditions. The initial focus was on production of biosurfactants in media containing glucose as substrate. Alkanes were subsequently investigated as an alternative substrate, since they are readily available in South Africa as byproducts from the petrochemical industry. The rhamnolipids produced from the culture collection strains were evaluated for their antibacterial activity against Mycobacterium aurum. A number of key factors were identified which were important for the development of a rhamnolipid production process. Of critical importance were the media conditions. Good production was achieved on glucose media containing a phosphate limitation, pH buffering around neutral pH and a high carbon concentration (2 % carbon). When Pseudomonas aeruginosa ATCC 9027 was cultured on this medium (a minimal salts phosphate limited medium with a Tris buffer), it produced 1.31 g/l rhamnose, equivalent to 4.0 g/l rhamnolipid. This rhamnolipid concentration is 2.7-fold higher that of 1.47 g/l reported in the literature with the same strain (cultured on a different phosphate limited medium The particular strain also proved to be a factor which influenced the yield of rhamnolipids. A rhamnose concentration of 0.43 g/l was obtained with Pseudomonas aeruginosa ATCC 27853 grown on MSM+Tris medium, compared to 1.31 g/l produced by Pseudomonas aeruginosa ATCC 9027 on the same medium. The most promising strain and medium, Pseudomonas aeruginosa ATCC 9027 and MSM+Tris medium, were evaluated under controlled conditions in an instrumented bioreactor. Nearly double the rate of growth and production were obtained in the bioreactor, indicating that production time can be shortened considerably under controlled conditions. However, when compared to shake flask studies, only a 4 % increase in growth and a 5 % increase in rhamnolipid production were achieved in the bioreactor, indicating that the yield was limited by the media components or process conditions. With media containing hexadecane as sole carbon source, negligible rhamnolipid production was achieved. Slow growth was observed and the stationary phase had not been reached even after 2 weeks of growth. It was shown that in glucose media rhamnolipid production only commenced in the stationary phase. Since the stationary phase was not reached during growth on hexadecane, rhamnolipids, which are known to increase the availability of alkanes through emulsification and solubilisation, could not be produced. A strategy was devised to accelerate growth on alkane media. A dual substrate medium containing both glucose and hexadecane was investigated. It was hypothesised that growth would be promoted by glucose leading to rhamnolipid production, which would then increase the uptake of hexadecane. Rhamnolipid was produced in the dual substrate experiments, but the hexadecane uptake was still poor. This was suggested to be due to the exposure of the cells to glucose in the inoculum or test flask, which hampered the ability of the cells to utilise hexadecane. It was reasoned that the ability to utilise hexadecane was determined by the cell hydrophobicity, which was influenced by the exposure to hydrophilic or hydrophobic substrates. Rhamnolipids from Pseudomonas aeruginosa ATCC 9027 and ATCC 27853 were shown to have antibacterial activity against Mycobacterium aurum. The largest zone of clearing of 45 mm was obtained with 4 g/l rhamnolipid from Pseudomonas aeruginosa ATCC 9027. The activity was shown to be directly related to the rhamnolipid concentration, highlighting the importance of maximising the biosurfactant yield when developing a process for the production of rhamnolipids as antimicrobial agents. Antifungal activity tests against Botrytis cinerea were inconclusive. Future studies should expand the antimicrobial application of rhamnolipids by testing their activity against a larger range of target organisms. In order to maximise the rhamnolipid yield in future studies, a fed batch process is proposed which would increase the cell density thereby increasing rhamnolipid production and prolonging the stationary phase, which was found to be the phase associated with rhamnolipid production. Different feeding strategies should be investigated, depending on the kinetics of substrate consumption. It is desirable to feed the smallest volume of substrate that is necessary with a high concentration in order to keep the dilution rate low and maximise the product concentration. A factorial design is recommended for this purpose. Further studies with alkanes as carbon source should be conducted using strains that have been maintained and cultured on media containing alkanes as sole carbon source. Alternative biosurfactant producing strains should also be investigated, which have higher natural cell hydrophobicities.

ESF1 is a 2.052 kDa antimicrobial peptide, mimicking the charge distribution and amphipathic alpha-helical structure of magainin, pGLa, a naturally occurring antimicrobial peptide. ESF1 has been reported to display high activity against Fusarium oxysporum f. sp lycopersici and F. oxysporum f. sp cubense race 4, the tomato and banana crop plant, wilt-causing pathogens, respectively. To assess whether this synthetic peptide can be heterologously expressed in yeast in significant quantities, and still retain full bioactivity, within a eukaryotic system, the ESF1 gene was designed and synthesized from five oligonucleotides, and cloned into pUC18. From the pUC18/ESF1 recombinant plasmid, the ESF1 gene sequence was amplified and cloned into the pBluescript-based vector, pVD4, downstream of the yeast pheromone mating factor alpha (MFa1) promoter, and in frame with the MFa1 signal peptide sequence for expression and secretion in yeast. The expression cassette comprising the MFa1 promoter and signal peptide sequence, and ESF1 gene was subsequently cloned into the yeast/ E. coli shuttle vector, pTG3828 and transformed into Saccharomyces cerevisiae. Chicken IgY antibodies against ESF1 peptide were raised and immunoaffinity purified. Following this, western dot blot analysis and mass spectrometry confirmed the presence of ESF1 in partial HPLC purified fractions of the recombinant yeast culture supernatant. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 2000.

Mini-dissertation submitted in partial compliance with the requirements of the Master's Degree in Technology: Homoeopathy, Durban Institute of Technology, 2003. / The aim of this study was to establish the efficacy of Withania somnifera in tincture, 1X and 6X homoeopathic dilutions (in 62% v/v ethanol) as an antimicrobial agent against the in vitro growth of Bacillus cereus, Pseudomonas aeruginosa, Klebsiella pneumonia, Escherichia coli and Staphylococcus aureus, as compared to 62% v/v ethanol only. The disc diffusion method was employed. W somnifera is indigenous to southern Africa and its use is well established amongst the traditional healers for many varied complaints. Infusions, decoctions and tinctures of the fresh and dry whole root are used. (Gericke and Van Wyk, 2000:150.) For this study 20 plates of Mueller-Hinton agar were inoculated with each bacteria, resulting in a total of 100 plates. Four dry discs previously impregnated with the test substances and two antibiotic discs were equidistantly placed on each plate and incubated at 3rC. The vancomycin and gentamycin discs were included to account for plate-to-plate variations in the sensitivity of the bacteria to the antimicrobial substances. The plates were observed at 18, 24 and 48-hour intervals. ' Statistical analysis was performed using the Friedman test to compare test and control substances at each observation interval. The Mann-Whitney-U test was used to compare the mean inhibition zones between test and control substances / M

Listeria monocytogenes is known to cause epidemic and sporadic cases of listeriosis. The present study investigated its occurrence, antibiotic sensitivity and serotyping of the organism in foods in various retail outlets in Gaborone, Botswana. Food samples were obtained randomly from selected supermarkets and street vendors from 5 geographical areas in Gaborone from May to September 2007. Listeria monocytogenes was isolated and positively identified by using morphological and biochemical tests. Furthermore, the organism was identified using multiplex PCR. From a total of 1324 food samples tested 57(4.3 %) were positive for Listeria monocytogenes. Out of the 57 isolates, 7 (12.3%), 3 (5.3%), 0 (0.0%), 27 (47.4%) and 20 (35.1%) were isolated from cheese, raw milk, meat (biltong), frozen cabbage and salad (coleslaw). From the 5 geographical areas selected for sampling in this study, Gaborone south recorded the most number 19 (33.3%) of L. monocytogenes isolates while Gaborone west recorded the least, 7 (12.3%). Most of the isolates (49%) belonged to serogroups 4a, 4b and 4c. These isolates were found mostly in cabbage. This was followed by serogroups 4b, 4d and 4e which comprised 30% of the isolates. This is in contrast to most studies that have found serotypes 1/2a and 1/2b to be the most common serotypes in food. That serotype 4b was detected in this study was a significant finding, because this is the number one serotype associated with human listeriosis. REP-PCR was used as a typing tool to characterize the L. monocytogenes strains. The method showed great promise as all of the L. monocytogenes strains were typable using this method, with good correlation between the REP-PCR profiles and the antibiotic resistant profiles. The findings reveal the presence of multi-drug resistant and virulent L. monocytogenes serotype 4b in ready to eat food in Gaborone, Botswana and highlight the need for education and training in food safety programmes. / Life and Consumer Sciences / M. Sc. (Microbiology (Life Sciences))