The spread of antibiotic-resistant microorganisms in the environment is recognized widely as an important public health issue, with concerns about future ability to treat infectious diseases. The main risk to public health is that the resistance genes are transferred from environmental bacteria to human pathogens. Safe water is one of the most important needs in public health in the twenty first century. The major health threat posed by drinking unsafe water is the transmission of infectious diseases, which are the leading causes of mortality and morbidity for children under the age of 5 and it is estimated to cause 1.5 million deaths annually in developing countries. In addition to the wide spread cases of water-borne diseases resulting from the contamination of water sources, concerns have been raised when these diseases fail to be cured due to development of resistance to most prescribed antibiotics by the contaminating microorganisms. It is now a well-established fact that E.coli is a significant cause of diarrheal illnesses both in infants and adults in many parts of the world. Data on clinical isolates is plenty while less attention has been given to environmental isolates of these enteric pathogens. Samples from the environment such as water may serve as probable reservoirs of these pathogens; this is compounded by the entry of functional compounds of antibiotics into waterways, through humans and animals that have ingested antibiotics. This is because antibiotics are not completely metabolized and may enter waterways through the waste products of these humans or animals.Studies on antimicrobial resistance is important in order to detect changes in patterns of resistance, implement control measures on the use of antimicrobial agents, and to prevent the spread of multidrug-resistant strains of bacteria. It also provides surveillance data for antibiotic resistances, necessary to define or update guidelines for empirical treatment, as well as a guide for appropriate antibiotic supplies. Study objectives: The objectives of this research were: (i) to determine the total and faecal coliform status of drinking water sources, as an indication of quality; (ii) to determine the bacteriological profile of bacteria flora in the drinking water sources; (iii) to determine prevalence and susceptibility profiles of antibiotic resistant water-borne E.coli; (iv) to investigate the virulence genes associated with multiple antibiotic resistant E. coli isolates; (v) to compare three laboratory based techniques: PCR, API 20E, and Culture based methods used for detection of E.coli and (vi) to determine the association between multiple antibiotic resistance and radiation sensitivity (D10). © University of South Africa 2014 VII Methodology: Four hundred and sixty four (464) water samples were collected for assessment between June 2011 and May 2012. The samples were collected from 57 sampling sites, from six different water sources including: boreholes (10), a canal (1), dams (15), hand-dug wells (15), a river (1), and streams (15). Total coliforms, faecal coliforms, and E. coli analysis were done by the MPN method. Bacteria isolation and identification were done using API 20E, conventional methods, and a PCR based DNA STRIP technology that allows simultaneous detection of virulence genes and confirmation of E. coli isolates. Antibiotic susceptibility testing was also conducted using the Kirby-bauer method. Radiation sensitivity was done using a cobalt 60 source. Results: The results obtained indicated that all the water sources were of poor quality in terms of microbial distributions with total coliform and faecal coliform counts ranging between 0 to 2.4x103 MPN/100ml. E. coli counts ranged between 10 to 7.9x101MPN/100ml. Disease risk assessment of the various water sources indicated that dam water sources presented a high disease risk, while borehole water sources had a low disease risk. A total of five hundred and twenty bacterial isolates (520) were obtained during the period of study. Three hundred and five (305) isolates representing 58.65% of the total were obtained during the dry season, as against (205) representing 41.35% in the rainy season. The most commonly occurring bacteria in the water samples was Klebsiella spp constituting 20%. The next most occurring organism was E. coli (18.7%). This was followed by Pseudomonas aeruginosa (15.61%), Enterobacter spp. (15.4%), Proteus vulgaris (13.1%), and Enterococcus faecalis (9.2%). The least isolated bacteria were Vibrio cholerae (1.2%) and Shigella spp. (1.2%). The prevalence of multi drug resistance E. coli was 49.48 %. E. coli isolates showed a high sistance patterns to the tested antibiotics. They were most resistant to penicillin (32.99%), cefuroxime (28.87%), erythromycin (23.71%), and tetracycline (21.45%). In contrast, they were susceptible to nitrofurantoin (93.8%), cefotaxime and amikacin (91.75%), gentamicin (90.7%), nalidixic acid (89.65%), ciprofloxacin (74.2%), chloramphenicol (69.07%), pipemidic acid (65.97%) and cefuroxime (52.58%). Sixty-three percent (63%) of the multidrug resistant E. coli strains recorded a multiple antibiotic resistance (MAR) index value of >0.2. Six (6%) percent of he multiple antibiotic resistant were eae virulence genes producing however, none of the E. coli isolates produced the stx1 and stx2 virulent gene. The analytical profile index (API) recorded specificity and sensitivity of 99.7% and 98.50 % respectively for the detection of E. coli. The © University of South Africa 2014 VIII culture/ biochemical based methods for detection of E coli recorded specificity of 81.82% and a sensitivity of 96.91%. There was no association (P> 0.05) between radiation sensitivity (D10) and antibiotic resistances. Conclusion: The study has confirmed that majority of the water sources used for drinking and domestic purposes in the study area are highly contaminated with high levels of faecal coliforms above the recommended standards. There were also resence of bacteria of public health importance in the water sources. Both animals and humans could be sources of faecal bacteria contamination of the drinking water sources. The study confirmed a high prevalence of multiple antibiotic resistances in E. coli isolates. The eae virulence gene was associated with some of the multiple resistant E. coli isolates. The study also concludes that API 20E has a high specificity and sensitivity close to that of the PCR. Lastly, There is no association between multiple antibiotic resistant indexes and radiation sensitivity (D10) of antibiotic resistant E. coli. / School of Environmental Sciences / D. Phil. (Environmental Science)

The spread of antibiotic-resistant microorganisms in the environment is recognized widely as an important public health issue, with concerns about future ability to treat infectious diseases. The main risk to public health is that the resistance genes are transferred from environmental bacteria to human pathogens. Safe water is one of the most important needs in public health in the twenty first century. The major health threat posed by drinking unsafe water is the transmission of infectious diseases, which are the leading causes of mortality and morbidity for children under the age of 5 and it is estimated to cause 1.5 million deaths annually in developing countries. In addition to the wide spread cases of water-borne diseases resulting from the contamination of water sources, concerns have been raised when these diseases fail to be cured due to development of resistance to most prescribed antibiotics by the contaminating microorganisms. It is now a well-established fact that E.coli is a significant cause of diarrheal illnesses both in infants and adults in many parts of the world. Data on clinical isolates is plenty while less attention has been given to environmental isolates of these enteric pathogens. Samples from the environment such as water may serve as probable reservoirs of these pathogens; this is compounded by the entry of functional compounds of antibiotics into waterways, through humans and animals that have ingested antibiotics. This is because antibiotics are not completely metabolized and may enter waterways through the waste products of these humans or animals.Studies on antimicrobial resistance is important in order to detect changes in patterns of resistance, implement control measures on the use of antimicrobial agents, and to prevent the spread of multidrug-resistant strains of bacteria. It also provides surveillance data for antibiotic resistances, necessary to define or update guidelines for empirical treatment, as well as a guide for appropriate antibiotic supplies. Study objectives: The objectives of this research were: (i) to determine the total and faecal coliform status of drinking water sources, as an indication of quality; (ii) to determine the bacteriological profile of bacteria flora in the drinking water sources; (iii) to determine prevalence and susceptibility profiles of antibiotic resistant water-borne E.coli; (iv) to investigate the virulence genes associated with multiple antibiotic resistant E. coli isolates; (v) to compare three laboratory based techniques: PCR, API 20E, and Culture based methods used for detection of E.coli and (vi) to determine the association between multiple antibiotic resistance and radiation sensitivity (D10). © University of South Africa 2014 VII Methodology: Four hundred and sixty four (464) water samples were collected for assessment between June 2011 and May 2012. The samples were collected from 57 sampling sites, from six different water sources including: boreholes (10), a canal (1), dams (15), hand-dug wells (15), a river (1), and streams (15). Total coliforms, faecal coliforms, and E. coli analysis were done by the MPN method. Bacteria isolation and identification were done using API 20E, conventional methods, and a PCR based DNA STRIP technology that allows simultaneous detection of virulence genes and confirmation of E. coli isolates. Antibiotic susceptibility testing was also conducted using the Kirby-bauer method. Radiation sensitivity was done using a cobalt 60 source. Results: The results obtained indicated that all the water sources were of poor quality in terms of microbial distributions with total coliform and faecal coliform counts ranging between 0 to 2.4x103 MPN/100ml. E. coli counts ranged between 10 to 7.9x101MPN/100ml. Disease risk assessment of the various water sources indicated that dam water sources presented a high disease risk, while borehole water sources had a low disease risk. A total of five hundred and twenty bacterial isolates (520) were obtained during the period of study. Three hundred and five (305) isolates representing 58.65% of the total were obtained during the dry season, as against (205) representing 41.35% in the rainy season. The most commonly occurring bacteria in the water samples was Klebsiella spp constituting 20%. The next most occurring organism was E. coli (18.7%). This was followed by Pseudomonas aeruginosa (15.61%), Enterobacter spp. (15.4%), Proteus vulgaris (13.1%), and Enterococcus faecalis (9.2%). The least isolated bacteria were Vibrio cholerae (1.2%) and Shigella spp. (1.2%). The prevalence of multi drug resistance E. coli was 49.48 %. E. coli isolates showed a high sistance patterns to the tested antibiotics. They were most resistant to penicillin (32.99%), cefuroxime (28.87%), erythromycin (23.71%), and tetracycline (21.45%). In contrast, they were susceptible to nitrofurantoin (93.8%), cefotaxime and amikacin (91.75%), gentamicin (90.7%), nalidixic acid (89.65%), ciprofloxacin (74.2%), chloramphenicol (69.07%), pipemidic acid (65.97%) and cefuroxime (52.58%). Sixty-three percent (63%) of the multidrug resistant E. coli strains recorded a multiple antibiotic resistance (MAR) index value of >0.2. Six (6%) percent of he multiple antibiotic resistant were eae virulence genes producing however, none of the E. coli isolates produced the stx1 and stx2 virulent gene. The analytical profile index (API) recorded specificity and sensitivity of 99.7% and 98.50 % respectively for the detection of E. coli. The © University of South Africa 2014 VIII culture/ biochemical based methods for detection of E coli recorded specificity of 81.82% and a sensitivity of 96.91%. There was no association (P> 0.05) between radiation sensitivity (D10) and antibiotic resistances. Conclusion: The study has confirmed that majority of the water sources used for drinking and domestic purposes in the study area are highly contaminated with high levels of faecal coliforms above the recommended standards. There were also resence of bacteria of public health importance in the water sources. Both animals and humans could be sources of faecal bacteria contamination of the drinking water sources. The study confirmed a high prevalence of multiple antibiotic resistances in E. coli isolates. The eae virulence gene was associated with some of the multiple resistant E. coli isolates. The study also concludes that API 20E has a high specificity and sensitivity close to that of the PCR. Lastly, There is no association between multiple antibiotic resistant indexes and radiation sensitivity (D10) of antibiotic resistant E. coli. / School of Environmental Sciences / D. Phil. (Environmental Science)