ISOLATION AND IDENTIFICATION OF PATHOGENIC BACTERIA FROM DRINKING WATER OF KHAIRPUR, SUKKUR AND ROHRI

Abstract

Little information is available on water borne diseases in Pakistan. This probably is due to absence of an infrastructure for detection and recording such infection and its source. There is no reason to believe that the risks of water born diseases are any different from those in the rest of the world. This possibility is supported by data presented in this study which show correlations of enteric infections in various cities to address the condition of sanitation, standards of living and education. The data reflects the incidence and public health impact of these diseases in the area under study. This research project describes the fecal indicator bacteria, different waterborne bacterial pathogens, pH, temperature and chlorine residual in municipal water intended for drinking and other domestic uses from Khairpur Sukkur and Rohri Sindh Pakistan. The work was done by collecting water samples twice in a month for three years of study period from 2005-2007. Drinking water microbiological quality was primarily determined by enumeration of “indicator organisms”, whose presence indicates faecal contamination. The presence of the indicators is often a key in assessing potential public health risks due to bacterial pathogens and is used in drinking water quality regulations and guidelines in many countries. The enumeration of fecal coliform specifically of Escherichia coli (indicator of fecal contamination) from human as well as animal sources per 100 ml of municipal water used for drinking purposes has been recommended by World Health Organization (WHO) worldwide to monitor the quality of drinking water. According to World Health Organization (WHO) guideline standards for drinking water total and fecal coliform the indicator of fecal contamination must not be detectable in any 100 ml samples. In present study the enumeration of fecal coliform bacteria by membrane filtration method revealed that the number of fecal coliform (E. coli) in water was higher than standard set by World Health Organization (WHO) and Pakistan. The number of fecal coliform (E. coli) was varying in different seasons. In winter months it was 2.69-3.17 log cfu/100 ml and in summer months it was 3.30-3.47 in more than 70% drinking water samples collected from municipal water of Khairpur. The drinking water samples collected from Sukkur were found with highest number of faecal coliform bacteria ranging from 3.30-3.45 log cfu/100 ml in winter months to 3.60- 3.84 log cfu/ 100 ml summer months in 75% water samples, and in the drinking water collected from Rohri the number of fecal coliform bacteria were also higher in summer months. In winter months the number of fecal coliform was 2.0 3.30 log cfu/ 100 ml and in summer months the number of fecal coliform was 3.39-3.45 log cfu/100 ml in water samples. The presence of fecal indicator bacteria (E. coli) in higher number in more than 75% water samples investigated was the indicative of possible presence of etiologic agents of other waterborne bacterial infectious diseases. The enumeration of total and fecal coliform bacteria showed that the municipal water of area under study was of inferior quality. The water sample tested were contained thousand of total and fecal coliform bacteria per 100 ml. the results of study suggested that the level of fecal contamination in municipal water was very high. The health impact of that unsafe drinking water could be the prevalence of waterborne diarrhoeal diseases in the population. A moderate correlation was observed between the Densities of fecal coliform and diarrhoeal cases in the in the area under study. Such water with a high number of total and fecal coliform could be the potential sources of waterborne bacterial pathogens. As it is evident from the study of 216 drinking water samples 12 species from 8 genera were isolated and identified. In this research different waterborne bacterial pathogens were isolated and identified with varying frequency by using the membrane filtration technique and Analytical Profile Index system for Enterobacteriaceae (API 20E). As the membrane filter technique (MFT) is highly reproducible, it can be used to test large volumes of samples and yield numerical results more rapidly than the multiple tube procedure; this technique was extremely useful in monitoring drinking water quality. In 1950s the membrane filter technique came into practice as a substitute for MPN method. The membrane filter technique is highly suitable for drinking water and different natural waters quality assessment. The API 20E system is considered the gold standard for identification of Enterobacteria. It has been reported in many studies that API 20 is a good method in its sensitivity with genus level identification (92%) and at the species level (72%) while working on genus Yesinia. The likelihood values from 62.5% to 99% of Salmonella enterica by API 20E has also been reported. In present study the species from family Enterobacteriaceae, Vibrionaceae, Aeromonadaceae, Pseudomondaceae were isolated. Some opportunistic bacteria like C. meningosepticum and Non- Fermenter species were also isolated from drinking water of three cities. In drinking water of Khairpur city the percentage of isolation rate was i.e. E. coli, 64%, P. mirabilis, 67% P. rettgeri 60%, P. stuarti 67%, C youngae 64%, K. oxytoca 49%, V. cholerae 29.16% and V. mimicus 47.22%, A. hydrophila 53%, P. aeruginosa 69.44%, C. meningosepticum 57% and Non- Fermenter species 61% were isolated. In drinking water of Sukkur city the E. coli, 69.44%, P. mirabilis, 65.28% P. rettgeri 71%, P. stuarti 61%, C. youngae 60%, K. oxytoca 54.16%, V. cholerae 22.22% and V. mimicus 50%, A. hydrophila 50%, P. aeruginosa 76%, C. meningosepticum 51.38% and Non- Fermenter species 57% were isolated. In drinking water of Rohri the E. coli, 76.38%, P. mirabilis, 68% P. rettgeri 69.44%, P. stuarti 67%, C youngae 69%, K. oxytoca 44.44%, V. cholerae 32% and V. mimicus 54.16%, A. hydrophila 51.33%, P. aeruginosa 78%, C. meningosepticum 50% and Non- Fermenter species 60% were isolated. In this study some new bacterial pathogens was isolated and identified from water samples tested for example C. meningosepticum P. rettgeri P. stuarti, C. youngae. These bacteria are not included in the list of bacterial pathogens of water previously. These findings could be valuable or a breakthrough in the inclusion of these species as waterborne pathogens. The isolation of waterborne bacterial pathogens at such a high percentage is of a grave concern, because contaminated water plays crucial role in the prevalence of waterborne and water related bacterial diseases outbreaks in the urban and rural population in Khairpur, Sukkur and Rohri. It is important to note that the municipal water of Khairpur and other cities were found contaminated with multi-drug resistant thermo-tolerant E. coli. The diseases i.e. Diarrhea, Dysentery, Gastroenteritis, Typhoid fever and Cholera and other water related nosocomial infection in health care setting may be the result of consumption of such polluted water with. The temperature of water samples was also determined to observe the seasonal effect on the bacterial population in fresh water environments. It was observed that when the temperature of water ranged from 25- 32oC the isolation rate of bacterial species in drinking water was 68-77% and when the temperature of water samples ranged from 17-22oC the isolation rate was 34- 42.61%. The temperature plays an important role in the microbial growth in biofilms where bacteria and other organism colonize, establish themselves and then produce permanent source of contamination in water distribution systems. The pH of water samples was also measured because the pH imparts its effect on water quality. Our result indicates that pH was in limits (6.5- 8.5) of WHO guideline standard for drinking water. When the residual chlorine in drinking water samples was determined, a very high number of drinking water samples were found without residual chlorine. In water samples collected from Khairpur, more than 80% of water samples were found without chlorine. More than 90 % water samples from drinking water of Sukkur were found without residual chlorine and also the same percentage of water samples was found without chlorine in city of Rohri Sindh Pakistan. It is evident from these results that the bacteriological quality of drinking water was not according to the standard of World Health Organization guidelines; for drinking water. The presence of high rate of waterborne pathogens in our study may be due to this reason. The disinfection treatment of drinking water has important role in reducing the waterborne epidemics. The inadequate disinfection treatment of municipal water results in the provision of unsafe drinking water to the people which can pose a great threat and risk of waterborne epidemics by bacterial pathogens to the population consuming it. Such negligence may result in a catastrophic disaster in the area. Water supplying authorities should take account of this situation and take measures for the provision of contamination free drinking water to prevent waterborne disease outbreaks by bacterial pathogens.