The main cause of waterborne diseases is bacteria and viruses which are commonly found in waste water and environmental water of both developing and developed countries (Cabral, 2010). Faecal material may contain various pathogens. Therefore, the more faecal material in the water means a greater risk to the people using it. The risks of a waterborne disease can be measured by the incidence of disease in the humans who have been in contact with the contaminated water (Pipes, 1982; Colford Jr et al., 2007). Although, there has been much investment to improve waste water treatment, point sources of waste water remain extremely problematic in developing countries, while waste water treatment and effluent disposal systems in developed nations can easily be overwhelmed as a result of adverse weather conditions (Kivaisi, 2001). Thus, in spite of the progress that has been achieved in waste water treatment in developed countries, while they strive to minimize the risk from bacteria, they still cannot eliminate them completely. Faecal pollution still affects beaches and coastal ecosystems all over the world (Solo-Gabriele et al., 2011).
In the UK, waste water treatment and receiving water quality is much better but there are commonly still problems on bathing beaches which are not easily explained. The result from diffuse sources such as the overflow of waste water, runoff from agricultural land, runoff from farms and urban
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environments and in situ animals such as dogs and birds which still contribute to the presence of bacteria in surface waters. Direct evidence of the presence of microbial pathogens can be obtained through monitoring processes. However, many of the pathogens are not easy to detect as they are frequently present in low numbers, but even a low concentration in polluted waters is hazardous to health for humans (Scott et al., 2004).
Epidemiological data have been used to identify pathogens of concern in a specific water body, and then used to target pathogen monitoring coupled with methods based on faecal source tracking as an additional rationale for this approach to try and maintain the water quality (Field and Samadpour 2007). Various microbiological, genotypic and phenotypic methods have previously been proposed to identify FIB for the purpose. Genetic methods can be used for differentiating between the lineages of bacteria within various animal hosts (Scott et al., 2002; 2004).
FIB have been used to detect faecal pollution based on the rationale that these indicators are indigenous to faecal material, and their presence in bathing water or the aquatic environments is indicative of faecal contamination (Goodridge et al., 2009). Microbial processes such as culture- dependent methods are now available to identify the presence of FIB in bathing water but these have problems associated with quickly changing diversity or communities of these FIB. Other methods are available to identify source(s) of bacteria such as MST methods but these are not suitable for all potential sources of faecal pollution, in particular there is no marker for some common sources of faecal contamination such as cats, goats and most wild animals. Thus the rationale for using MST methods here is to identify the
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source(s) or host of the FIB from which causes intermittent faecal pollution was derived. Hence, this study explores the development and application of analytical techniques to support improved management of contaminated bathing waters.
1.11.1 The aim of this study
Therefore, in the light of this situation, where routine pollution by human sewage is much reduced but not eliminated, the aim of this study is to evaluate, use and develop better MST techniques for the detection of faecal pollution from humans and other animals which enable determination of the presence and source of such pollution so that monitoring and management can be improved. This work has built on a previous study (Joseph, 2009) carried out to monitor bathing water quality in the Kingsbridge estuary, Devon, UK which has shown intermittent spikes of bacterial pollution and obvious bacterial pollution in water and sediment on beach in Devon. In addition, the identification of the nature and scale of the reservoirs of indicator bacteria in the sediment of some beaches and its influence on bathing water quality was studied.
1.11.2 The objectives
In order to achieve this aim, the following specific objectives were established:
i. To use and assess different methods of tracking faecal water pollution at Kingsbridge estuary, South Sands, Devon, UK, namely current culture based EU methods and PCR-based on Bacteroides spp. of human genetic marker and other origins (described in Chapter 3).
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ii. To develop new dog-specific conventional PCR and qPCR primer sets to detect dog faecal pollution in bathing water and test at a tourist bathing beach where dogs are permitted access and banned (Bigbury- on-Sea, described in Chapter 4).
iii. To investigate the survival and persistence of Bacteroides PCR genetic markers when compared to culture-dependent recovery and to study the possible adherence to sediment under controlled laboratory conditions (described in Chapter 5).
iv. To make recommendation on the application of these new methods for bacterial monitoring.
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Chapter Two
61 2.1 Introduction
The general materials and methods in this chapter were used unless otherwise indicated. This chapter will deal with the sample collection, media, culture-dependent techniques and molecular methods including DNA extraction, DNA amplification of samples using polymerase chain reaction (PCR) and quantitative PCR (qPCR). Samples of bathing water, coastal sediment and faeces were collected from Devon, United Kingdom. Culture- dependent analysis of water and sediment were conducted for isolation and enumeration of the FIB specifically E. coli, Enterococci and Bacteroides species.