24 1.6.6 Vibrio sp.
Vibrio sp. are non-spore forming, motile gram negative rods with a single flagellum defined as being facultative anaerobes and oxidase positive. They are frequently found in a range of water sources, usually transmitted via the faecal–oral route with infections generally caused by the ingestion of faecal-contaminated water and food (Cabral, 2010). There are a number of pathogenic species, including V. cholerae, V. parahaemolyticus and V. vulnificus with V.
cholerae being the major pathogen of concern in freshwater environments. Whilst non- toxigenic V. cholerae is widely distributed in water environments, toxigenic strains are not distributed as widely with only the O1 and O139 serovars known to cause the related-cholera symptoms due to the production of the cholera endotoxin. Infected individuals are known to have alterations in ionic fluxes across the intestinal mucosa leading to severe water and electrolyte loss (WHO, 2011b; 2002).
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viruses, thus allowing these coliphages to serve as suitable indicators of viral contamination (DWAF, 1996b). Structural and morphological analysis indicate the presence of a nucleic acid molecule, more specifically the genome, which is surrounded by a protein coat also known as the capsid. Generally, somatic coliphages have been found to outnumber F-RNA coliphages in water environments by a factor of approximately 5 and cytopathogenic human viruses by a factor of about 500 (WHO, 2011b). Contaminant source identification together with their physical structure, composition and morphology have allowed phages to serve as a means to distinguish between faecal pollution of human and animal origin and is an ideal indicator of the presence of human enteric viruses.
1.7.1 Somatic coliphages
Somatic coliphages are DNA viruses and consist of a wide range of phages belonging to members of the families - Myoviridae, Siphoviridae, Podoviridae and Microviridae with a vast range of morphological types (WHO, 2008). They replicate more frequently in the gastrointestinal tract of warm-blooded animals with a population range of less than 10 to 108 plaque-forming units per gram. They have also been known to replicate in water environments and are easily detectable by simple and inexpensive plaque assays (Grabow, 2001). These phages possess the ability to infect hosts such as E. coli and other closely related members of the Enterobacteriaceae family by attaching to receptors permanently located on the cell wall of hosts. These phages tend to initiate infection of the respective E.
coli host by adsorbing to receptors located in the host cell wall (DWAF, 1996b).
26 1.7.2 Male-specific F-RNA coliphages
F-RNA coliphages are defined as single stranded RNA phages that are morphologically similar to that of picornaviruses. These phages possess an icosahedral capsid and belong to the family Leviviridae which have further been divided into four serological groups allowing for source contamination identification. Male-specific F-RNA coliphages have been classified into four main groups based on their serological and physiochemical properties, namely; MS-2, f2, and JP501 in Group 1; GA, BZ13 and JP34 in Group 2; Qβ, VK and TW18 in Group 3 and SP, F1 and TW28 in Group 4 with Groups 2 and 3 being associated with human faecal contamination and Groups 1 and 4, associated with animal contamination (Sundram et al., 2005). These phages initiate infection in hosts that possess the fertility (F+) plasmid and which produce the required fertility fimbriae. Of importance is the production of these fimbriae, which are only produced when hosts are in the exponential phase and at temperatures greater than 30 ºC, indicating the restriction of these hosts to grow within the gastrointestinal tract thus serving as a more reliable indicator of faecal contamination (WHO, 2008). As a result of their mode of replication and host specificity, F-RNA coliphages are generally excreted by a lower percentage of humans and animals compared to somatic coliphages. Schaper et al. (2002) confirmed this with F-specific RNA phages being detected in 10% of human, 45% of bovine and 60% of porcine faecal samples. In addition, detection methods are not as simple as that for somatic coliphages due to specific requirements of the host bacterium which needs to be grown at temperatures greater than 30 ºC and in the log phase to ensure that fertility fimbriae to which the phage attaches are present, thus indicating the need for timeous preparation of host cultures (Grabow, 2001; WHO, 2011b). Also, numerous studies have revealed that F-RNA phage counts outnumber enteric viruses by a factor of approximately 100 in wastewater and raw water sources. However, one of the major
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disadvantages is that currently utilised F-RNA coliphage detection schemes are not as simple due to the requirement of adequate host preparation prior to detection.
1.7.3 Phages that infect Bacteroides fragilis
Considerable attention has been given to Bacteroides fragilis as an indicator of water quality and disinfection processes as they are found solely in human faeces and they also show resistance to a range of chlorine disinfection steps when compared to other pathogens, such as poliovirus, rotavirus, certain coliphages, E. coli and Streptococcus faecalis. Two groups of B. fragilis phages are used as indicators in water quality assessments. They differ by their respective bacterial host strains, Bacteroides HSP40 and RYC2056 which inhabit the gasterointestinal tract. The first group belongs to the family Siphoviridae and are restricted solely to the human gasterointestinal tract and hence their detection within the environment provides a strong indication of human faecal contamination. The second group however includes a wider range of phages which are detected in both human and animal faeces (WHO, 2008). However, as with all indicators, one of the major disadvantages is that HSP40 phages are excreted by approximately 10 – 20% of humans in certain parts of the world and hence occur in relatively low numbers in sewage, polluted water environments and drinking water sources, indicating that their absence does not confirm the absence of other pathogens such as enteric viruses. In addition, detection methods have proven to be more complex and expensive than those for somatic and F-RNA coliphages due to the need for antibiotics and anaerobic environments (Grabow, 2001).
28 1.8 Current guidelines for treated effluent
Guidelines range from providing required public advice, numerical data as well as a respective classification system. Various guidelines and water quality criteria have been set in both local and international committees, however, due to the vast differences in methodology and development; these values tend to differ greatly. Whilst some guidelines tend to exhibit the maximum concentration of a particular contaminant others attempt to define the ideal concentration thus leading to confusion. In addition, these guidelines need to be flexible and adaptable to suit local, regional and national scenarios by taking the current socio-economic and environmental conditions into consideration (WHO, 2003a). This should be followed by subsequent transposition of guidelines into legally enforceable national standards by Government. Despite an outdated guideline for treated effluent being discharged into any catchment or river (Table 1.4), a South African Green Drop Certification Program was recently started by the Department of Water Affairs in an attempt to regularly monitor and improve the wastewater sector. This program allows local municipalities to generate information from data pertaining to their treatment plant efficiency and effluent characteristics, in order to monitor and report back, regarding their wastewater management systems. In addition, it provides the respective water regulators with an overview of required information allowing for improved trend monitoring and decision making as well as providing the public with access to relevant information regarding their regions (DWA, 2011).
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Table 1.4: Currently used guidelines by the eThekwini Municipality (South Africa) for treated effluent being discharged into a receiving catchment
PARAMETER A B
Colour/ Odour/ Taste None None
pH 5.5 – 7.5 5.5 - 9.5
Dissolved Oxygen (mg/l) 75% saturation 75% saturation
Faecal Coliforms (CFU/100 ml) 0 0
Temperature (°C) 25 35
Chemical Oxygen Demand (mg/l) 30 75
Electrical Conductivity (mS/m) 250 250
Total Suspended Solids (mg/l) 10 90
Sodium Content (mg/l) 50 90
Soap/ Oil/ Grease None 2.5
Residual Chlorine (mg/l) 0 0.1
Free/ Saline Ammonia (mg/l) 1 1
Nitrate (mg/l) 1.5 None
Orthophosphate (mg/l) 1 1
Adapted from Government Gazette, 1984; (A): Guidelines for effluent being discharged into any catchment area/ river or a tributary (B): Guidelines for effluent being discharged into any area other than that specified by A.
1.9 Enteric viruses commonly detected in the water environment and associated