LIST OF ABBREVIATIONS
CHAPTER 5: DISCUSSION
5.4 Characterisation and identification of presumptive faecal bacteria and HPC isolates
up and wash faecal material and other chemical constituents into the Loopspruit River. The results depicted in Figure 4.8 show that the WWTP contributed both E. coli and enterococci during the wet season and urban land use during the dry seasons. Agricultural activities contributed a moderate amount of E. coli and enterococci during the wet seasons and only enterococci during the dry seasons.
Hall et al. (2014) used multivariate statistical methods to determine the fate of faecal pollution in surface water of the Watauga River watershed in northeast Tennessee, USA. The authors found that during wet warm seasons, faecal coliform concentrations increased significantly in comparison with the dry seasons. These findings correlate with a study by Kostyla et al. (2015) that focused on the seasonal variation of faecal contaminations in developing countries. The differences between the wet warm and dry cold seasons are the result of climatic changes.
Increased rainfall with more runoff events and higher temperature both contribute to higher faecal coliform counts during the wet season (Hall et al., 2014; Kim et al., 2017; Sibanda et al., 2013).
5.4 Characterisation and identification of presumptive faecal
The enterococci species E. faecalis and E. faecium pose risks to humans causing nosocomial infections and may also cause endocarditis, urinary tract infections and bacteraemia (Fernández- Guerrero et al., 2002; Ratanasuwan et al., 1999). These organisms pose risk to humans who are exposed to E. faecalis and E. faecium containing waters with serious infections. Possible sites that may be at health risk include the urban areas such as MU01, VA07 and GP08, and KW03 at a WWTP and informal settlement.
5.4.2 Identification of Clostridia isolates
Due to the anaerobic nature of Clostridia, they are commonly found in the intestine of humans and animals but are not limited to these environments. Twenty-seven of the 31 Clostridia isolates were identified to species level and included Clostridium perfringens, C. baratii C. nitritogenes, C.
bifermentans and C. sordellii. These same species were isolated from the Schoonspruit River, Crocodile River and Groot Marico River by Fourie (2017). The presence of Clostridia is indicative of possible faecal contamination that may occur based on the surrounding land uses of the sample sites. These land uses include urban settings (MU01 and VA07), Agriculture (TS04 and KA05) and a WWTP (KW03).
Clostridium perfringens has been isolated from WWTPs, pigs and chicken farms as well as from plants (Abia, et al., 2015a; Álvarez-Pérez et al., 2018; Rimoldi et al., 2015; Voidarou et al., 2011;
Watcharasukarn et al., 2009). Clostridia were present at sample sites MU01, KW03, TS04, KD06, VA07 and GP08. Sample sites KD06, VA07 and GP08 are downstream of chicken farms. In a previous study, C. baratii and C. sordellii were isolated from chicken faeces (Rimoldi et al., 2015) upstream of sites KW03, TS04, KA05, KD06 and VA07.
A common Clostridium pathogen, Clostridium perfringens is usually found in the intestinal tract of humans and animals. C. perfringens’ pathogenicity ranges from gastroenteritis as a result of food poisoning to necrotising gas-gangrene (Irikura et al., 2015).
C. perfringens was present at sites MU01, KW03, TS04, KD06, VA07 and GP08. Sites TS04, KD06, VA07 and GP08 are all downstream of a WWTP. Wastewater effluent can possibly introduce this pathogen into the aquatic environment. Clostridium perfringens produces endospores that are highly resistant to the treatment processes and environmental stressors (Ajonina et al., 2015). This presents a cause for concern that this organism may cause health risks to both humans and animals.
5.4.3 Identification of presumptive E. coli isolates
The identified isolates included Shewanella xiamenensis (from MU01), Aeromonas hydrophila (from MU01, TS04 and GP08), E. coli (form MD02, KD03, KA05 and VA07), Serratia marcescens (from MU01), Klebsiella aerogenes (from KW03), Enterobacter cloacae (from TS04, KD06 and GP08), and Citrobacter freundii (from TS04). The expected outcome was to identify E. coli;
however, various other isolates were identified. The reason to only isolate E. coli is that it serves as a faecal indicator. The Escherichia, Citrobacter, Enterobacter, Serratia and Klebsiella genera are all part of the Enterobacteriaceae taxonomic Family. Shewanella and Aeromonas both follow the same taxonomic classification of their genera but differ in the Order classification with Alteromonadales and Aeromondales, respectively. E. coli produce a yellow slant with a yellow butt, is positive for gas production and does not produce H2S. Enterobacter, Klebsiella, Shewanella and Aeromonas all share these biochemical traits with E. coli. Citrobacter and Serratia tested negatively one parameter of the TSI test (black colour chance meaning hydrogen sulphide production). Citrobacter is positive for H2S and Serratia do not produce gas. These discrepancies may be due to observational errors. 16S rRNA sequencing is therefore needed for definitive identification of the organism.
An important consideration is the possible presence of Verocytotoxin-producing E. coli known as Enterohemorrhagic E. coli (EHEC) such as E. coli O157. This strain has been isolated in the North West Provence by Ateba et al., (2008), from cattle, pigs and human faeces. The E. coli O157 strain was found to be prevalent in cattle. Therefore, water sources that are frequented by grazing cattle could possibly contribute to the spread of E. coli O157. Hunter (2003), stated that E. coli O157:H7 may cause haemorrhagic enteritis or haemolytic uremic syndrome in humans.
5.4.4 Identification of HPC isolates
HPC isolates in the study were identified as Flavobacterium sp., Sphingobacterium sp., Chryseobacterium indoltheticum, Arcicella rigui, Arthrobacter psychrochitiniphilus, Beta proteobacteria, Janthinobacterium lividum, Rheinheimera sp., Aeromonas sp. and Pseudomonas sp. Which were similar HPC bacterial species found by Bezuidenhout et al. (2017); Carstens, (2012); Jordaan et al. (2019) in surrounding study areas. Opportunistic pathogens identified in the Loopspruit River from HPC bacteria were also observed in previous studies in the Mooi River comprising of Pseudomonas, Acinetobacter, Aeromonas and Flavobacterium (Horn et al., 2016).
Flavobacterium are known for their industrially important compounds (Enisoglu-Atalay et al., 2018) and enzymes. These include enzymes that can degrade agar, alginate, chitin, pectin, xylan,
environment, the hydrolysis of cellulose, xylan, and chitin is for the most part upheld by bacteria and fungi (Berlemont, 2017). Sphingobacterium sp. was isolated from soil in a rice paddy in Changzhou (Jiangsu Province, China) (Cai et al., 2015). This organism may have originated from agricultural settings; however, the microorganisms were isolated from sample site MD02, which is an urban area and situated upstream of the agricultural farms.
Fortunately, there were no human gastrointestinal diseases associated with Pseudomonas (WHO, 2011). Pseudomonas aeruginosa is a waterborne opportunistic pathogen that poses a risk to immuno-compromised populations (Wang et al., 2012). Increased levels of Pseudomonas may cause taste, odour and turbidity problems in water (WHO, 2011).
Aeromonas sp. and Pseudomonas sp. was present at all the sample sites of the Loopspruit River.
In a study by Kivanc et al. (2011) the authors found Aeromonas sp. in drinking water, tap water and environmental water samples. They also found that Aeromonas sp. were more prevalent during the cold and dry months. Pseudomonas sp. are commonly found in groundwater and plants. Pseudomonas sp. are considered to be a plant and animal pathogen (Cui et al., 2019).