Investigating the presence of microbial pathogens in the Umhlangane River, Durban, South Africa.
The use of rivers for recreational and domestic practices makes it imperative to scrutinize the water quality circulating within the surrounding communities. The presence of potential pathogens in the Umhlangane River was monitored at five points (Phoenix industrial: P1; upstream KwaMashu wastewater/residential: P2; natural wetlands: P3; Riverhorse Valley industrial/business estate: P4; and Springfield industrial: P5) on a monthly basis from October 2013 to September 2014. Commonly measured physico-chemical parameters were determined according to standard protocols. Bacterial indicators were enumerated using the membrane filtration technique. A tangential flow filtration process was set up to remove the bacteria and to concentrate the virus populations from 25 ℓ of river water samples. Somatic and F⁺RNA coliphages were enumerated using plaque assays. The virus-like particles (VLPs) were estimated using epifluorescence microscopy and viral morphology was observed using transmission electron microscopy (TEM). The potential infectious nature of the concentrated viruses was assessed using cytopathic effect (CPE) of tissue culture. The specific detection of some virus populations was determined by a two round nested-PCR reaction using virus-specific primer sets and confirmed by sequencing. Chemical and biological oxygen demand (COD; BOD) fluctuated at all sampling points and months with BOD ranging from 0.48 mg/ℓ (Riverhorse Valley; April 2014) to 12.4 mg/ℓ (Phoenix industrial; June 2014), respectively. The highest COD content was recorded at the Phoenix industrial site in May with 269 mg/ℓ. The total dissolved solid (TDS) content and electrical conductivity (E.C.) fluctuated throughout all sampling months and points with all measurements exceeding the Department of Water Affairs recommended limits of 0 – 100 mg/ℓ and 0 – 15 mS/m, respectively. High counts of E. coli (EC), total and faecal coliform (TC; FC) and Shigella (SHIG) were recorded at the industrial sites in Phoenix and Springfiled and upstream of the KwaMashu WWTP in Phoenix while the total heterotrophic bacteria (THB) depicted the highest and lowest counts at the Phoenix industrial natural wetland sites ranging from 14.9 x 10⁶ cfu/100mℓ to 1.3 x 10⁶ cfu/100mℓ, respectively. Somatic and F⁺RNA coliphages produced its highest counts at the industrial site in Phoenix ranging from 765 pfu/mℓ and 585 pfu/mℓ in January 2014, respectively. Direct VLP counts were substantially lower (105 vlp/mℓ; April 2014) than the plaques produced by the somatic and F⁺RNA coliphages. Morphological changes of HEK293, Vero and Hep-G2 cell lines were indicative of a positive CPE for viral concentrates. Apart from visualization of bacteriophages belonging to the Siphoviridae, Myoviridae and Podoviridae families, presumptive Picornaviridae, Adenoviridae, Herpesviridae, Coronaviridae, Reoviridae, Polyomaviridae and Orthomyxoviridae VLPs were revealed based on size and comparisons to electron micrographs of known viruses. Adenovirus, polyomavirus, and hepatitis A and C virus-specific nested primers revealed the detection of these waterborne pathogens in the Umhlangane River. Moreover, sequence data confirmed the presence of these virus populations by comparisons made in GenBank. An increase in the amount of chemical pollutants entering the water would allow for the high COD, BOD and changing E.C. and TDS levels. Elevated THB populations at all sampling points and months indicate poor water quality. High EC, TC, FC and SHIG are indicative of possible faecal pollution, which could be attributed to faecal contamination entering the catchment. The presence of these indicators as well as the somatic and F⁺RNA coliphages could be due to anthropogenic activities, changing climatic conditions and the excreta of infected and non-infected individuals entering the river. Viruses or phages in the river water samples are morphologically diverse. Phage diversity further indicates diversity in their bacterial counterparts. The presence of various VLPs revealed by TEM together with substantial CPE on human tissue cell lines and the confirmation of adenoviruses, polyomaviruses as well as hepatitis A and C viruses by molecular detection and sequencing data raise the health concerns of the river system. The present study highlights the importance of routine environmental surveillance of human enteric viruses for a better understanding of the actual burden of these viral infections on those who might be using the water directly without treatment.
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