Browsing by Author "Reyneke, Brandon"

Now showing 1 - 5 of 5
  • Thumbnail Image
    Item
    Application of solar pasteurization for the treatment of harvested rainwater
    (Stellenbosch : Stellenbosch University, 2017-03) Reyneke, Brandon; Khan, Wesaal; Khan, Sehaam; Stellenbosch University. Faculty of Science. Dept. of Microbiology.
    ENGLISH ABSTRACT: Rainwater harvesting has been earmarked by South African governmental authorities as an intervention strategy that could alleviate the pressures on existing centralised water distribution systems, especially in rural areas and urban informal settlements, where insufficient waste removal and potable water infrastructure are available. However, numerous studies have indicated that harvested rainwater may not be safe to use for all daily water requirements, as numerous chemical and microbial contaminants may be associated with stored tank water. Rainwater treatment technologies, including solar pasteurization (SOPAS), have subsequently been investigated (Chapter 1). In order to determine whether decentralised rainwater harvesting SOPAS systems may be a viable alternative in providing the inhabitants of informal settlements with a supplementary water source, two small- (Sites 1 and 2) and one large-scale (Site 3) rainwater harvesting SOPAS systems were installed in Enkanini informal settlement, Stellenbosch, South Africa (Chapter 2). The microbial and chemical quality of the unpasteurized and pasteurized (produced by the respective systems) rainwater was monitored using conventional water quality monitoring techniques, including the culturing of indicator organisms, screening for selected indigenous rainwater pathogens using the polymerase chain reaction (PCR) and quantitative PCR (qPCR) assays and the monitoring of anion and cation concentrations. Additionally, the operational sustainability of the systems and water usage by the participating households were monitored. Chemical analyses indicated that all anions and cations were within the limits stipulated by various national and international drinking water quality guidelines, with the exception of zinc which contravened the respective guidelines before (mean: 3919 μg/L) and after (mean: 3964 μg/L) pasteurization at both Sites 1 and 2. In addition, the arsenic concentrations measured at Site 3 before (mean: 18.69 μg/L) and after (mean: 18.30 μg/L) pasteurization exceeded the respective drinking water guidelines. The increased zinc concentrations were attributed to the galvanised zinc roofing material installed at Sites 1 and 2, while the increased arsenic concentrations may be attributed to a roofing treatment or paint utilised to cover the catchment area at Site 3. Microbial analyses indicated that pasteurization temperatures of 53 °C (small-scale systems) and 55 °C (large-scale system) were required to reduce Escherichia coli and total and faecal coliforms to below the detection limit [< 1 colony forming units (CFU)/100 mL]. However, minimum pasteurization temperatures of 66 °C (small-scale systems) and 71 °C (large-scale system), were required to reduce the heterotrophic plate count (HPC) to within drinking water limits (1.0 × 104 CFU/100 mL). Of the opportunistic pathogens detected using PCR assays, Legionella spp. was the most prevalent pathogen detected in the small-scale systems [unpasteurized (100%) and pasteurized (91%)] and the large-scale system [unpasteurized (83%) and stored pasteurized tank water (100%)]. Quantitative PCR analysis then indicated that while the gene copies of Legionella spp., Pseudomonas spp. and Salmonella spp. were reduced during SOPAS, the organisms were still detected at the highest pasteurization temperatures analysed for each site (Site 1 – 85 °C; Site 2 – 66 °C; Site 3 – 79 °C). Additionally, the application of a metabolic responsiveness adenosine triphosphate (ATP) assay (BacTiter-GloTM Microbial Cell Viability Assay) indicated the presence of metabolically active cells in all pasteurized rainwater samples analysed. Results also indicated that the systems required limited maintenance and the small-scale systems in particular were able to provide the participating households with an alternative warm water source that could be utilised for numerous domestic purposes. As various limitations have been associated with the use of culture-based analyses for the monitoring of water quality, the aim of Chapter 3 was to compare molecular-based viability assays [ethidium monoazide bromide (EMA)-qPCR, propidium monoazide (PMA)-qPCR and DNase treatment in combination with qPCR] as well as the metabolic responsiveness ATP assay to culturing analysis for their ability to accurately determine cell viability in bacterial monocultures following heat treatment. Three Gram-negative (Legionella spp., Pseudomonas spp. and Salmonella spp.) and two Gram-positive (Staphylococcus spp. and Enterococcus spp.) bacteria commonly associated with water sources were selected as test organisms. Of the various concentrations of EMA and PMA analysed, 6 μM EMA and 50 μM PMA were identified as the optimal dye concentrations as low log reductions were recorded (viable and heat treated samples) in comparison to the no viability treatment control. Comparison of the results obtained for all the molecular viability assays (6 μM EMA, 50 μM PMA and DNase treatment) then indicated that the 6 μM EMA concentration was comparable to both the 50 μM PMA and the DNase treatment for the analysis of most of the test organisms (viable and heat treated). In addition, the results for the culturing analysis (CFU) of the viable S. typhimurium as well as the viable and heat treated samples of L. pneumophila and P. aeruginosa were comparable to the gene copies detected using molecular-based viability assays. However, the CFU in the heat treated samples of S. typhimurium were significantly lower than the gene copies detected using DNase in combination with qPCR, with no gene copies or CFU detected in the heat treated samples of S. aureus and E. faecalis. In contrast, while the ATP assays indicated the presence of metabolically active cells in the viable and heat treated samples, the ATP assay also indicated the presence of metabolically active cells in samples that had been autoclaved (negative viability control). It was thus concluded that molecular-based assays may be used to supplement culture based analysis for the comprehensive identification of the viable microbial population in water samples (before and after treatment).
  • Thumbnail Image
    Item
    Comparative analysis of solar pasteurization versus solar disinfection for the treatment of harvested rainwater
    (BioMed Central, 2016-12-09) Strauss, Andre; Dobrowsky, Penelope Heather; Ndlovu, Thando; Reyneke, Brandon; Khan, Wesaal
    Background: Numerous pathogens and opportunistic pathogens have been detected in harvested rainwater. Developing countries, in particular, require time- and cost-effective treatment strategies to improve the quality of this water source. The primary aim of the current study was thus to compare solar pasteurization (SOPAS; 70 to 79 °C; 80 to 89 °C; and ≥90 °C) to solar disinfection (SODIS; 6 and 8 hrs) for their efficiency in reducing the level of microbial contamination in harvested rainwater. The chemical quality (anions and cations) of the SOPAS and SODIS treated and untreated rainwater samples were also monitored. Results: While the anion concentrations in all the samples were within drinking water guidelines, the concentrations of lead (Pb) and nickel (Ni) exceeded the guidelines in all the SOPAS samples. Additionally, the iron (Fe) concentrations in both the SODIS 6 and 8 hr samples were above the drinking water guidelines. A >99% reduction in Escherichia coli and heterotrophic bacteria counts was then obtained in the SOPAS and SODIS samples. Ethidium monoazide bromide quantitative polymerase chain reaction (EMA-qPCR) analysis revealed a 94.70% reduction in viable Legionella copy numbers in the SOPAS samples, while SODIS after 6 and 8 hrs yielded a 50.60% and 75.22% decrease, respectively. Similarly, a 99.61% reduction in viable Pseudomonas copy numbers was observed after SOPAS treatment, while SODIS after 6 and 8 hrs yielded a 47.27% and 58.31% decrease, respectively. Conclusion: While both the SOPAS and SODIS systems reduced the indicator counts to below the detection limit, EMA-qPCR analysis indicated that SOPAS treatment yielded a 2- and 3-log reduction in viable Legionella and Pseudomonas copy numbers, respectively. Additionally, SODIS after 8 hrs yielded a 2-log and 1-log reduction in Legionella and Pseudomonas copy numbers, respectively and could be considered as an alternative, cost-effective treatment method for harvested rainwater.
  • Thumbnail Image
    Item
    Exploring the antimicrobial resistance profiles of WHO critical priority list bacterial strains
    (BMC (part of Springer Nature), 2019-12-23) Havenga, Benjamin; Ndlovu, Thando; Clements, Tanya; Reyneke, Brandon; Waso, Monique; Khan, Wesaal
    Background: The antimicrobial resistance of clinical, environmental and control strains of the WHO “Priority 1: Critical group” organisms, Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa to various classes of antibiotics, colistin and surfactin (biosurfactant) was determined. Methods: Acinetobacter baumannii was isolated from environmental samples and antibiotic resistance profiling was performed to classify the test organisms [A. baumannii (n = 6), P. aeruginosa (n = 5), E. coli (n = 7) and K. pneumoniae (n = 7)] as multidrug resistant (MDR) or extreme drug resistant (XDR). All the bacterial isolates (n = 25) were screened for colistin resistance and the mobilised colistin resistance (mcr) genes. Biosurfactants produced by Bacillus amyloliquefaciens ST34 were solvent extracted and characterised using ultra-performance liquid chromatography (UPLC) coupled to electrospray ionisation mass spectrometry (ESI–MS). The susceptibility of strains, exhibiting antibiotic and colistin resistance, to the crude surfactin extract (cell-free supernatant) was then determined. Results: Antibiotic resistance profiling classified four A. baumannii (67%), one K. pneumoniae (15%) and one P. aeruginosa (20%) isolate as XDR, with one E. coli (15%) and three K. pneumoniae (43%) strains classified as MDR. Many of the isolates [A. baumannii (25%), E. coli (80%), K. pneumoniae (100%) and P. aeruginosa (100%)] exhibited colistin resistance [minimum inhibitory concentrations (MICs) ≥ 4mg/L]; however, only one E. coli strain isolated from a clinical environment harboured the mcr-1 gene. UPLC-MS analysis then indicated that the B. amyloliquefaciens ST34 produced C13–16 surfactin analogues, which were identified as Srf1 to Srf5. The crude surfactin extract (10.00 mg/mL) retained antimicrobial activity (100%) against the MDR, XDR and colistin resistant A. baumannii, P. aeruginosa, E. coli and K. pneumoniae strains. Conclusion: Clinical, environmental and control strains of A. baumannii, P. aeruginosa, E. coli and K. pneumoniae exhibiting MDR and XDR profiles and colistin resistance, were susceptible to surfactin analogues, confirming that this lipopeptide shows promise for application in clinical settings.
  • Thumbnail Image
    Item
    A global review of the microbiological quality and potential health risks associated with roof-harvested rainwater tanks
    (Nature Research, 2019) Hamilton, Kerry; Reyneke, Brandon; Waso, Monique; Clements, Tanya; Ndlovu, Thando; Khan, Wesaal; DiGiovanni, Kimberly; Rakestraw, Emma; Montalto, Franco; Haas, Charles N.; Ahmed, Warish
    A broad body of literature has been published regarding roof-harvested rainwater quality around the world. In particular, the presence of fecal indicator bacteria and pathogenic microorganisms has raised concerns regarding the acceptability of rainwater for potable and non-potable uses. As the use of molecular assays has improved understanding of the diverse microbial communities present in rainwater tanks and their role in providing benefits or harm to human health, a comprehensive review is needed to summarize the state of the science in this area. To provide a summary of microbial contaminants in rainwater tanks and contextual factors, a comprehensive review was conducted here to elucidate the uses of rainwater, factors affecting water quality, concentrations of fecal indicators and pathogens, the attribution of pathogens to host sources using microbial source tracking, microbial ecology, human health risks determined using epidemiological approaches and quantitative microbial risk assessment, and treatment approaches for mitigating risks. Research gaps were identified for pathogen concentration data, microbial source tracking approaches for identifying the sources of microbial contamination, limitations to current approaches for assessing viability, treatment, and maintenance practices. Frameworks should be developed to assess and prioritize these factors in order to optimize public health promotion for roof-harvested rainwater.
  • Thumbnail Image
    Item
    The potential of solar disinfection in combination with bacteriophage biocontrol to reduce the health risks associated with contaminated harvested rainwater
    (Stellenbosch : Stellenbosch University, 2020-04) Reyneke, Brandon; Khan, Wesaal; Khan, Sehaam; Stellenbosch University. Faculty of Science. Dept. of Microbiology.
    ENGLISH ABSTRACT: Domestic rainwater harvesting may aid developing countries in achieving Sustainable Development Goal 6, which aims to ensure universal access to safe and affordable water. However, numerous microbial pathogens are associated with rainwater and treatment is required to reduce the potential health risk associated with using this water source. Solar disinfection (SODIS) has subsequently been identified as an easy-to-use and cost-effective treatment, where natural sunlight inactivates microorganisms through ultraviolet (UV) radiation and solar mild-heat. The small volumes of water that can effectively be treated (2 L) and decreased efficiency under poor weather conditions, have however been identified as key limitations of SODIS. The aim of Chapter 2 (published in Science of the Total Environment) was thus to monitor the efficiency of two large-volume batch solar reactor prototypes designed by the research consortium of the European Union (EU) Horizon 2020 project, Water Sustainable Point-of-Use Treatment Technologies (WATERSPOUTT). The systems were installed in a local informal settlement (Site 1; 88 L Prototype I) and rural farming community (Site 2; 140 L Prototype II) and were connected to rainwater harvesting systems. Due to space availability, a first-flush diverter system was installed at site 2. Conventional water quality monitoring techniques [chemical (anions, cations, physico-chemical parameters) and microbial (indicator organism and opportunistic pathogen culture-based analysis)] then indicated that the chemical quality of the untreated and treated rainwater adhered to national and international drinking water quality guidelines. While the microbial quality of the untreated rainwater exceeded the guideline limits at both sites, an 8 h solar exposure (6 and 8 h assessed) effectively reduced the indicator organism and monitored opportunistic pathogen counts to below the detection limit, with the exception of 43% of the samples collected from the Prototype I reactor (site 1), where heterotrophic bacteria still exceeded the guideline limit. An increased treatment efficiency was subsequently obtained at site 2 and was attributed to the first-flush diverter system (installed at this site) reducing the level of microbial contaminants entering the rainwater harvesting tank. As microorganisms may enter a viable but non culturable state under unfavourable conditions (such as those experienced during SODIS treatment), nucleic acid binding dyes were combined with quantitative polymerase chain reaction (qPCR) assays to monitor the removal of various opportunistic pathogens, including Escherichia coli (E. coli), Legionella spp. and Pseudomonas spp., amongst others. A mean overall reduction of 75% in the target organisms was then obtained for both solar reactors. It is however important to note that based on drinking water quality guidelines, the solar reactor prototypes were able to provide small households with the minimum essential daily water requirement for human activities of 25 L per person per day. Additionally, a generic Water Safety Plan was compiled for use in combination with the solar reactor prototypes to aid in ensuring the quality of the treated water. The detection of intact and potentially viable cells following solar reactor treatment however, highlighted the need for quantitative microbial risk assessment (QMRA) of the rainwater. However, as numerous microbial pathogens may be present within harvested rainwater, Illumina next-generation sequencing was coupled with ethidium monoazide (EMA) viability pre-treatment to elucidate the total viable microbial community in the untreated and treated rainwater (Chapter 3). Legionella, Pseudomonas, Mycobacterium, Clostridium sensu stricto and Escherichia were subsequently identified as the most frequently detected pathogenic or opportunistic pathogenic genera within the untreated and treated rainwater samples collected from both sites. Legionella pneumophila (L. pneumophila), Pseudomonas aeruginosa (P. aeruginosa) and E. coli were then selected as the target organisms for the health-risk assessment of the untreated and treated rainwater. Treatment with the solar reactor prototypes effectively reduced the health risk posed by E. coli and P. aeruginosa, when using rainwater for bathing/washing, garden work, cleaning and washing laundry (by hand), to below the one extra infection per 10 000 persons per year (1 × 10-4) benchmark limit. However, for both organisms the risk associated with using treated rainwater for drinking, still exceeded the benchmark limit. Additionally, while the solar reactor treatment reduced the risk associated with garden hosing and showering based on the presence of L. pneumophila, the risk estimate for using the treated rainwater for showering still exceeded the annual benchmark limit. The water produced by the large-volume batch solar reactor prototypes may thus safely be used for various domestic activities commonly practised within the target communities (e.g. washing/bathing, cleaning the home, etc.) in water scarce regions of sub-Saharan Africa. As Pseudomonas was identified as one of the dominant bacterial genera persisting following treatment using the solar reactor prototypes, bacteriophages targeting Pseudomonas spp. were isolated and characterised for the biocontrol pre-treatment of rainwater (Chapter 4; published in Environmental Science: Water Research and Technology). Bacteriophage PAW33 was isolated and characterised as a Podoviridae bacteriophage, with broad-spectrum activity against P. aeruginosa strains, and was applied in small-scale rainwater pre-treatment trials for 8 h and 24 h, to restrict the proliferation of an environmental P. aeruginosa S1 68 strain. The bacteriophage pre-treated samples were then subjected to treatment in small-scale SODIS compound parabolic collector (SODIS-CPC) systems for 4 h under natural sunlight. Although culture- and molecular-based analyses for the 8 h trial indicated similar total log reductions of P. aeruginosa S1 68 for the bacteriophage pre-treated and non-pre-treated samples, results for the 24 h trial (followed by SODIS) indicated that a higher log reduction was recorded for the pre-treated sample (4.61 log) in comparison to the non-pre-treated sample (3.91 log), using culture-based analysis, with comparable results obtained using EMA-qPCR. Gene expression analysis indicated that PAW33 pre-treatment for 24 h influenced the ability of P. aeruginosa S1 68 to initiate stress response mechanisms (decreased expression of the recA and lexA genes) during the SODIS-CPC treatment and resulted in the decreased expression of the phzM gene (virulence factor responsible for pyocyanin production). Bacteriophage PAW33 thus displays promise as a pre-treatment strategy of rainwater as it restricts the proliferation of P. aeruginosa and may increase the efficiency of primary disinfection methods.