Detection of viable hookworm ova from wastewater and sludge

Abstract

Infectious helminths are a worldwide major public health problem. In terms of morbidity, approximately 3 × 10 9 people are infected by soil-transmitted helminths (STHs) influencing rates of malnutrition and failure to thrive. All of this increases the global disease burden (GDB) by up to 5.9 × 10 6 Daily Adjusted Life Years (DALYs). Helminth infections are more often seen as a major public health issue for developing countries however, concerns have been expressed in many of the developed nations because of the increased use of treated wastewater and sludge with no clear way of knowing the infection potential that can be attributed from such water.Guidelines have been established to minimise the potential public health risk associated with wastewater and sludge reuse. For example, the WHO guideline specified ≤ 1 ova (Ascaris lumbricoides) per L liquid (wastewater) or 4 g dry solid (sludge) for unrestricted use. Various wastewater treatment processes have been recommended to remove the helminths ova from wastewater depending upon its reuse. However, detection methods used to quantify viable helminths ova from wastewater has limitations. Therefore there is always a potential public health risk associated with reuse of wastewater and sludge.In this research, a real-time PCR method was developed. The new PCR method is rapid and specific. The method was found to be able to detect less than one ovum in one litre treated wastewater and approximately four ova in one litre raw wastewater and ~ 4 g sludge.The real-time PCR method was modified to a quantitative PCR (qPCR) method and further used to quantify hookworm ova from wastewater and sludge. The qPCR had estimated an average of 1.1, 8.6 and 67.3 ova for treated wastewater that was seeded with 1, 10 and 100 ova, respectively. The gene copy numbers obtained for 1, 10 and 100 ova by qPCR varied significantly (P < 0.05) within the tested samples indicating that absolute quantification of ova may not be accurate. Despite the difficulty quantifying accurate numbers of hookworm ova, the lower limit of quantification (LLOQ) of the qPCR method was 30 gene copies. This was a lot less than the gene copies produced by one ovum. Therefore, the qPCR method has potential to use for complying with wastewater guidelines.. iiAlthough the overall aim of this research was to develop a sensitive and specific method for quantitative detection of viable hookworm ova from wastewater, the importance of recovering the ova from wastewater and sludge samples for accurate detection was identified. While determining appropriate recovery rates was not an aim of this research, a suitable method that could be used to standardise research outcomes for further study was established. Therefore, ova recovery rate by different rapid methods was evaluated for further experiments. The result indicated that the ova recovery rate was higher for the treated wastewater (0.2 -50%) than the raw wastewater (0.3 -35%) and sludge (0.02 -4.7%) samples. A significant difference (P < 0.05) was observed between...