Comparison of concentration methods for rapid detection of hookworm ova in wastewater matrices using quantitative PCR

P Gyawali; W Ahmed; P Jagals; J P S Sidhu; S Toze

doi:10.1016/j.exppara.2015.09.002

Comparison of concentration methods for rapid detection of hookworm ova in wastewater matrices using quantitative PCR

Exp Parasitol. 2015 Dec:159:160-7. doi: 10.1016/j.exppara.2015.09.002. Epub 2015 Sep 8.

Authors

P Gyawali¹, W Ahmed², P Jagals³, J P S Sidhu⁴, S Toze⁴

Affiliations

¹ CSIRO Land and Water, Ecosciences Precinct, 41 Boggo Road, QLD 4102, Australia; School of Public Health, University of Queensland, Herston Road, QLD 4006, Australia. Electronic address: pradip.gyawali@csiro.au.
² CSIRO Land and Water, Ecosciences Precinct, 41 Boggo Road, QLD 4102, Australia; Faculty of Science, Health and Education, University of the Sunshine Coast, Maroochydore, DC, QLD 4558, Australia.
³ School of Public Health, University of Queensland, Herston Road, QLD 4006, Australia.
⁴ CSIRO Land and Water, Ecosciences Precinct, 41 Boggo Road, QLD 4102, Australia; School of Public Health, University of Queensland, Herston Road, QLD 4006, Australia.

PMID: 26358269
DOI: 10.1016/j.exppara.2015.09.002

Abstract

Hookworm infection contributes around 700 million infections worldwide especially in developing nations due to increased use of wastewater for crop production. The effective recovery of hookworm ova from wastewater matrices is difficult due to their low concentrations and heterogeneous distribution. In this study, we compared the recovery rates of (i) four rapid hookworm ova concentration methods from municipal wastewater, and (ii) two concentration methods from sludge samples. Ancylostoma caninum ova were used as surrogate for human hookworm (Ancylostoma duodenale and Necator americanus). Known concentration of A. caninum hookworm ova were seeded into wastewater (treated and raw) and sludge samples collected from two wastewater treatment plants (WWTPs) in Brisbane and Perth, Australia. The A. caninum ova were concentrated from treated and raw wastewater samples using centrifugation (Method A), hollow fiber ultrafiltration (HFUF) (Method B), filtration (Method C) and flotation (Method D) methods. For sludge samples, flotation (Method E) and direct DNA extraction (Method F) methods were used. Among the four methods tested, filtration (Method C) method was able to recover higher concentrations of A. caninum ova consistently from treated wastewater (39-50%) and raw wastewater (7.1-12%) samples collected from both WWTPs. The remaining methods (Methods A, B and D) yielded variable recovery rate ranging from 0.2 to 40% for treated and raw wastewater samples. The recovery rates for sludge samples were poor (0.02-4.7), although, Method F (direct DNA extraction) provided 1-2 orders of magnitude higher recovery rate than Method E (flotation). Based on our results it can be concluded that the recovery rates of hookworm ova from wastewater matrices, especially sludge samples, can be poor and highly variable. Therefore, choice of concentration method is vital for the sensitive detection of hookworm ova in wastewater matrices.

Keywords: Concentration methods; Hookworm ova; Public health; Recovery rate; Wastewater and sludge.

Publication types

Comparative Study
Research Support, Non-U.S. Gov't

MeSH terms

Ancylostoma / genetics
Ancylostoma / isolation & purification*
Animals
Centrifugation / standards
DNA, Helminth / isolation & purification
DNA, Ribosomal Spacer / analysis
Dogs
Feces / parasitology
Filtration / standards
Humans
Ovum
Queensland
RNA, Ribosomal, 5.8S / genetics
Real-Time Polymerase Chain Reaction / standards
Reference Standards
Reproducibility of Results
Sensitivity and Specificity
Sewage / parasitology
Ultrafiltration / methods
Ultrafiltration / standards
Wastewater / parasitology*
Water Purification / methods
Water Purification / standards*
Western Australia

Substances

DNA, Helminth
DNA, Ribosomal Spacer
RNA, Ribosomal, 5.8S
Sewage
Waste Water