Quantitative detection of viable helminth ova from raw wastewater, human feces, and environmental soil samples using novel PMA-qPCR methods

Gyawali, Pradip; Ahmed, Warish; Sidhu, J. P. S.; Nery, Susana Vaz; Clements, Archie C. A.; Traub, Rebecca J.; McCarthy, James S.; Llewellyn, Stacey; Jagals, Paul; Toze, Simon

doi:10.1007/s11356-016-7039-9

Cited by 27 publications

(23 citation statements)

References 54 publications

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“…Molecular methods are under development for detection of STH in stool and biosolids [23,24]. Additionally, two recent studies used molecular methods for detection of hookworm species and Ascaris lumbricoides in soil [25,26]. Molecular methods should be also be created and tested for detection of Ascaris and Trichuris in soil.…”

Section: Introductionmentioning

confidence: 99%

Detecting and enumerating soil-transmitted helminth eggs in soil: New method development and results from field testing in Kenya and Bangladesh

et al. 2017

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Globally, about 1.5 billion people are infected with at least one species of soil-transmitted helminth (STH). Soil is a critical environmental reservoir of STH, yet there is no standard method for detecting STH eggs in soil. We developed a field method for enumerating STH eggs in soil and tested the method in Bangladesh and Kenya. The US Environmental Protection Agency (EPA) method for enumerating Ascaris eggs in biosolids was modified through a series of recovery efficiency experiments; we seeded soil samples with a known number of Ascaris suum eggs and assessed the effect of protocol modifications on egg recovery. We found the use of 1% 7X as a surfactant compared to 0.1% Tween 80 significantly improved recovery efficiency (two-sided t-test, t = 5.03, p = 0.007) while other protocol modifications—including different agitation and flotation methods—did not have a significant impact. Soil texture affected the egg recovery efficiency; sandy samples resulted in higher recovery compared to loamy samples processed using the same method (two-sided t-test, t = 2.56, p = 0.083). We documented a recovery efficiency of 73% for the final improved method using loamy soil in the lab. To field test the improved method, we processed soil samples from 100 households in Bangladesh and 100 households in Kenya from June to November 2015. The prevalence of any STH (Ascaris, Trichuris or hookworm) egg in soil was 78% in Bangladesh and 37% in Kenya. The median concentration of STH eggs in soil in positive samples was 0.59 eggs/g dry soil in Bangladesh and 0.15 eggs/g dry soil in Kenya. The prevalence of STH eggs in soil was significantly higher in Bangladesh than Kenya (chi-square, χ2 = 34.39, p < 0.001) as was the concentration (Mann-Whitney, z = 7.10, p < 0.001). This new method allows for detecting STH eggs in soil in low-resource settings and could be used for standardizing soil STH detection globally.

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Section: Introductionmentioning

confidence: 99%

Detecting and enumerating soil-transmitted helminth eggs in soil: New method development and results from field testing in Kenya and Bangladesh

et al. 2017

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“…As the PMA concentration increased from 150 to 200 lM (groups 9 and 10), the gene copy number fell by a statistically significant amount (P < 0.05), which suggested that a PMA concentration as high as 200 lM exerted toxicity towards the viable bacteria, producing a mortality rate of 63%. The toxicity could be explained by the infiltration of excessive PMA to the viable cells (Gyawali et al, 2016). Therefore, it is necessary to determine the maximum PMA concentration for specific samples, especially for manure, sludge and soil (Gyawali et al, 2016).…”

Section: Resultsmentioning

confidence: 99%

New insights into the kinetics of bacterial growth and decay in pig manure–wheat straw aerobic composting based on an optimized PMA–qPCR method

Huang

Sun

et al. 2019

Microbial Biotechnology

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Summary Aerobic composting is a bacteria‐driven process to degrade and recycle wastes. This study quantified the kinetics of bacterial growth and decay during pig manure–wheat straw composting, which may provide insights into microbial reaction mechanisms and composting operations. First, a propidium monoazide–quantitative polymerase chain reaction ( PMA – qPCR ) method was developed to quantify the viable bacteria concentration of composting samples. The optimal PMA concentration and light exposure time were 100 μM and 8 min respectively. Subsequently, the concentrations of total and decayed bacteria were quantified. Viable and decayed bacteria coexisted during the entire composting period (experiments A and B), and the proportion of viable bacteria finally fell to only 35.1%. At the beginning, bacteria grew logarithmically and decayed rapidly. Later, the bacterial growth in experiment A remained stable, while that of experiment B was stable at first and then decomposed. The duration of the stable stage was positively related to the soluble sugar content of composting materials. The logarithmic growth and rapid decay of bacteria followed Monod equations with a specific growth (0.0317 ± 0.0033 h −1 ) and decay rate (0.0019 ± 0.0000 h −1 ). The findings better identified the bacterial growth stages and might enable better prediction of composting temperatures and the degree of maturation.

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“…Nevertheless, non-human helminths parasites have been detected in fresh water [25], soil, and wastewater samples by qPCR [26]. …”

Section: Introductionmentioning

confidence: 99%

Quantitative PCR and Digital PCR for Detection ofAscaris lumbricoidesEggs in Reclaimed Water

Soto

Santísima-Trinidad

Bornay-Llinares

et al. 2017

BioMed Research International

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The reuse of reclaimed water from wastewater depuration is a widespread and necessary practice in many areas around the world and must be accompanied by adequate and continuous quality control. Ascaris lumbricoides is one of the soil-transmitted helminths (STH) with risk for humans due to its high infectivity and an important determinant of transmission is the inadequacy of water supplies and sanitation. The World Health Organization (WHO) recommends a limit equal to or lower than one parasitic helminth egg per liter, to reuse reclaimed water for unrestricted irrigation. We present two new protocols of DNA extraction from large volumes of reclaimed water. Quantitative PCR (qPCR) and digital PCR (dPCR) were able to detect low amounts of A. lumbricoides eggs. By using the first extraction protocol, which processes 500 mL of reclaimed water, qPCR can detect DNA concentrations as low as one A. lumbricoides egg equivalent, while dPCR can detect DNA concentrations as low as five A. lumbricoides egg equivalents. By using the second protocol, which processes 10 L of reclaimed water, qPCR was able to detect DNA concentrations equivalent to 20 A. lumbricoides eggs. This fact indicated the importance of developing new methodologies to detect helminth eggs with higher sensitivity and precision avoiding possible human infection risks.

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Quantitative detection of viable helminth ova from raw wastewater, human feces, and environmental soil samples using novel PMA-qPCR methods

Cited by 27 publications

References 54 publications

Detecting and enumerating soil-transmitted helminth eggs in soil: New method development and results from field testing in Kenya and Bangladesh

Detecting and enumerating soil-transmitted helminth eggs in soil: New method development and results from field testing in Kenya and Bangladesh

New insights into the kinetics of bacterial growth and decay in pig manure–wheat straw aerobic composting based on an optimized PMA–qPCR method

Quantitative PCR and Digital PCR for Detection ofAscaris lumbricoidesEggs in Reclaimed Water

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