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Evaluation of microbial viability techniques on monitoring Ascaris suum in wastewater

Raw wastewater is a vital reusable water source particularly for agricultural lands in developing countries. However, wastewater also acts as a reservoir for microbial pathogens that pose a significant public health risk. To evaluate these risks, microbial presence in wastewater needs to be properly assessed by accurate and sensitive methods. Currently, the presence of microbial pathogens in wastewater are evaluated by culture-based methods, LIVE/DEAD BacLight™ viability staining, and PMA-qPCR. The culture-based method involves incubating a wastewater sample for several days before analyzing the amount of embryonated ova under a microscope. The BacLight™ staining method, which also requires a microscope, distinguishes viable from non-viable cells based on membrane integrity. Meanwhile, PMA-qPCR is a unique PCR-based method that relies on inhibiting PCR amplification of DNA from non-viable cells after treatment with PMA, a photoreactive DNA binding dye. The Ascaris species of parasitic worms are extremely prevalent in contaminated wastewater and are used as indicators to evaluate the effectiveness of water treatment processes. At present, there are no comparative studies that directly assess the effectiveness of each method for monitoring viable Ascaris ova in raw wastewater.

In a recent issue of Water Science & Technology, Ravindran et al. studied the efficacy of PMA-qPCR on monitoring viable Ascaris suum ova in wastewater relative to LIVE/DEAD® BacLight™ staining and the culture-based method. Results from all three methods show no significant difference in percentage viability of A. suum in raw wastewater, suggesting comparable detection efficiencies. In addition, the authors note significant drawbacks of microscopy-based viability methods due to their inherent variability and time-consumption. In contrast, PMA-qPCR offers comparable assessment with greater speed and specificity than the other two-methods when evaluating A. suum ova viability in raw wastewater.

Viability of A. suum ova in raw wastewater evaluated by culture-based, BacLight Live/Dead staining and PMA-qPCR. Credit: Ravindran et al. doi:10.2166/wst.2019.286


Full Citation

Ravindran, V. B., Shahsavari, E., Soni, S. K. & Ball, A. S. Viability determination of Ascaris ova in raw wastewater: a comparative evaluation of culture-based, BacLight Live/Dead staining and PMA-qPCR methods. Water Sci. Technol. 1–10 (2019). doi:10.2166/wst.2019.286

Learn more about Biotium’s PMA and PMAxx™ Viability PCR Dyes as well as our bacterial and yeast viability stains for microscopy or flow cytometry.