Performance of viral and bacterial genetic markers for sewage pollution tracking in tropical Thailand

Human Fecal Pollution Monitoring and Microbial Risk Assessment for Water Reuse Potential in a Coastal Industrial–Residential Mixed-Use Watershed

“…In this study, HAdV40/41 distribution functions were estimated from crAssphage distributions, as supported by a significant correlation between these two DNA markers in wastewater (Farkas et al, 2019;Sirikanchana et al, 2020;Tandukar et al, 2020;Wu et al, 2020). Moreover, HPyVs and crAssphage have been reported for their significant correlation in wastewater and environmental waters (Sangkaew et al, 2021), thus supporting the fact that similar estimated risks of HAdV40/41 were observed in this study when predicted from either HPyVs or crAssphage. Human-specific MST markers, such as crAssphage (Crank et al, 2019) and HF183 (Boehm et al, 2015;Ahmed et al, 2018), have been used for QMRA; they provide the advantage of higher abundance than waterborne pathogens (Zhang et al, 2019).…”

“…GenBac3 was detected in all samples with concentrations from 4.68 to 8.24 log 10 copies/100 ml (Table 1 and Figure 2). HPyVs were detected in all but two samples (MLOQs of 2.10 and 2.40 log 10 copies/100 ml), with a maximum of 5.10 log 10 copies/100 ml, while crAssphage was in all but three samples (MLOQs of 2.40 and 3.00 log 10 copies/100 ml) at concentrations up to 5.21 log 10 copies/100 ml, as previously reported (Sangkaew et al, 2021). HAdV40/41 was found only in three residential sites from Event 1 at MP1 (2.82 log 10 copies/100 ml), MP3 (2.59 log 10 copies/100 ml), and MP10 (3.87 log 10 copies/100 ml), while the rest were non-detects with an MLOQ of 2.40 log 10 copies/100 ml.…”

“…Human fecal contamination in this study was identified by the HPyVs and crAssphage markers of up to five orders in log 10 copies/100 ml in this study, which were relatively lower than those presented in untreated sewage in Thailand (three to six and five to seven orders, respectively) (Kongprajug et al, 2019b; Sangkaew et al, 2021) and in the Tha Chin river (three to seven orders for crAssphage) (Kongprajug et al, 2019b), but slightly higher than in beach water in Thailand (up to four and three orders, respectively) (Sangkaew et al, 2021). The levels of HPyVs and crAssphage in Thailand's freshwater were equivalent to those found in other geographical areas, as previously discussed (Sangkaew et al, 2021). Notably, marker abundance is also affected by the decay rate, which can be dependent on the water type (freshwater or marine water) and other environmental stressors (e.g., sunlight and indigenous microbiota) at different levels (Booncharoen et al, 2018;Korajkic et al, 2019).…”

“…A general fecal marker, GenBac3 (Siefring et al, 2008), two human-specific fecal markers, human polyomavirus BK and JC (HPyVs) (McQuaig et al, 2009) and crAssphage (Stachler et al, 2017), and adenovirus types 40/41 (HAdV40/41) (Ko et al, 2005) were used in this study (Supplementary Table 2). The abovementioned MST markers were selected because they have been investigated and validated in Thailand (Kongprajug et al, 2019a(Kongprajug et al, , 2020Sangkaew et al, 2021), and HAdV40/41 was chosen because it has been identified as the second most prevalent following the subgroup C from child patients with acute gastroenteritis in Thailand (Kumthip et al, 2019). A 20-µl qPCR reaction contained 0.8 µl of each 10 µM forward and reverse primers, 0.4 µl of 10 µM probe, 2 µl of extracted DNA, 6 µl of 1 µg/µl BSA, and 10 µl of the 2X iTaq Universal Probes Supermix (Bio-Rad, United States).…”

“…The GenBac3 assay was administered, and C q values for each dilution were plotted against the DNA concentration, and an R 2 of <0.90 suggested significant inhibition. Three field blanks and three method blanks were also investigated with HPyV and crAssphage assays (Sangkaew et al, 2021).…”