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Pathogen Characterization of Fresh and Stored Biosolids and Implications of a Screening Level Microbial Risk Assessment

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A limited study of enteric pathogens from different stages of mesophilic anaerobic digestion (MAD), i.e., primary sludge, and liquid and dewatered (cake) biosolids, was conducted in six wastewater treatment plants (WWTP) in Ontario, Canada. In addition, bacterial pathogens were re-analysed for regrowth after 2 to 3 days storage in a laboratory at 30°C. Pathogen datasets were used in a preliminary screening level quantitative microbial risk assessment (QMRA). The QMRA assumed conservative human health exposure scenarios for ingestion of soil containing freshly surface-applied or incorporated sewage biosolids, and also considered the aerosol pathway. Culture methods (Most Probable Number or plate count) were used to analyse for bacterial pathogens, including: Campylobacter spp.Salmonella spp.Listeria monocytogenes and Yersinia enterocolitica; as well as fecal indicators: fecal coliforms, E. coli, enterococci and Clostridium perfringens. Two additional pathogens Cryptosporidium parvum and Giardia lamblia, were quantified in a select few samples by polymerase chain reaction coupled with Most Probable Number (PCR-MPN). Salmonella and Listeria were found frequently in primary sludge and liquid digested biosolids (70–100% of samples) but less so in dewatered cake (50–60%). Yersinia was found less frequently (in 20–30% of samples) across all treatment stages. Giardia was found in 80% of cake biosolids while Cryptosporidium was found in 20%. E. coli reduction during MAD was more than 2-log (at 4 of the 6 plants), while reduction of bacterial pathogens was variable, as shown in other literature. Increases in bacteria levels immediately upon dewatering (i.e., “reactivation”), were generally not significant. Salmonella and E. coli densities increased 1 to 3-log after 2-3 days storage at 30°C, signifying potential for regrowth. Significant correlations between E. coli and Salmonella were observed after the regrowth period (r >0.55, p<0.02). Pathogen datasets from dewatered biosolids were used in a preliminary screening level QMRA, using Monte Carlo analyses. For QMRA simulations pathogen data were fit to log normal distributions using censored regression, which statistically accounts for the non-detects in pathogen data. QMRA results must be interpreted with caution given the limited datasets and conservative nature of the assumptions applied; QMRA results were intended solely for research, to identify data gaps and inform priorities. QMRA revealed that median risks of enteric infection and illness from indirect ingestion of aerosolized particulates were orders of magnitude lower than those from direct ingestion. Median risks of gastrointestinal illness from Salmonella were very low, even after regrowth (e.g., 10-16). Giardia and Cryptosporidium posed higher risks compared to Salmonella, but estimates for these protozoan pathogens were based on very few samples (n=9) which were highly censored. In addition these data were derived from molecular detection methods that did not distinguish viable and/or infectious organisms. Comparison of risks from different biosolids land application scenarios illustrated that incorporation in the top 15 cm of soil could potentially reduce risks by greater than two orders of magnitude. Results support the need for future research on pathogen regrowth in biosolids under field conditions, improved detection methods for Giardia and Cryptosporidium, and future QMRA studies, to better quantify potential human health risks associated with pathogens in land applied biosolids.

Keywords: Cryptosporidium; Giardia; Listeria; QMRA; Salmonella; biosolids; pathogens; quantitative microbial risk assessment; regrowth; wastewater treatment

Document Type: Research Article


Publication date: 2009-01-01

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