Quantitative Microbial Risk Assessment for Acute Gastrointestinal Inflammation and viral Hepatitis A in Michigan, U.S.A., 2014 – 2018

4 Poster Track: Applied Research » 4 Health, safety and hygiene

Speaker: Florian Thymen Kleinhoven

Summary – for publication in conference brochure:

Vaccination, in addition to safe personal and environmental hygiene practices, contributes to the reduction of faecal-oral transmission of pathogens. Epidemiological data used often relies on the availability of sanitation facilities to map the primary pathways of person-to-person transmission. This study alternatively uses microbial indicator organisms, such as C. perfringens to assess the primary routes of transmission. This study determined significant correlation of logarithmically normalized C. perfringens abundance with enteroviruses (p < 0.05). This study determined a much more constant ratio of enteroviruses to C. perfringens ranging from 4% to 30% under aerobic sampling conditions.

Introduction, methods, results and discussion:

Enteroviruses are classified as a group of pathogens causing water-washed diseases, which are difficult to track because of the many different routes of transmission. Molecular detection methods of viral genetic material have often proven insufficient because of their relative expense and time requirements. Alternatively, indicator organisms, such as C. perfringens, may be able to provide an approximate substitute. Historical assessments of natural and municipal water and wastewater showed that C. perfringens remains relatively resistant to chlorination and environmental factors in which C. perfringens correlated with various types of enteric pathogens in 61% of samples. Additionally, evidence has suggested that C. perfringens presents an exceedingly complicated adaptive response when introduced into aquatic environments with high concentrations of reactive dissolved oxygen species. As a result, C. perfringens is found in much higher numbers than either fecal coliform or strepto- and enterococci in aerated wastewater streams. Another potential benefit is that other than fecal bacteria, C. perfringens has been shown not to replicate extensively in the natural environment. Additionally, the presence of C perfringens in sediment has been shown to be heavily dependent on environmental factors and therefore minimally impacts the measurability in water samples. This study aimed to verify to what extent C. perfringens enumeration may correlate with the degree of fecal contamination present in the water sample. The occurrence of pathogens in hydrologic systems was represented using the binary logistic regression in this study. Previous research has shown that the abiotic and environmental factors of a catchment may provide a viable indicator of the potential presence of enteroviruses. This study compared the constancy of the ratio between C. perfringens and enteroviruses between anoxic and aerobic sampling conditions throughout different stages of water sanitation facilities. The degree of correlation between the indicator organisms and enterovirus content was established by computing the likelihood ratio and Pearson correlation coefficient. Excreta-related microorganism content, including fecal coliforms, strepto- and enterococci, and C. perfringens were obtained from previous studies.The analysis of variance indicated that there was a highly significant correlated relationship between C. perfringens and enterovirus counts under anoxic sampling conditions (p < 0.001). However, although the correlation under aerobic sampling conditions was still highly significant, the 95% confidence interval was significantly smaller, while the degree of correlation between fecal coliform bacteria and enteroviruses outweighed that of C. perfringes (p < 0.05). Previous evidence suggests that under aerobic conditions, the degree of mineralization in the water sample might be greater, affecting the viability of less environmentally resilient organisms, including C. perfringens. The surprisingly high correlation coefficient between fecal coliforms and enteroviruses in these samples may be difficult to explain because of fecal coliform bacteria’s ability to replicate in aerobic environments. Additionally, the results suggest that the accumulation of C. perfringens spores in the sediment combined with the rapid die-off of strepto- and enterococci may represent an inadequately favorable picture of C. perfringens as an indicator. Nevertheless, C. perfringens poses a promising fecal indicator organism as a result of the spores’ hardiness in highly acidic or alkaline environments. 

Conclusions and implications:

The disproportionate fluctuation in the constancy ratio between C. perfringens or fecal strepto- and enterococci and enteroviruses shows that neither is an ideal indicator. Nevertheless, the results show that the degree of correlation between each of the indicator organisms assayed and the presence of fecal contamination is highly significant (p < 0.01). The dependency of the number of enteroviruses on the number of C. perfringens spores in the environment indicates a higher likelihood ratio than previously quantified. Specifically, amples taken under aerobic conditions showed C. perfringens' comparative hardiness under extreme environmental conditions. The general consensus of these results is that the certainty of the constancy ratio decreases with an increased number of indicator organisms in the sample, indicating a higher likelihood of predicting the pathogen content as higher dilution implies a lower individual indicator organism count. Future research should focus on quantifying the concentration gradients of indicator organisms from the enumeration of C. perfringens spores or other organisms with highly constant ratios between different locations.  These dose-independent data may be used to predict the responses in populations reliant to greater extents on groundwater sources for freshwater in these regions. 

Relevant references:

Duncan D. and G.P. Alabaster (1995)
“An environmental classification of housing-related diseases in developing countries”
Journal of Tropical Medicine and Hygiene
Duncan D. and Robert G.A. Feachem (1999)
“Water- and excreta-related diseases: Unitary Environmental Classification”
Journal of Environmental Engineering
J.I. and Thomas P. Curtis (1983)
“The Removal of Excreted Bacteria and Viruses in Deep Waste Stabilization Ponds”
Water Science and Technology
Joan B. and D.E. Huffman (2001)
“Reduction of Enteric Microorganisms at the Upper Occoquan Sewage Authority Water Reclamation Plant”
Water Environment Research
M.S. and E.D. Ebel (2014)
"Estimating the correlation between concentrations of two species of bacteria with censored microbial testing data". International Journal of Food Microbiology

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