Elucidating fecal pollution patterns in alluvial water resources by linking standard fecal indicator bacteria to river connectivity and genetic microbial source tracking.

A novel concept for fecal pollution analysis was applied at alluvial water resources to substantially extend the information provided by fecal indicator bacteria (FIB). FIB data were linked to river connectivity and genetic microbial source tracking (MST). The concept was demonstrated at the Danube River and its associated backwater area downstream of the city of Vienna, using a comprehensive 3-year data set (10 selected sites, n = 317 samples). Enumeration of Escherichia coli (ISO 16649-2), intestinal enterococci (ISO 7899-2) and Clostridium perfringens (ISO 14189) revealed a patchy distribution for the investigation area. Based on these parameters alone a clear interpretation of the observed fecal contamination patterns was not possible. Comparison of FIB concentrations to river connectivity allowed defining sites with dominating versus rare fecal pollution influence from the River Danube. A strong connectivity gradient at the selected backwater sites became obvious by 2D hydrodynamic surface water modeling, ranging from 278 days (25%) down to 5 days (<1%) of hydraulic connectivity to the River Danube within the 3-year study period. Human sewage pollution could be identified as the dominating fecal source at the highly connected sites by adding information from MST analysis. In contrast, animal fecal pollution proofed to be dominating in areas with low river connectivity. The selection of genetic MST markers was focusing on potentially important pollution sources in the backwater area, using human (BacHum, HF183II), ruminant (BacR) and pig (Pig2Bac) -associated quantitative PCR assays. The presented approach is assumed to be useful to characterize alluvial water resources for water safety management throughout the globe, by allocating fecal pollution to autochthonous, allochthonous, human or animal contamination components. The established river connectivity metric is not limited to bacterial fecal pollution, but can be applied to any type of chemical and microbiological contamination.

Poikilothermic Animals as a Previously Unrecognized Source of Fecal Indicator Bacteria in a Backwater Ecosystem of a Large River.

Quantitative information regarding the presence of Escherichia coli, intestinal enterococci, and Clostridium perfringens in poikilotherms is notably scarce. Therefore, this study was designed to allow a systematic comparison of the occurrence of these standard fecal indicator bacteria (SFIB) in the excreta of wild homeothermic (ruminants, boars, carnivores, and birds) and poikilothermic (earthworms, gastropods, frogs, and fish) animals inhabiting an alluvial backwater area in eastern Austria. With the exception of earthworms, the average concentrations of E. coli and enterococci in the excreta of poikilotherms were equal to or only slightly lower than those observed in homeothermic excreta and were 1 to 4 orders of magnitude higher than the levels observed in the ambient soils and sediments. Enterococci reached extraordinarily high concentrations in gastropods. Additional estimates of the daily excreted SFIB (E. coli and enterococcus) loads (DESL) further supported the importance of poikilotherms as potential pollution sources. The newly established DESL metric also allowed comparison to the standing stock of SFIB in the sediment and soil of the investigated area. In agreement with its biological characteristics, the highest concentrations of C. perfringens were observed in carnivores. In conclusion, the long-standing hypothesis that only humans and homeothermic animals are primary sources of SFIB is challenged by the results of this study. It may be necessary to extend the fecal indicator concept by additionally considering poikilotherms as potential important primary habitats of SFIB. Further studies in other geographical areas are needed to evaluate the general significance of our results. We hypothesize that the importance of poikilotherms as sources of SFIB is strongly correlated with the ambient temperature and would therefore be of increased significance in subtropical and tropical habitats and water resources.IMPORTANCE The current fecal indicator concept is based on the assumption that the standard fecal indicator bacteria (SFIB) Escherichia coli, intestinal enterococci, and Clostridium perfringens multiply significantly only in the guts of humans and other homeothermic animals and can therefore indicate fecal pollution and the potential presence of pathogens from those groups. The findings of the present study showed that SFIB can also occur in high concentrations in poikilothermic animals (i.e., animals with body temperatures that vary with the ambient environmental temperature, such as fish, frogs, and snails) in an alluvial backwater area in a temperate region, indicating that a reconsideration of this long-standing indicator paradigm is needed. This study suggests that poikilotherms must be considered to be potential primary sources of SFIB in future studies.