In addition to the traditional BOD/SS measures of effluent strength, the microbial
makeup of the spiking effluents is also important. There are virtually thousands of
pathogenic and non pathogenic organisms that could be present in septage and therefore
enumerated as part of this project. To keep the project costs within reason, we chose to
use the traditional total coliform, fecal coliform and E. coli indicators of fecal pollution as
well as two innovative indicators, enterococci and male specific coliphages.
Total coliform and E. coli counts were performed using the standard minimal
media ONPG-MUG enzyme based test system (Colilertam) with the Quantitraym
enumeration system. Total coliform is the indicator that has been traditionally used by
water microbiologists since 1920 to detect microbial contamination of wells. While total
coliform does, in fact, detect outside contamination of a water well system, its ability to
accurately predict the presence of fecal material is limited. None the less, it is important
to include this indicator since Wisconsin groundwater standards are still based on total
coliform occurrence. E. coli is included since it does accurately predict the presence of
fecal contamination in a water sample, and thus, is considered an excellent predictor of
imminent health risk from consuming water from a positive well.
Fecal coliform is a traditionally used indicator of fecal contamination in water.
While it has generally been replaced by E. coli, we are including it in this study to tie the
data to previously published studies. The detection method used was the standard
membrane filter fecal coliform test (MFFCC).
While E. coli has generally been proven to be an excellent indicator organism and
public health tool, its utility can be challenged in situations where contaminated water is
filtered through soil. The surface characteristics and shape of E. coli allow it to be easily
removed as it moves through a soil column, unlike the small round virus often associated
with waterborne disease outbreaks, that might move through more readily. For this
proposal we chose to include two additional indicators, enterococci and coliphage, that
behave more like the viral pathogens in their ability to move through soil.
Enterococci are small round bacteria found in the intestinal track of warm blooded
animals. For fifteen years, they have been suggested by the USEPA as the best indicator
organism to be used in testing of recreational waters, because of the strong correlations
with their presence in water with the occurrence of illness in swimmers. Enterococci
counts were performed using a proprietary defined substrate enzyme based test system
called Enterolertm combined with the Quantitrayam enumeration system.
Male specific coliphages have been suggested by several researchers as the fecal
indicator of choice because of their increased likelihood of behaving like a pathogenic
virus in ground water contamination situations. Coliphage are viruses that infect E. coli
bacteria. Male specific coliphages are viruses of E. coli that infect the bacterial cells
through sex pili appendages that are only found on the bacteria when they are at or near
body temperatures. In other words, the detection of male specific coliphages is strong
evidence of fecal contamination from a warm blooded animal. Additionally, since the*
coliphages are in fact viruses, they behave in the environment much as human pathogenic
virus do. For this study, samples were extracted with freon or chloroform. Recovered
phage were then detected using the double agar layer plating method using a host
organism vulnerable to infection by male specific coliphages.
Since the main focus of the research was to characterize the passage of microbes
through mound sand, the microbiological indicator flora of the effluents was determined