Enumeration of Coliform Bacteria in Wastewater Solids Using Defined Substrate Technology

Commercially available defined substrate technology for the enumeration of coliform bacteria was investigated for its applicability to waste activated sludge (WAS) solids. The defined substrate, Colilert, in conjunction with the QuantiTray enumeration system, was compared with multiple‐tube fermentation (MTF). Multiple‐tube fermentation analyses (U.S. Environmental Protection Agency [U.S. EPA]‐accepted methods for the enumeration of coliforms in wastewater solids) were conducted on settled wastewater solids using lauryl tryptose broth as the presumptive phase and brilliant‐green lactose bile broth for the confirmed phase of total coliform enumeration. Both EC medium for fecal coliform identification and EC‐4‐methyl‐umbelliferyl‐β‐D‐glucuronide (MUG) media for Escherichia coli for the MTF completed phase were also conducted on the WAS. Samples analyzed included raw WAS and WAS that had been pasteurized at 70 °C for 15 and 30 minutes, respectively. Twenty replicates of each sample were measured to demonstrate statistical equivalence of the total coliform numbers for all three WAS samples (raw, 15‐minute pasteurization, and 30‐minute pasteurization). Results show that both systems provided statistically equal total coliform most probable numbers (MPNs) for all samples tested. As expected, fecal coliform numbers identified by the MTF technique were statistically higher than E. coli enumerated by Colilert–QuantiTray. However, the E. coli MPN generated by the MTF method was found to be statistically equal to the Colilert–QuantiTray numbers. Because of its ease of use and reduced time to obtain the analysis, the Colilert–QuantiTray system can be useful in enumerating coliforms in WAS. However, more data are required to definitively show equivalence of the commercially available defined substrate technology to the U.S.‐EPA accepted protocol. It is hoped that the results of this study will help form a database for the acceptance of the Colilert–QuantiTray system for WAS bacterial analyses.

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