A comparison of methods used to enumerate Escherichia coli in conventionally treated sewage sludge

Abstract

Aims: This study examined the suitability of three analytical methods for isolating and enumerating Escherichia coli from conventionally treated sewage sludge. Methods and Results: Crude sewage, mesophilic anaerobic digested (MAD) sludge, and final product sludge samples were taken from six sewage treatment works for analysis. Two of the three methods tested were membrane filtration techniques, utilizing chromogenic E. coli/coliform (CEC) media and membrane-lactose glucuronide agar (MLGA); the third method was a most probable number (MPN) technique utilizing Colilert in Quantitray 2000 (Idexx). The methods were evaluated for variation, consistency, false-positive and false-negative results, as well as method correlation. The methods gave good and consistent recovery of E. coli for a range of conventionally treated sewage matrices. All of the methods had a false-positive rate of <3%, although MLGA had a high false-negative rate (35AE5%) compared with Colilert (3AE81%) and the CEC method (6AE75%). This resulted in slightly lower presumptive counts but comparable numbers of confirmed counts. Conclusions: The three detection methods tested, chromogenic, MLGA and Colilert gave comparable recoveries, and did not vary by greater than one order of magnitude (1 log). Significance and Impact of the Study: Forthcoming revisions to the Use of Sludge in Agriculture Regulations (1989) will categorize sewage sludge as untreated, conventionally treated or enhanced treated in accordance to microbiological standards. The standard will be based upon numbers of E. coli removed through the sludge treatment process and the numbers remaining in the final product. It is recommended that the Colilert 2000 (Idexx, Westbrook, Maine) and CEC methods would be equally suitable to assess the reduction of indigenous E. coli in conventionally treated sludges, and that MLGA be used with follow-up confirmatory testing.

Evaluation of the effectiveness of a commercially available defined substrate medium and enumeration system for measuring Escherichia coli numbers in faeces and soil samples*

“…The comparable recovery of E. coli with Colilert is not surprising as previous studies have shown the effectiveness of Colilert for recovering E. coli from a range of different matrices (Berger 1991;Covert et al 1992;Frampton and Restaino 1993;Eccles et al 2004).…”

“…The Colilert method has been validated for use on various food and water samples as summarized in the review by Frampton and Restaino (1993). In addition the Colilert method has recently been shown to be an appropriate means of enumerating E. coli in sewage sludge (Eccles et al 2004). In this paper we used samples containing faecal material and soil to compare the recovery of E. coli by the Colilert method with a miniaturized MPN method that used traditional microbiological media.…”

Are the defined substrate-based methods adequate to determine the microbiological quality of natural recreational waters?

“…In addition, b-galactosidase has been found to be present in non-coliform strains, which makes the level of enzymatic activity a result of the proportion of target and non-target bacteria in rapid enzymatic assays without a cultivation step (Tryland & Fiksdal 1998 (Fricker et al 1997;Eckner 1998;Yakub et al 2002;Buckalew et al 2006). However, since the Colilert w system requires short incubation periods and is easy to apply, it can be useful for enumerating coliforms from natural waters and wastewaters (Grasso et al 2000;Kramer & Liu 2002;Eccles et al 2004). Studies carried out with tropical marine and freshwater samples resulted in a disparity between the numbers estimated by Colilert w and membrane filtration (Chao et al 2003).…”

Modified Moore swab optimization and validation in capturing E. coli O157:H7 and Salmonella enterica in large volume field samples of irrigation water

“…Before inoculation and after water filtration, turbidity levels were determined using the most probable number (MPN) of total coliforms and E. coli using IDEXX Colilert reagent in the QuantiTray 2000 format (IDEXX Lab. ; Westbrook, ME) in two sub-samples of 100 mL each (Eccles, Searle, Holt, & Dennis, 2004).…”