Desk studies on feasibility of horizontal standard rapid methods for detection of E. coli (including E. coli O157) and Salmonella

The emerging methods becoming available for the rapid detection and enumeration of E. coli (including E. coli O157) and Salmonella in sludges, soil and treated biowastes have been evaluated with a view to possible future standardisation. The main methods that are available for the detection and enumeration of E. coli (including E. coli O157) and Salmonella have been developed largely for analysis of food and water and can be broadly divided into four groups. Proprietary Quantitray® technology, equivalent to the 5-tube most probable number (MPN) technique, employing disposable plastic trays for enumeration of E. coli and Salmonella. Immunological, involving a short or overnight pre-enrichment of the target organism followed by specific detection of cellular antigen in either a lateral flow device or following immunomagnetic capture. Molecular, involving PCR amplification of target DNA sequences from low numbers of cells, or preferably following a short pre-enrichment of the organism to amplify numbers and demonstrate viability prior to molecular detection. Physico-chemical, involving techniques such as measurement of impedance changes during enrichment and growth in appropriate media. The merits of each are described, in relation to their suitability for use with sludge, soil and biowastes. Since the majority of agar and MPN broth techniques take between 24-96 hours for identification and enumeration, we define “rapid” as any technique that detects, and if possible, enumerates the target organism in under 24 hours.All of the methods described have strengths and weaknesses, dependent on not only the Regulators’ types of requirements for sludge, soil and biowaste analysis but also their sensitivity, specificity, speed and cost. It is unlikely therefore that there can be only one methodology applicable to both E. coli (and E. coli O157) and Salmonella detection. Nevertheless, it is considered feasible to formulate horizontal standards to cover rapid analysis of E. coli and Salmonella in sludge, soil, soil improvers, growing media, and biowaste. None of the methods have been extensively evaluated for sewage sludge, soils or biowastes. As such, there is an urgent need for their modification and evaluation as part of the next phase of the Project Horizonta

Desk studies on feasibility of horizontal standard rapid methods for detection of Clostridium perfringens and enterococci in sludges, soil, soil improvers, growing media and biowastes

The existing methods currently available for the detection and enumeration of clostridia and enterococci in sludges and treated biowastes have been evaluated with a view to possible standardisation. The main methods used for the detection and enumeration of Clostridium and Enterococcus spp. have been developed largely for analysis of food and water and can be broadly divided into three groups. Quantification of colonies on agar media; most probable number (MPN) quantification in indicator broth using conventional test tube technology; and proprietary Quantitray® technology equivalent to the 5-tube MPN technique employing disposable plastic trays for enumeration of enterococci. The merits of each are described. At least one report has suggested that the use of m-CP agar medium, which is used in the reference method in the European Union, is not suitable for recovering C. perfringens spores from groundwater. This questions its possible use as a method for detecting C. perfringens in sludge, soil, soil improvers, growing media, and biowaste.Indeed, all of the methods described for detection of C. perfringens and enterococci have strengths and weaknesses, dependent on not only the Regulators’ types of requirements for sludge, soil and biowaste analysis but also their sensitivity, specificity, speed and cost. Nevertheless, it is considered feasible to formulate horizontal standards to cover analysis of C. perfringens and enterococci in sludge, soil, soil improvers, growing media, and biowaste. However, none of the methods have been extensively evaluated for these waste types. As such, there is an urgent need for their modification and evaluation as part of the next phase of the Project Horizonta