|Callahan, Mary Theresa|
|East, Cheryl - Roberts|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 3/22/2013
Publication Date: N/A
Technical Abstract: Introduction: Mature, finished compost made from various feedstocks should undergo testing for the presence of Escherichia coli O157:H7 to ensure thermal destruction of the pathogen during composting. Immunomagnetic separation (IMS) –based methods may provide an assay which can be conducted within 24-48 h to determine presence or absence of E. coli O157:H7 in finished compost. Purpose: 1) To compare the ability of two IMS methods – automated recirculating (AR) and direct plating (DP) - to recover E. coli O157:H7 from compost samples containing biosolids, manure, or yardwastes; 2) to determine the ability of compost samples to support regrowth of E. coli O157:H7 in compost. Materials and Methods: Twenty-nine USCC-STA certified compost samples were collected from across the U.S. Aliquots (400g) of each sample were inoculated with stx-2 positive E. coli O157:H7 at 10^1 to 10^2 cfu/g. Homogenized samples were incubated 5 h at 37oC for DP, then serially diluted and incubated 2 h at room temperature, then 42oC for 6 h. For both methods, samples were incubated overnight at 4oC before addition of immunomagnetic beads. Sample recirculation (30 min) and DNA purification from immunomagnetic beads, was followed by real-time PCR targeting the stx2 gene. For DP, immunomagnetic beads were added to diluted samples, mixed, removed, and then plated onto chromagarO157. Results: The recirculation and DP methods both recovered E. coli O157:H7 from 30/ 30 (100%) samples. PCR detection of E. coli O157:H7 was greatly enhanced by the removal of polyphenols and humic acids from DNA samples. Regrowth of E. coli O157:H7 in compost varied based on feedstock. Significance: IMS recovery, coupled with either direct plating or real-time PCR detection, is an efficient method to recover E. coli O157:H7 from finished compost within 24 – 48 h. Regrowth potential of E. coli O157:H7 in finished composts is affected by physicochemical parameters of compost.