Comparing Apples to Apples: Validating an Elution-based Defined Substrate Method to Enumerate Escherichia coli on Tree Fruit

Introduction: Agricultural water, in particular irrigation and cooling water, has been identified as a potential route for introduction of microbial contamination to apples.  However, a full understanding of the degree to which microbial contamination is introduced to fruit surfaces is limited by a lack of simple, efficient, and accurate methods to quantify microbial contamination on tree fruit. This study investigates a quick and easy-to-perform assay that incorporates surface elution and defined substrate technology to quantify E. coli on apples. 

Purpose: This study aims to evaluate the efficacy of a surface elution method for the enumeration of E. coli on apples, utilizing IDEXX Colilert^®-18 and QuantiTtray^®/2000.

Methods: Unwaxed apple samples were inoculated with E. coli at low, medium, and high seeding levels, each differing by an order of magnitude. Inoculated apples were massaged in a Whirl-Pak^® with 100ml of PBS for 5 minutes. The eluate was mixed with Colilert^®-18 substrate and sealed in a QuantiTray^®/2000. After incubation at 37°C for 18 hours, an MPN for E. coli was generated for each sample based on the number of positive wells (those that fluoresced under UV light).

Results: Using the surface elution method, the average recovery rate was 11.5% at the lowest inoculation level (~10² E. coli per sample; n = 16). At the medium inoculation level (~10³ E. coli per sample), the average recovery was 11.2% (n = 16). Samples in the highest seeding level (~10⁴ E. coli per sample) had an average recovery of 4.1% (n = 10). Apple samples ranged in weight from 112.1g to 198.5g.

Significance: The E. coli enumeration method evaluated in this study will aid researchers in conducting quantitative microbial risk assessment studies on apple handling and processing. Future studies utilizing this method may inform food safety policies to minimize the risk of apple microbial contamination.