Validation of Test Portion Pooling for the Detection of Listeria spp. and L. monocytogenes in Dairy Products

Introduction: Listeria monocytogenes is a major foodborne pathogen. Testing of multiple portions of a same food item is often required to verify the effectiveness of a HACCP plan. It will be advantageous to the laboratories to combine these test portions and process as one sample as it will result in efficiency. 

Purpose: Validate pooling of test portions for the detection of L. monocytogenes and Listeria spp. in dairy products as no scientific evidence currently exists to support this practice. 

Methods: Six matrices, namely, pudding, yogurt, brie cheese, 2% milk, ice cream and infant formula were spiked separately with stressed L. monocytogenes and Listeria spp. in 25 g and pooled test portions (375 g/ 250 g/125 g). ISO 11290-1:1996 Amd1:2004 and Rapid L’mono methods were followed until confirmation step for the detection of the spiked organisms. Performance of a method in pooled test portions was considered to be satisfactory if the relative limit of detection (RLOD₅₀; LOD₅₀ [pooled test portion] / LOD₅₀ [25-g test portion]) obtained was ≤ 2.5 (acceptance criteria following draft ISO 16140-2).

Results: Acceptable RLOD₅₀ (≤ 2.5) values were consistently obtained when test portions were pooled up to 125 g for all matrices tested. Obtained RLOD₅₀ values ranged from 0.719 to 1.666 and 0.568 to 2.435 for the detection of L. monocytogenes and Listeria spp., respectively, using both methods. LOD₅₀ values obtained from the testing of 125-g test portions was approximately 1 CFU for most of the matrices. Though satisfactory RLOD₅₀ values were obtained for milk (0.568 – 1.132) and infant formula (0.874 – 1.666) matrices when pooled up to 375 g, it was not the case for other food items.

Significance: Pooling of test portions to up to 125 g for the detection of L. monocytogenes and Listeria spp. by two culture methods in dairy products has been validated.