Purpose: A proficiency testing (PT) study was conducted to evaluate performance of laboratories in detecting Shigella in spiked sausage utilizing qPCR and/or culture methods.
Methods: Prior to shipping the samples were tested for in-house validation of sample preparation techniques, homogeneity and stability testing. At least 3 replicates of samples containing S. flexneri or S. dysenteriae at 66 - 100, 665- 1000 and 6650 – 10000 CFU/25g with or without confounding enterics were tested by qPCR and culture methods. A set composed of 4 Shigella spiked (10000 CFU/25g) samples, two non-Shigella spiked samples and two un-spiked samples were distributed to 49 laboratories for Shigella analyses using qPCR for screening and/or culture method for confirmation.
Results: During in-house validation, a total of 22 and 30 samples at each inoculation level were tested for Shigella flexneri and Shigella dysenteriae, respectively. S. flexneri (17% - 67%) and S. dysenteriae ( <34% ) were accurately identified with Biolog Shigella and R&F Shigella chromogenic, MacConkey, Xylose Lysine Deoxycholate and/or Hektoen media; however 100% of the samples were accurately detected with qPCR. In the proficiency study, 22% and 78% of participants analyzed samples by culture and qPCR procedures, respectively. Culture methods detected 36 – 82% and qPCR method detected 79 – 100% of the spiked samples. Of the positive qPCR results, only 29 - 87% of were culturally confirmed.
Significance: The real-time PCR method is a highly sensitive and selective assay which can be used as a screening tool. The low recovery rate of Shigella isolates indicated that the current culture method needs to improve in two areas: (1) Develop more selective enrichment to cut down background flora. (2) Adopt or develop differential and/or selective agars for better isolation.