An assessment of reference method selective broths and plating media using 19 Listeria spp. highlights the importance of including diverse species in Listeria spp. method evaluations

Abstract

Reference methods developed for L. monocytogenes are commonly used for Listeria spp. detection. Improved method performance data are needed, since the genus Listeria has expanded from 6 to 26 species and now includes several Listeria sensu lato species, which can show phenotypes distinct from Listeria sensu stricto . Here, we evaluated growth of 19 Listeria spp., including 12 recently described sensu lato species, using the media specified by (i) the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual , (ii) the U.S. Department of Agriculture (USDA) Microbiology Laboratory Guidebook , and (iii) the International Organization for Standardization (ISO). The FDA enrichment procedure allowed all species to grow to detectable levels (≥ 4 log 10 ), yielded the highest mean growth (7.58 log 10 ), and was the only procedure where no sensu lato species yielded significantly higher bacterial growth than a sensu stricto species. With the USDA or ISO enrichment procedures several sensu lato species yielded significantly higher bacterial growth than either L. seeligeri or L. ivanovii , suggesting that these two sensu stricto species could be outgrown by sensu lato species. On selective and differential agars, L. seeligeri, L. ivanovii, and L. grayi yielded atypical colony morphologies and/or showed inhibited growth (which may lead to incorrect classification of a sample as negative), while several newly described sensu lato species grew well and showed typical morphologies. Overall, our study shows that the ability to detect different Listeria spp. can be impacted by the specific broth and selective and differential agars used. Our data will aid with selection of media and detection methods for environmental Listeria monitoring programs and facilitate selection of methods that are most likely to detect the targeted Listeria groups (e.g., Listeria sensu stricto, which appear to be the most appropriate index organisms for the pathogen L. monocytogenes ).