Bacterial community structure analysis on Listeria monocytogenes inoculated spinach leaves is affected by PCR based methods to exclude chloroplast co-amplification
This study investigated the bacteriome of spinach leaves, stored under EURL challenge conditions for 9 days after inoculation with L. monocytogenes using two methods of excluding chloroplast co-amplification (COMPETE, BLOCK) at the PCR step as well as a post-PCR chloroplast sequence filter option (CONTROL). While all three approaches have demonstrated a change of bacterial communities over time, the pPNA based BLOCK approach resulted in greater diversity similarities to the CONTROL option. The COMPETE solution with a specifically designed primer to prevent chloroplast amplification had a strong underrepresentation of the Planctomycetota phylum and to a lesser extend underrepresentation of Chloroflexi and Verrucomicrobiota due to the inheritance of the selected primer region that allowed to deselect chloroplast co-amplification. However, the COMPETE approach had the lowest level of chloroplast co-amplification. Higher growth potential of L. monocytogenes from day 7 to 9 co-occurred with higher relative abundances of Pseudomonadaceae and lower numbers of Lactobacillales, suggesting that particular phylogenetic groups may support growth of L. monocytogenes. While chloroplast co-amplification with spinach in the present study was relatively modest and a purely filter based chloroplast removal was sufficient, other leafy vegetables may require one of the tested co-amplification prevention solutions.