Fig. 3

Assessing the recovery of FFPE bacterial DNA by quantitative PCR and 16S rRNA sequencing. a Evaluating PCR recovery of FFPE bacterial DNA from Protoblocks fixed for 24 h (green) or 48 h (cyan) and compared with the recovery of paired NF samples (red). (i) % of absolute PCR recovery (% shown above corresponding box). A 2-log fold decrease in recovery is observed for FFPE-treated samples, which was found to be statistically significant in all cases as per 1 sample Wilcoxon signed-rank test. In addition, longer fixation periods lead to a significantly greater reduction in recovery (p = 0.04). (ii) % deviation in recovery after compensating for 10-fold loss in recovery. Input = 0 (dotted line). % deviation shown above corresponding box. Significant deviation from input values, even after compensation for 10-fold decrease shown in all FFPE-treated samples. (In all cases, p = + < 0.1, * < 0.05, ** < 0.01 and *** < 0.001). b Sample composition bar plot of calculated input of bacterial cells added to Protoblock and 16S rRNA gene sequence analysis of Protoblocks fixed for 24 h or 48 h. c Average concentration of DNA purified from samples