qRT-PCR verification of S. flexneri chromosomal genes induced under anaerobic growth conditions and the role of FNR in the process. Strand specific qRT-PCR analysis of mRNA levels of S. flexneri M90T chromosomal genes shown to be induced under anaerobiosis in RNA-seq analysis. Panel A shows transport and metabolic genes, and panel B acid resistance, OMP and regulatory genes. Data were calculated as the n-fold difference relative to polA (2-ΔCt, where ΔCt represents the difference in threshold cycle between the target and control genes). Results are shown in relation to the wild-type strain 2-ΔCt levels under aerobic conditions, here referred to as 1. Thus, values greater than 1 indicate increased transcription under anaerobiosis, and lower than 1 indicate the opposite. Significant differences were detected when wild-type 2-ΔCt levels under aerobic and anaerobic conditions, or wild-type vs. Δfnr 2-ΔCt levels under anaerobiosis were compared. ns = non-significant, P < 0.05, *; P <0.01, **; n = 4; Mann–Whitney test. Error bars show Standard Deviation (SD).