qRT-PCR verification of S. flexneri chromosomal genes repressed under anaerobic growth conditions and the role of FNR in the repression. Strand specific qRT-PCR analysis of mRNA levels of S. flexneri M90T chromosomal genes shown to be repressed in RNA-seq analysis. 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 wild-type 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 indicates 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. P <0.01, **; n = 4; Mann–Whitney test. Error bars show Standard Deviation (SD).