Exposure to oxygen and light generates photooxidative stress by the bacteriochlorophyll a mediated formation of singlet oxygen ((1)O(2)) in Rhodobacter sphaeroides. Our study reports the genome-wide search for small RNAs (sRNAs) involved in the regulatory response to (1)O(2). By using 454 pyrosequencing and Northern blot analysis, we identified 20 sRNAs from R. sphaeroides aerobic cultures or following treatment with (1)O(2) or superoxide (O(-)(2)). One sRNA was specifically induced by (1)O(2) and its expression depends on the extracytoplasmic function sigma factor RpoE. Two sRNAs induced by (1)O(2) and O(-)(2) were cotranscribed with upstream genes preceded by promoters with target sequences for the alternative sigma factors RpoH(I) and RpoH(II). The most abundant sRNA was processed in the presence of (1)O(2) but not by O(-)(2). From this and a second sRNA a conserved 3'-segment accumulated from a larger precursor. Absence of the RNA chaperone Hfq changed the half-lives, abundance and processing of (1)O(2)-affected sRNAs. Orthologues of three sRNA genes are present in different alpha-proteobacteria, but the majority was unique to R. sphaeroides or Rhodobacterales species. Our discovery that abundant sRNAs are affected by (1)O(2) exposure extends the knowledge on the role of sRNAs and Hfq in the regulatory response to oxidative stress.