Fig 1. CBS is the key enzyme for H₂S production in V. cholerae.

(A). Schematic illustration of the proposed pathways for H₂S biogenesis in V. cholerae. CBS, CSE and 3MST are key enzymes in cysteine degradation that are conserved from bacteria to mammals, while NADPH-dependent sulfite reductase (CysJI) catalyzes the last step in assimilatory sulfite reduction. (B). Cysteine degrading enzyme CBS is essential for H₂S production. To investigate the contribution of cysteine degradation (cbs, cse, and 3mst) and assimilatory sulfite reduction (cysI and cysJ) to H₂S synthesis in V. cholerae, H₂S production was detected with Pb(Ac)₂ paper strips with (upper) or without (bottom) supplementation of 200 μM L-cysteine hydrochloride in the culture medium. Stained paper strips were scanned and quantified with ImageJ. Average H₂S level of the wild-type (WT) was set to 100% for subsequent normalization. Three replicates were sampled for each strain. Asterisks indicate statistically significant differences compared to WT as found by t-test (***, p-value < 0.001). (C). Real-time detection of intracellular H₂S signals. H₂S signal was detected by WSP-5 (working concentration 15 μM, green signal) for bacteria that were cultivated in LB. Picture captured in the 488–524 channel was merged with the bright field image. Scale bar indicates length of 10 μm. Bottom chart exhibits end-point detection of H₂S production for the corresponding strains in the top chart.