An organotrophic Sideroxydans reveals potential iron oxidation marker genes

To understand the ecophysiology and the role of iron-oxidizing bacteria (FeOB) in various ecosystems, we need to identify marker genes of the iron oxidation pathway to track activity in situ . The Gallionellaceae Sideroxydans sp. CL21, an autotrophic iron-oxidizing bacteria isolated from a peatland, is unusual amongst FeOB isolates in its genomic potential to utilize organic compounds as energy sources. Thus, it offers the unique opportunity to elucidate which genes are expressed under litho- versus organotrophic conditions. We demonstrated the growth of Sideroxydans sp. CL21 on organic substrates (lactate and pyruvate) and inorganic substrates (Fe(II), magnetite, thiosulfate, and S(0)). Thus, cells were capable of lithoautotrophic, organotrophic, and potentially organoheterotrophic growth. Surprisingly, when lactate-grown cells were given Fe(II), mid-log phase cells were unable to oxidize iron, while late-log phase cells oxidized iron. To elucidate iron oxidation pathways, we compared gene expression between mid-log (non-iron-oxidizing) and late-log (iron-oxidizing) lactate-grown cells. Genes for iron oxidases ( cyc2, mtoA ) were highly expressed at both time points, so did not correspond to iron oxidation capability, making them unsuitable marker genes of iron oxidation activity by themselves. However, genes encoding periplasmic and inner membrane cytochromes were significantly upregulated in cells capable of iron oxidation. These genes include mtoD , cymA/imoA , and a cluster of Fe(II)-responsive genes ( ircABCD ). These findings suggest Gallionellaceae regulate their iron oxidation pathways in multiple stages, with iron oxidase-encoding genes proactively expressed. Other genes encoding electron carriers are upregulated only when iron oxidation is needed, which makes these genes (i.e. ircABCD ) good prospective indicators of iron oxidation ability.