© 2015 Kato, Ohkuma, Powell, Krepski, Oshima, Hattori, Shapiro, Woyke and Chan. Neutrophilic microaerophilic iron-oxidizing bacteria (FeOB) are thought to play a significant role in cycling of carbon, iron and associated elements in both freshwater and marine iron-rich environments. However, the roles of the neutrophilic microaerophilic FeOB are still poorly understood due largely to the difficulty of cultivation and lack of functional gene markers. Here, we analyze the genomes of two freshwater neutrophilic microaerophilic stalk-forming FeOB, Ferriphaselus amnicola OYT1 and Ferriphaselus strain R-1. Phylogenetic analyses confirm that these are distinct species within Betaproteobacteria; we describe strain R-1 and propose the name F. globulitus. We compare the genomes to those of two freshwater Betaproteobacterial and three marine Zetaproteobacterial FeOB isolates in order to look for mechanisms common to all FeOB, or just stalk-forming FeOB. The OYT1 and R-1 genomes both contain homologs to cyc2, which encodes a protein that has been shown to oxidize Fe in the acidophilic FeOB, Acidithiobacillus ferrooxidans. This c-type cytochrome common to all seven microaerophilic FeOB isolates, strengthening the case for its common utility in the Fe oxidation pathway. In contrast, the OYT1 and R-1 genomes lack mto genes found in other freshwater FeOB. OYT1 and R-1 both have genes that suggest they can oxidize sulfur species. Both have the genes necessary to fix carbon by the Calvin-Benson-Basshom pathway, while only OYT1 has the genes necessary to fix nitrogen. The stalk-forming FeOB share xag genes that may help form the polysaccharide structure of stalks. Both OYT1 and R-1 make a novel biomineralization structure, short rod-shaped Fe oxyhydroxides much smaller than their stalks; these oxides are constantly shed, and may be a vector for C, P, and metal transport to downstream environments. Our results show that while different FeOB are adapted to particular niches, freshwater and marine FeOB likely share common mechanisms for Fe oxidation electron transport and biomineralization pathways.