Phylogenetic and genetic variation among Fe(II)-oxidizing acidithiobacilli supports the view that these comprise multiple species with different ferrous iron oxidation pathways

Abstract

Autotrophic acidophilic iron-and sulfur-oxidizing bacteria of the genus Acidithiobacillus constitute a heterogeneous taxon encompassing a high degree of diversity at the phylogenetic and genetic levels, though currently only two species are recognized (Acidithiobacillus ferrooxidans and Acidithiobacillus ferrivorans). One of the major functional disparities concerns the biochemical mechanisms of iron and sulfur oxidation, with discrepancies reported in the literature concerning the genes and proteins involved in these processes. These include two types of high-potential iron-sulfur proteins (HiPIPs): (i) Iro, which has been described as the iron oxidase; and (ii) Hip, which has been proposed to be involved in the electron transfer between sulfur compounds and oxygen. In addition, two rusticyanins have been described: (i) rusticyanin A, encoded by the rusA gene and belonging to the well-characterized rus operon, which plays a central role in the iron respiratory chain; and (ii) rusticyanin B, a protein to which no function has yet been ascribed. Data from a multilocus sequence analysis of 21 strains of Fe(II)-oxidizing acidithiobacilli obtained from public and private collections using five phylogenetic markers showed that these strains could be divided into four monophyletic groups. These divisions correlated not only with levels of genomic DNA hybridization and phenotypic differences among the strains, but also with the types of rusticyanin and HiPIPs that they harbour. Taken together, the data indicate that Fe(II)-oxidizing acidithiobacilli comprise at least four distinct taxa, all of which are able to oxidize both ferrous iron and sulfur, and suggest that different iron oxidation pathways have evolved in these closely related bacteria. INTRODUCTIONThe majority of obligately chemolithoautotrophic acidophilic bacteria that can oxidize ferrous iron [Fe(II)], sulfur and reduced inorganic sulfur compounds (RISCs) have for many years been considered a priori to be strains of the well-documented species Acidithiobacillus ferrooxidans (At. ferrooxidans). The genus Acidithiobacillus has generally been considered to be a member of the Gammaproteobacteria, though recent phylogenetic analysis suggests it arose after divergence from the Alphaproteobacteria but before the Betaproteobacteria/Gammaproteobacteria split (Williams et al., 2010). There have been numerous reports suggesting that Fe(II)-oxidizing acidithiobacilli are a heterogeneous collection of bacteria with sufficient genetic variability to warrant classification as more than one species (Harrison, 1982(Harrison, , 1984Novo et al., 1996;Amils et al., 1998;Selenska-Pobell et al., 1998;Paulino et al., 2001;Karavaiko et al., 2003;Mitchell et al., 2003;Bergamo et al., 2004; Akbar et al., 2005;Waltenbury et al., 2005;Peng et al., 2006;Ni et al., 2007Ni et al., , 2008a. For instance, a Abbreviations: BV, bootstrap value; HiPIP, high potential iron-sulfur protein; MLSA, multilocus sequence analysis; PP, posterior probability; RISC, reduced inorganic sulfur compound.3T...

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