Abstract

Acidic environments are characterised by low concentrations of bioavailable N-containing compounds. Therefore, the ability of indigenous microorganisms to fix nitrogen, represents an important trait and a key role for the community. Acidithiobacillus ferrooxidans, Acidithiobacillus ferrivorans, Leptospirillum ferrooxidans and L. ferrodiazotrophum are considered to be the only lithoautotrophic acidophiles that are able to fix dinitrogen (Norris et al, 1995; Tyson et al, 2005, Hallberg et al, 2010). Analysis of the genome of L. ferriphilum strain Sp-Cl revealed the presence genes involved in the nitrogen fixation, though previously, L. ferriphilum has not been considered to be a diazotroph. To asses the ability of members of the Leptospirillum genus to fix nitrogen, some strains of all three known species were screened for the presence of the nifH gene by PCR. Thirteen strains of Leptospirillum spp. were grown in autotrophic basal salts medium (pH 1.5), 50 mM of ferrous iron as energy source, at 37°C. Cell lysates were prepared to be used as template in 16S rRNA and nifH amplification as previously described (Hallberg et al., 2006; Ueda et al., 1995). All strains of L. ferrooxidans and L. ferrodiazatrophum yielded positive PCR products for nifH gene, with amplicons of the expected molecular weight (290 bp) being obtained. In contrast, 3 out of 5 strains of L. ferriphilum (including type strain) gave appropiate-size PCR products. The sequences of the amplified nifH fragments showed high similarity amongst all strains examinated. This study suggests that some strains of L. ferriphilum, like other Leptospirillum spp., have the potential to fix dinitrogen. While this important trait appears to be characteristic of all strains of L. ferrooxidans, it appears to be more sporadically distributed among strains of L. ferriphilum. Given the importance of L. ferriphilum in commercial mineral bioprocessing this ability may have important consequences for biomining operations