Diversity and community structure of culturable arsenic-resistant bacteria across a soil arsenic gradient at an abandoned tungsten-tin mining area

We studied the bacterial diversity at a single location (the Terrubias mine; Salamanca province, Spain) with a gradient of soil As contamination to test if increasing levels of As would (1) change the preponderant groups of arsenic-resistant bacteria and (2) increase the tolerance thresholds to arsenite As(III)] and arsenate As(V)] of such bacteria. We studied the genetic and taxonomic diversity of culturable arsenic-resistant bacteria by PCR fingerprinting techniques and 16S rRNA gene sequencing. Then, the tolerance thresholds to As(III) and As(V) were determined for representative strains and mathematically analyzed to determine relationships between tolerances to As(III) and As(V), as well as these tolerances with the soil contamination level. The diversity of the bacterial community was, as expected, inversely related to the soil As content. The overall preponderant arsenic-resistant bacteria were Firmicutes (mainly Bacillus spp.) followed by γ-Proteobacteria (mainly Pseudomonas spp.), with increasing relative frequencies of the former as the soil arsenic concentration increased. Moreover, a strain of the species Rahnella aquatilis (γ-Proteobacteria class) exhibited strong endurance to arsenic, being described for the first time in literature such a phenotype within this bacterial species. Tolerances of the isolates to As(III) and As(V) were correlated but not with their origin (soil contamination level). Most of the strains (64%) showed relatively low tolerances to As(III) and As(V), but the second most numerous group of isolates (19%) showed increased tolerance to As(III) rather than to As(V), even though the As(V) anion is the prevalent arsenic species in soil solution at this location. To our knowledge, this is the first study to report a shift towards preponderance of Gram-positive bacteria (Firmicutes) related to high concentrations of soil arsenic. It was also shown that, under aerobic conditions, strains with relatively enhanced tolerance to As(III) predominated over the most As(V)-tolerant ones.