GLOBAL BIOGEOGRAPHY AND DIVERSITY OF EXTREMELY ACIDIC CAVE-DWELLING ACIDOPHILIC COMMUNITIES

Hydrogen sulfide-rich caves host extremely acidic (pH 0-1) wall biofilms where gypsum replacement crusts isolate sulfide-oxidizing microbial communities from buffering by the parent limestone. Given the extreme acidity and subsurface location of these biofilms, and in light of earlier work showing strong geographic patterns among the dominant Acidithiobacillus populations, we hypothesized that the structure and diversity of the biofilm communities themselves would be impacted by geographic isolation. Using a combination of fluorescence in situ hybridization (FISH) and 16S rRNA gene cloning, we analyzed 26 extremely acidic “snottite” biofilm samples from 4 sulfidic cave systems in Italy and Mexico. All biofilms had low biodiversity and were dominated by Acidithiobacillus spp. Other abundant taxa included Ferroplasma and other Thermoplasmatales archaea (0-15% of cells), Acidimicrobium spp. (0-15% of cells) and rare phylotypes including Sulfobacillus spp. and members of the TM6 and TM7 bacterial lineages. Biofilms from the Mexican caves had substantially higher diversity compared with those from the Italian caves, and subsequent 16S rRNA gene sequencing using high-throughput techniques confirmed the same abundant taxa listed above and revealed additional diversity including substantial populations of Metallibacterium, Erythrobacter, and Saccharibacter spp. Across large geographic distances, snottites from caves nearest to each other had more similar community structure. Even though H₂S gas concentrations in cave air at the biofilm sample locations ranged over two orders of magnitude, sample locations with similar gas compositions did not have similar community compositions. Our results suggest that stochastic colonization events during community assembly in distant regions and/or subsequent isolation due to barriers to dispersal and colonization are important controls on the cave biofilm communities. At smaller spatial scales within cave systems, gas concentrations and other environmental variables such as predation, energy fluxes, or nutrient availability likely play a role in determining biofilm community compositions.