BACTERIAL ROLE IN THE FORMATION OF POOL FINGERS IN CARBONATE CAVE POOLS

The degree to which bacteria play a role in the development of cave formations (a.k.a. speleothems), such as pool fingers that hang down from shelfstone in carbonate cave pools, is currently the subject of debate. To shed further light on this debate, we undertook a study of the biodiversity of pools containing cave carbonate pool precipitates in five New Mexico caves to look for the presence of known calcium carbonate precipitating bacteria, using molecular and cultivation techniques. In order to determine the diversity among the bacteria found in association with these calcite speleothems, pool water was filtered aseptically through a 0.2 micron filter from on or near the carbonate deposits. DNA was extracted from these filtered samples, and the 16S rRNA gene was amplified and cloned to create clone libraries for sequencing. Additional pyrosequencing also was performed. Analysis of clone and pyrosequencing library sequences has suggested that novel bacteria (<90% similar to any known bacteria based on genetic sequences) occur in areas of low human visitation (<100 visitors per year). The results also suggest that bacterial communities associated with these calcite formations are unique between caves. To test whether bacteria were present that possess the capability to precipitate calcite, carbonate surfaces below the water line of pools in Lechuguilla Cave and in Snowy River in Fort Stanton Cave were swabbed and subcultured to produce putatively pure cultures and then transferred to B4 medium to assess calcite production. Gram-staining of cultures with active precipitation showed bacilli and clustered coccoid-shaped bacteria, suggesting there are a variety of bacteria that may be precipitating calcite.

Using phylogenetic and community analyses, we showed that differences exist among communities of bacteria in the five New Mexico caves, and that many novel bacteria are present. Our results also suggest that multiple bacteria are present in caves that can precipitate calcite and that further research into the role that bacteria play in calcite speleothem formation is warranted.