Patterns of microbes within caves and across regions: Similarities where microbes are most stable deep in the aphotic zone.

Background/Question/Methods  Caves are often a stable environment.  Microorganisms in deep portions of caves that are no longer undergoing active formation are surviving in an aphotic environment under constant temperatures and humidity.  Although stable, resources, especially carbon, are limited.  We sampled multiple cave sites from Belize, Illinois, Iowa and Virginia at three depths, including directly at the mouth, midway in low light, and deep in the aphotic region.  A limestone mine in Southern Illinois was used as a control.  We employed a polyphasic approach to compare microbiological results from sediment samples.   The analyses performed were aerobic heterotrophic plate counts, direct microscopic enumeration, colony morphology-based diversity indices and BiOLOG physiological fingerprinting.  BiOLOG analysis included both the use of GN and GP identification micro-plates and the three panel Eco-plates used to describe the bacterial community carbon usage at the various depths from each cave.   

Results/Conclusions BiOLOG GN/GP plate analysis indicated a common presence of Actinomycete genus types in nearly all samples.  Various species of Fracisella, Rhizobium and Vibrio genus were also found in mid to deep sites in several temperate zone caves.   Bacterial total direct counts were universally at or above 1 x 10⁸ cells/g sediment except for the mine, where cell numbers peaked at low 10⁶ levels.  Culture based counts were often two to three orders of magnitude lower than their respective total direct counts, although patterns between caves were similar.   Trends show microbial numbers dropped consistently across all caves with decreasing % culturablity with depth.  This suggests a larger proportion of microbes in the deep aphotic region consist of chemolithotrophic bacteria compared to near surface sites.  A principal components analysis of the 31 potential carbon sources found in the BiOLOG eco-plates show the microbes associated with the interior of the cave group together, irrespective of region, including temperate vs. tropical caves.  This may be the result of chemolithotrophic activity providing carbon sources not available, or at different ratios, than at the mouth of the caves.