ABSTRACT: Volcanic caves are filled with colorful microbial mats on the walls and ceilings. These volcanic caves are found worldwide, and studies are finding vast bacteria diversity within these caves. One group of bacteria that can be abundant in volcanic caves, as well as other caves, is Actinobacteria. As Actinobacteria are valued for their ability to produce a variety of secondary metabolites, rare and novel Actinobacteria are being sought in underexplored environments. The abundance of novel Actinobacteria in volcanic caves makes this environment an excellent location to study these bacteria. Scanning electron microscopy (SEM) from several volcanic caves worldwide revealed diversity in the morphologies present. Spores, coccoid, and filamentous cells, many with hair-like or knobby extensions, were some of the microbial structures observed within the microbial mat samples. In addition, the SEM study pointed out that these features figure prominently in both constructive and destructive mineral processes. To further investigate this diversity, we conducted both Sanger sequencing and 454 pyrosequencing of the Actinobacteria in volcanic caves from four locations, two islands in the Azores, Portugal, and Hawai'i and New Mexico, USA. This comparison represents one of the largest sequencing efforts of Actinobacteria in volcanic caves to date. The diversity was shown to be dominated by Actinomycetales, but also included several newly described orders, such as Euzebyales, and Gaiellales. Sixty-two percent of the clones from the four locations shared less than 97% similarity to known sequences, and nearly 71% of the clones were singletons, supporting the commonly held belief that volcanic caves are an untapped resource for novel and rare Actinobacteria. The amplicon libraries depicted a wider view of the microbial diversity in Azorean volcanic caves revealing three additional orders, Rubrobacterales, Solirubrobacterales, and Coriobacteriales. Studies of microbial ecology in volcanic caves are still very limited. To rectify this deficiency, the results from our study help fill in the gaps in our knowledge of actinobacterial diversity and their potential roles in the volcanic cave ecosystems.
