At least two-thirds of commercial antibiotics today are derived from Actinobacteria, more specifically from the genus Streptomyces. Antibiotic resistance and new emerging diseases pose great challenges in the field of microbiology. Cave systems, in which actinobacteria are ubiquitous and abundant, represent new opportunities for the discovery of novel bacterial species and the study of their interactions with emergent pathogens. White-nose syndrome is an invasive bat disease caused by the fungus Pseudogymnoascus destructans, which has killed more than six million bats in the last 7 years. In this study, we isolated naturally occurring actinobacteria from white-nose syndrome (WNS)-free bats from five cave systems and surface locations in the vicinity in New Mexico and Arizona, USA. We sequenced the 16S rRNA region and tested 632 isolates from 12 different bat species using a bilayer plate method to evaluate antifungal activity. Thirty-six actinobacteria inhibited or stopped the growth of P. destructans, with 32 (88.9%) actinobacteria belonging to the genus Streptomyces. Isolates in the genera Rhodococcus, Streptosporangium, Luteipulveratus, and Nocardiopsis also showed inhibition. Twenty-five of the isolates with antifungal activity against P. destructans represent 15 novel Streptomyces spp. based on multilocus sequence analysis. Our results suggest that bats in western North America caves possess novel bacterial microbiota with the potential to inhibit P. destructans. IMPORTANCE This study reports the largest collection of actinobacteria from bats with activity against Pseudogymnoascus destructans, the fungal causative agent of white-nose syndrome. Using multigene analysis, we discovered 15 potential novel species. This research demonstrates that bats and caves may serve as a rich reservoir for novel Streptomyces species with antimicrobial bioactive compounds.KEYWORDS Actinobacteria, bats, caves, Pseudogymnoascus, Streptomyces, white-nose syndrome P resently, 90% of antibiotics are derived from microorganisms within the phylum Actinobacteria (1-3). The family Streptomycetaceae is particularly known for the production of chitinases capable of hydrolyzing the cell wall (4) and targeting ergosterol in the cell membrane of fungi (5). The rapid development of antibiotic resistance and the emergence of infectious diseases in humans and other animals, including bats and amphibians, have prompted the study of the bacterial microbiome and use for probiotic treatment in vertebrates (6, 7).

Microbial mats are a prominent feature in many Hawaiian lava caves, but little research has been done on these communities. Since 2008, we have sampled 16 lava caves on the Big Island of Hawai`i for microbial communities for scanning electron microscopy (SEM) analysis, cultivation, and DNA sequencing. These caves occurred in areas of Hawai`i that varied in rainfall from 47-401 cm per year. Sampled communities included microbial mats of various colors from white to tan, yellow, and orange; white mats floating on puddles in the floor; and butterscotch-colored organic ooze. We also sampled "microbes that masquerade as minerals" to determine whether mineral deposits contained substantial microorganisms. SEM studies revealed diverse morphologies across the lava caves, with coccoid and filamentous shapes predominating. Culture media inoculated with microbial mat or mineral deposits on site in Hawaiian lava caves revealed morphologies consistent with Actinobacteria and many cultures demonstrated the presence of fugitive dyes that were aqueously soluble. DNA analysis revealed that the white wall microbial mats differed from the yellow, pink, and orange mats, which were more similar to each other. Actinobacteria dominated the latter deposits. Overall, the type of sample (mat versus mineral versus surface soil) made the greatest composition difference.

Background Antibiotic-producing Streptomyces bacteria are ubiquitous in nature, yet most studies of its diversity have focused on free-living strains inhabiting diverse soil environments and those in symbiotic relationship with invertebrates. Results We studied the draft genomes of 73 Streptomyces isolates sampled from the skin (wing and tail membranes) and fur surfaces of bats collected in Arizona and New Mexico. We uncovered large genomic variation and biosynthetic potential, even among closely related strains. The isolates, which were initially identified as three distinct species based on sequence variation in the 16S rRNA locus, could be distinguished as 41 different species based on genome-wide average nucleotide identity. Of the 32 biosynthetic gene cluster (BGC) classes detected, non-ribosomal peptide synthetases, siderophores, and terpenes were present in all genomes. On average, Streptomyces genomes carried 14 distinct classes of BGCs (range = 9–20). Results also revealed large inter- and intra-species variation in gene content (single nucleotide polymorphisms, accessory genes and singletons) and BGCs, further contributing to the overall genetic diversity present in bat-associated Streptomyces. Finally, we show that genome-wide recombination has partly contributed to the large genomic variation among strains of the same species. Conclusions Our study provides an initial genomic assessment of bat-associated Streptomyces that will be critical to prioritizing those strains with the greatest ability to produce novel antibiotics. It also highlights the need to recognize within-species variation as an important factor in genetic manipulation studies, diversity estimates and drug discovery efforts in Streptomyces.

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