Antifungal Bacteria on Woodland Salamander Skin Exhibit High Taxonomic Diversity and Geographic Variability

Abstract

ABSTRACT Diverse bacteria inhabit amphibian skin; some of those bacteria inhibit growth of the fungal pathogen Batrachochytrium dendrobatidis . Yet there has been no systematic survey of anti- B. dendrobatidis bacteria across localities, species, and elevations. This is important given geographic and taxonomic variations in amphibian susceptibility to B. dendrobatidis . Our collection sites were at locations within the Appalachian Mountains where previous sampling had indicated low B. dendrobatidis prevalence. We determined the numbers and identities of anti- B. dendrobatidis bacteria on 61 Plethodon salamanders (37 P. cinereus , 15 P. glutinosus , 9 P. cylindraceus ) via culturing methods and 16S rRNA gene sequencing. We sampled co-occurring species at three localities and sampled P. cinereus along an elevational gradient (700 to 1,000 meters above sea level [masl]) at one locality. We identified 50 anti- B. dendrobatidis bacterial operational taxonomic units (OTUs) and found that the degree of B. dendrobatidis inhibition was not correlated with relatedness. Five anti- B. dendrobatidis bacterial strains occurred on multiple amphibian species at multiple localities, but none were shared among all species and localities. The prevalence of anti- B. dendrobatidis bacteria was higher at Shenandoah National Park (NP), VA, with 96% (25/26) of salamanders hosting at least one anti- B. dendrobatidis bacterial species compared to 50% (7/14) at Catoctin Mountain Park (MP), MD, and 38% (8/21) at Mt. Rogers National Recreation Area (NRA), VA. At the individual level, salamanders at Shenandoah NP had more anti- B. dendrobatidis bacteria per individual (μ = 3.3) than those at Catoctin MP (μ = 0.8) and at Mt. Rogers NRA (μ = 0.4). All salamanders tested negative for B. dendrobatidis . Anti- B. dendrobatidis bacterial species are diverse in central Appalachian Plethodon salamanders, and their distribution varied geographically. The antifungal bacterial species that we identified may play a protective role for these salamanders. IMPORTANCE Amphibians harbor skin bacteria that can kill an amphibian fungal pathogen, Batrachochytrium dendrobatidis . Some amphibians die from B. dendrobatidis infection, whereas others do not. The bacteria that can kill B. dendrobatidis , called anti- B. dendrobatidis bacteria, are thought to influence the B. dendrobatidis infection outcome for the amphibian. Yet how anti- B. dendrobatidis bacterial species vary among amphibian species and populations is unknown. We determined the distribution of anti- B. dendrobatidis bacterial species among three salamander species ( n = 61) sampled at three localities. We identified 50 unique anti- B. dendrobatidis bacterial species and found that all of the tested salamanders were negative for B. dendrobatidis . Five anti- B. dendrobatidis bacterial species were commonly detected, suggesting a stable, functional association with these salamanders. The number of anti- B. dendrobatidis bacteria per individual varied among localities but not among co-occurring salamander species, demonstrating that environment is more influential than host factors in structuring the anti- B. dendrobatidis bacterial community. These anti- B. dendrobatidis bacteria may serve a protective function for their salamander hosts.