Action: Use antifungal skin bacteria or peptides to reduce chytridiomycosis infection
- Three of four randomized, replicated, controlled studies in the USA found that adding antifungal bacteria to the skin of salamanders or frogs exposed to the chytrid fungus did not reduce chytridiomycosis infection rate or death. One found that adding antifungal bacteria to frogs prevented infection and death. One randomized, replicated, controlled study in the USA found that adding antifungal skin bacteria to soil significantly reduced chytridiomycosis infection rate of red-backed salamanders.
- One randomized, replicated, controlled study in Switzerland found that treatment with antimicrobial skin peptides before or after infection with chytridiomycosis did not significantly increase survival of common toads.
- Three randomized, replicated, controlled studies in the USA found that adding antifungal skin bacteria to chytrid infected amphibians reduced weight loss.
The chytrid fungus Batrachochytrium dendrobatidis infects the outer layer of amphibian skin. A number of bacterial species of amphibian skin have been found to inhibit the chytrid fungus in experiments (e.g. Harris et al. 2006; Becker et al. 2010; Lam et al. 2011). There is also some evidence that anti-microbial peptides, which are secreted into mucus and thought to help protect against colonization by skin pathogens, may provide some resistance to chytrid infections (e.g. Pask et al. 2012; 2013). It is therefore possible that adding such anti-fungal species or peptides to amphibian skin or to their environment may reduce the effects of the disease.
Becker, M. H. & Harris, R.N. 2010. Cutaneous bacteria of the redback salamander prevent morbidity associated with a lethal disease. PLoS One, 5, e10957.
Harris R.N., James T.Y., Lauer A., Simon M.A. & Patel A. (2006) The amphibian pathogen Batrachochytrium dendrobatidis is inhibited by the cutaneous bacteria of amphibian species. EcoHealth, 3, 53–56.
Lam B.A., Walton D.B. & Harris R.N. (2011) Motile zoospores of Batrachochytrium dendrobatidis move away from antifungal metabolites produced by amphibian skin bacteria. EcoHealth, 8, 36–45.
Pask J.D., Cary T.L. & Rollins-Smith L.A. (2013) Skin peptides protect juvenile leopard frogs (Rana pipiens) against chytridiomycosis. Journal of Experimental Biology, 216, 2908–2916.
Pask J.D., Woodhams D.C. & Rollins-Smith L.A. (2012) The ebb and flow of antimicrobial skin peptides defends northern leopard frogs (Rana pipiens) against chytridiomycosis. Global Change Biology, 18, 1231–1238.
Supporting evidence from individual studies
A randomized, replicated, controlled study in a laboratory in California, USA (Harris et al. 2009) found that adding antifungal bacteria (Janthinobacterium lividum) to the skins of mountain yellow-legged frog Rana muscosa prevented death from chytridiomycosis. Infected frogs treated with the antifungal skin bacteria all survived, gained 33% body mass and had no chytrid zoospores on their skin. In contrast, five of six exposed to chytrid zoospores alone lost weight and died; the sixth had severe chytridiomycosis. Treatment with Janthinobacterium lividum increased colonization by the skin bacteria and did not result in reduced growth or death. There were three treatments each with six frogs: exposure to chytrid zoospores (300 zoospores/15 ml for 24 h); exposure to antifungal skin bacteria (26 x 106 cells/ml for 30 min) and exposure to skin bacteria and 48 hours later chytrid zoospores. There were also 10 untreated control frogs. Before treatments, animals were rinsed in 3% hydrogen peroxide and sterile Provosoli medium to reduce natural skin bacteria. Frogs were weighed and tested for antifungal skin bacteria and chytrid before and every two weeks after treatment until day 139.
A randomized, replicated, controlled study in a laboratory in Virginia, USA (Harris et al. 2009) found that the severity, but not the infection rate, of chytridiomycosis was reduced by adding chytrid-inhibiting skin bacteria to the skin of red-backed salamanders Plethodon cinereus. Infection rate did not differ significantly between those with added bacteria (Pseudomonas reactans; 80%) and those with chytrid alone (60%). Numbers of zoospore equivalents on infected individuals were also similar (with bacteria: 6; chytrid alone: 10). However, by day 46, salamanders with the bacteria had lost significantly less body mass (15%) than those with chytrid alone (30%) and a similar amount to controls (bacteria or medium alone: 8%). Following inoculation with skin bacteria, 89% of 18 individuals tested positive for the bacteria. Individuals were randomly assigned to one of four exposure treatments: anti-chytrid skin bacteria, chytrid zoospores, bacteria followed by chytrid zoospores three days later or solution alone. Sample sizes were 5, 20, 20 and 5 respectively. Individuals were tested for chytrid on day 1 and 14 and for skin bacteria on day 1 and 10. Salamanders were bathed with 5 ml of solution containing bacteria (3 x 109 cells/ml) for two hours and/or a solution with chytrid (3 x 106 zoospores/5 ml) for 24 hours.
A randomized, replicated, controlled study in a laboratory in the USA (Becker et al. 2011) found that although the chytrid-inhibiting skin bacteria Janthinobacterium lividum colonized skin temporarily, it did not reduce or delay death of chytrid infected Panamanian golden frogs Atelopus zeteki. All infected frogs died within four months, whereas all control frogs survived. Although mortality and overall chytrid load did not differ between frogs exposed and not exposed to the bacteria, at death those exposed had significantly lower numbers of chytrid zoospores (1.5 x 105 vs 1.3 x 106). Colonization by the bacteria was successful on 95% of frogs. However, by day 39 bacterial cell counts had declined (<2.8 x 105 cells/frog), infection with chytrid had increased (>13,000 zoospore equivalents/frog) and frogs began to die. Frogs were randomly assigned to one of four exposure treatments: anti-chytrid skin bacteria, chytrid zoospores, bacteria followed by chytrid or water alone. Sample sizes were 7, 20, 20 and 7 respectively. Bacteria were isolated from four-toed salamanders Hemidactylium scutatum. Frogs were swabbed every two weeks for 120 days to test for chytrid and bacteria.
A randomized, replicated, controlled study in 2010 in a laboratory in Virginia, USA (Muletz et al. 2012) found that infection rate of red-backed salamanders Plethodon cinereus with chytridiomycosis was significantly lower following exposure to chytrid-inhibiting skin bacteria in the soil. Infection rate was 40% with exposure to the bacteria Janthinobacterium lividum compared to 83% without. All salamanders exposed tested positive for the skin bacteria up until day 29, but by day 42 it was no longer detected. Salamanders infected with chytrid had significantly higher densities of bacteria than uninfected individuals. Fifteen randomly selected wild caught salamanders were exposed to skin bacteria in soil followed by chytrid in solution. Twelve were exposed to chytrid alone, six to skin bacteria in soil alone and five were unexposed controls. Each tank received 150 g of soil, which had 1.5 ml of skin bacteria suspension (2.9 x 107 colony-forming units/dry g soil) or pond water. Janthinobacterium lividum was isolated from the skin of four-toed salamanders Hemidactylium scutatum. Salamanders were tested for chytridiomycosis and the skin bacteria on days 8, 13, 20, 29 and 42.
A randomized, replicated, controlled study in 2007 in a laboratory in Virginia, USA (Woodhams et al. 2012) found that survival of mountain yellow-legged frogs Rana muscosa naturally infected with chytridiomycosis was not increased by adding chytrid-inhibiting skin bacteria. Survival of frogs treated with bacteria was 50% compared to 39% for infected controls. Infection was not cleared in surviving frogs. However, weight loss was reduced with treatment (0.1 vs 0.4 g/week). Wild-caught frogs were randomly assigned to treatments. Twenty were bathed in water containing bacteria (Pedobacter cryoconitis) isolated from mountain yellow-legged frog and 13 control frogs in water alone for two hours. Frogs were swabbed and tested at seven and 13 days after treatment.
A randomized, replicated, controlled study in 2010 in a laboratory in Switzerland (Woodhams, Geiger, Reinert, Rollins-Smith, Lam, Harris, Briggs, Vredenburg & Voyles 2012) found that survival of common toad Bufo bufo toadlets was not significantly increased by treatment with antimicrobial skin peptides before or after infection with chytridiomycosis, although treatment may have cured infection in some individuals. Survival of toads treated with peptides immediately before or eight days after infection was not significantly different from chytrid infected controls (12 vs 18%). However, none of the three treated toadlets that survived to 35 days were infected with chytridiomycosis, compared to all three of the untreated infected controls. Peptide treatment alone did not reduce survival compared to uninfected controls (64% vs 58%). Captive toadlets were randomly assigned to treatments. Seventeen were infected with chytridiomycosis alone. Seventeen were treated with skin peptides from edible frogsPelophylax esculentus (2 minute bath in 400 μg/ml peptide solution) immediately before infection and 17 on day eight following infection. Twenty four were uninfected controls, 12 of which were bathed with peptides. Swabs were taken and tested for the chytrid fungus on day 35.
- Harris R.N., Brucker R.M., Walke J.B., Becker M.H., Schwantes C.R., Flaherty D.C., Lam B.A., Woodhams D.C., Briggs C.J., Vredenburg V.T. & Minbiole K.P.C. (2009) Skin microbes on frogs prevent morbidity and mortality caused by a lethal skin fungus. The ISME Journal, 3, 818-824
- Harris R.N., Lauer A., Simon M.A., Banning J.L. & Alford R.A. (2009) Addition of antifungal skin bacteria to salamanders ameliorates the effects of chytridiomycosis. Diseases of Aquatic Organisms, 83, 11–16
- Becker M.H., Harris R.N., Minbiole K.P.C., Schwantes C.R., Rollins-Smith L.A., Reinert L.K., Brucker R.M., Domangue R.J. & Gratwicke B. (2011) Towards a better understanding of the use of probiotics for preventing chytridiomycosis in Panamanian golden frogs. EcoHealth, 8, 501-506
- Muletz C.R., Myers J.M., Domangue R.J., Herrick J.B. & Harris R.N. (2012) Soil bioaugmentation with amphibian cutaneous bacteria protects amphibian hosts from infection by Batrachochytrium dendrobatidis. Biological Conservation, 152, 119–126
- Woodhams D.C., Geiger C.C., Reinert L.K., Rollins-Smith L.A., Lam B., Harris R.N., Briggs C.J., Vredenburg V.T. & Voyles J. (2012) Treatment of amphibians infected with chytrid fungus: learning from failed treatments with itraconazole, antimicrobial peptides, bacteria, and heat therapy. Diseases of Aquatic Organisms, 98, 11-25