Dataset Information

Development and worldwide use of non-lethal, and minimal population-level impact, protocols for the isolation of amphibian chytrid fungi.

ABSTRACT: Parasitic chytrid fungi have emerged as a significant threat to amphibian species worldwide, necessitating the development of techniques to isolate these pathogens into culture for research purposes. However, early methods of isolating chytrids from their hosts relied on killing amphibians. We modified a pre-existing protocol for isolating chytrids from infected animals to use toe clips and biopsies from toe webbing rather than euthanizing hosts, and distributed the protocol to researchers as part of the BiodivERsA project RACE; here called the RML protocol. In tandem, we developed a lethal procedure for isolating chytrids from tadpole mouthparts. Reviewing a database of use a decade after their inception, we find that these methods have been applied across 5 continents, 23 countries and in 62 amphibian species. Isolation of chytrids by the non-lethal RML protocol occured in 18% of attempts with 207 fungal isolates and three species of chytrid being recovered. Isolation of chytrids from tadpoles occured in 43% of attempts with 334 fungal isolates of one species (Batrachochytrium dendrobatidis) being recovered. Together, these methods have resulted in a significant reduction and refinement of our use of threatened amphibian species and have improved our ability to work with this group of emerging pathogens.

SUBMITTER: Fisher MC

PROVIDER: S-EPMC5958081 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Development and worldwide use of non-lethal, and minimal population-level impact, protocols for the isolation of amphibian chytrid fungi.

Fisher Matthew C MC Ghosh Pria P Shelton Jennifer M G JMG Bates Kieran K Brookes Lola L Wierzbicki Claudia C Rosa Gonçalo M GM Farrer Rhys A RA Aanensen David M DM Alvarado-Rybak Mario M Bataille Arnaud A Berger Lee L Böll Susanne S Bosch Jaime J Bosch Jaime J Clare Frances C FC A Courtois Elodie E Crottini Angelica A Cunningham Andrew A AA Doherty-Bone Thomas M TM Gebresenbet Fikirte F Gower David J DJ Höglund Jacob J James Timothy Y TY Jenkinson Thomas S TS Kosch Tiffany A TA Lambertini Carolina C Laurila Anssi A Lin Chun-Fu CF Loyau Adeline A Martel An A Meurling Sara S Miaud Claude C Minting Pete P Ndriantsoa Serge S O'Hanlon Simon J SJ Pasmans Frank F Rakotonanahary Tsanta T Rabemananjara Falitiana C E FCE Ribeiro Luisa P LP Schmeller Dirk S DS Schmidt Benedikt R BR Skerratt Lee L Smith Freya F Soto-Azat Claudio C Tessa Giulia G Toledo Luís Felipe LF Valenzuela-Sánchez Andrés A Verster Ruhan R Vörös Judit J Waldman Bruce B Webb Rebecca J RJ Weldon Che C Wombwell Emma E Zamudio Kelly R KR Longcore Joyce E JE Garner Trenton W J TWJ

Scientific reports 20180517 1

Parasitic chytrid fungi have emerged as a significant threat to amphibian species worldwide, necessitating the development of techniques to isolate these pathogens into culture for research purposes. However, early methods of isolating chytrids from their hosts relied on killing amphibians. We modified a pre-existing protocol for isolating chytrids from infected animals to use toe clips and biopsies from toe webbing rather than euthanizing hosts, and distributed the protocol to researchers as pa ...[more]

PMID: 29773857

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Project description:While research on frog chytrid fungus Batrachochytrium dendrobatidis (Bd), an infectious disease that threatens amphibian diversity, continues to advance worldwide, little progress has been made in Japan since around 2010. The reason for this is, which we pointed out in 2009, that the origin of frog chytrid fungus may be in the East Asian region, including Japan based on the Bd ITS-DNA variation, and as few cases of mass mortality caused by this fungus have been observed in wild amphibian populations in Japan, the interest of the Japanese government and the general public in Bd has waned. However, we believe that organizing the data obtained so far in Japan and distributing the status of frog chytrid fungus in Japan to the world will provide useful insight for future risk management of this pathogen. We collected more than 5500 swab samples from wild amphibians throughout Japan from 2009 to 2010. Then, we investigated the infection status using the Nested-PCR method. We sequenced the obtained DNA samples and constructed a maximum-parsimony (MP) tree to clarify the phylogenetic diversity of Bd. We detected Bd infection in 11 (nine native and two alien) amphibian species in Japan and obtained 44 haplotypes of Bd ITS-DNA. The MP tree showed a high diversity of Bd strains in Japan, suggesting that some strains belong to Bd-GPL and Bd-Brazil. Except for local populations of the Japanese giant salamanders Andrias japonicus in Honshu Island and the sword tail newts Cynops ensicauda in Okinawa Island, the Bd infection prevalence in native amphibian species was very low. The alien bullfrog Aquaranacatesbeiana had high Bd infection rates in all areas where they were sampled. No Bd infection was detected in other native amphibians in the areas where giant salamanders, sword tail newts, and bullfrogs were collected, suggesting that many native amphibians are resistant to Bd infection. The sword tail newt of Okinawa Island had both the highest infectious incidence and greatest number of haplotypes. The giant salamanders also showed relatively high infection prevalence, but the infected strains were limited to those specific to this species. These two Caudata species are endemic to a limited area of Japan, and it was thought that they may have been refugia for Bd, which had been distributed in Japan Islands for a long time.

Dataset Information

Development and worldwide use of non-lethal, and minimal population-level impact, protocols for the isolation of amphibian chytrid fungi.

Publications

Development and worldwide use of non-lethal, and minimal population-level impact, protocols for the isolation of amphibian chytrid fungi.

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