Genetic evidence for a high diversity and wide distribution of endemic strains of the pathogenic chytrid fungus Batrachochytrium dendrobatidis in wild Asian amphibians

Abstract

Population declines and extinctions of amphibians have been attributed to the chytrid fungus Batrachochytrium dendrobatidis (Bd), especially one globally emerging recombinant lineage ('Bd-GPL'). We used PCR assays that target the ribosomal internal transcribed spacer region (ITS) of Bd to determine the prevalence and genetic diversity of Bd in South Korea, where Bd is widely distributed but is not known to cause morbidity or mortality in wild populations. We isolated Korean Bd strains from native amphibians with low infection loads and compared them to known worldwide Bd strains using 19 polymorphic SNP and microsatellite loci. Bd prevalence ranged between 12.5 and 48.0%, in 11 of 17 native Korean species, and 24.7% in the introduced bullfrog Lithobates catesbeianus. Based on ITS sequence variation, 47 of the 50 identified Korean haplotypes formed a group closely associated with a native Brazilian Bd lineage, separated from the Bd-GPL lineage. However, multilocus genotyping of three Korean Bd isolates revealed strong divergence from both Bd-GPL and the native Brazilian Bd lineages. Thus, the ITS region resolves genotypes that diverge from Bd-GPL but otherwise generates ambiguous phylogenies. Our results point to the presence of highly diversified endemic strains of Bd across Asian amphibian species. The rarity of Bd-GPL-associated haplotypes suggests that either this lineage was introduced into Korea only recently or Bd-GPL has been outcompeted by native Bd strains. Our results highlight the need to consider possible complex interactions among native Bd lineages, Bd-GPL and their associated amphibian hosts when assessing the spread and impact of Bd-GPL on worldwide amphibian populations.

Overview of chytrid emergence and impacts on amphibians

“…Bd is widely distributed throughout Asia [67] and has been present in the region since at least 1911 [68], but no reports exist of disease-driven mortality or population declines [67]. Many distinct genotypes are being described from the regions, including Japan [69], Korea [70] and India [71] as well as Bsal [58]. The growing diversity of chytrid pathogens of amphibians from Asia suggests a likely site of origin for these pathogens.…”

Enhanced call effort in Japanese tree frogs infected by amphibian chytrid fungus

“…Yet, other species, despite themselves harbouring Bd infections, survive in the midst of epizootics affecting amphibian communities. In Asia, enzootic Bd strains commonly are found on amphibians, but hosts typically bear low infection burdens [9]. Asian Bd strains may have had a long historical association with amphibian hosts [10].…”

Batrachochytrium dendrobatidis detected in Kihansi spray toads at a captive breeding facility (Kihansi, Tanzania)

“…Morphological lesions and PCR results confirmed that chytridiomycosis caused the observed mortalities. Preliminary analysis showed that the Bd 5.8S rRNA gene and partial flanking ITS1 and ITS2 nucleotide sequences obtained in this study were not 100% identical to any other Bd nucleotide sequences available in GenBank, although they were very similar to isolates from Ecuador, Japan, USA, Brazil, Korea, and South Africa (James et al 2006, Goka et al 2009, McCracken et al 2009, Schloegel et al 2012, Bataille et al 2013. Possible introduction of Bd into the Udzungwa Mountains arising from stocking of rainbow trout Oncorhynchus mykiss has been speculated, although further fieldwork and re-sampling is necessary before conclusions can be reached about the nature and origin of Bd in this area (Moyer & Weldon 2006).…”

“…A diagnostic polymerase chain reaction (PCR) assay for sensitive and specific detection of Bd has been developed using primers that amplify a region of the 5.8S ribosomal RNA gene and the flanking internal transcribed spacer regions (ITS1 and ITS2; Annis et al 2004). Sequencing of this fragment of the 5.8S rRNA gene has been previously used to infer phylogeny of Bd , Goka et al 2009, Schloegel et al 2012, Bataille et al 2013.…”