First detection of the amphibian chytrid fungus Batrachochytrium dendrobatidis in free-ranging populations of amphibians on mainland Asia: survey in South Korea

Abstract

Chytridiomycosis, a disease that has caused amphibian population declines globally and elevated many species of anurans to endangered or threatened status, has recently been declared an internationally notifiable disease. Batrachochytrium dendrobatidis (Bd), the amphibian chytrid fungus causing this disease, has not been previously reported in Korea or on mainland Asia. Thirty-six frog specimens representing 7 species were collected from the wild in South Korea and examined for Bd using standard PCR. Bd was detected in 14 (38.8%) samples from 3 species (Bufo gargarizans, Hyla japonica, and Rana catesbiana). Skin sections from all 14 PCR-positive frogs were examined using 2 staining techniques: haematoxylin and eosin (H&E) and Bd immunoperoxidase (IPX). In histological sections, zoosporangia were found in 6 frogs, with lower sensitivity for H&E (21%) than for IPX (46%). Intensity of infection, based on histopathology, was low in all frogs. These results confirm that Bd is present in South Korea and, hence, on the Asian mainland. Studies are urgently required to determine the impact of chytridiomycosis on Korean amphibians, and to map the distribution of Bd in Korea and other Asian mainland countries. KEY WORDS: Chytridiomycosis · Batrachochytrium dendrobatidis · Amphibian decline · Korea · Fungus Resale or republication not permitted without written consent of the publisherDis Aquat Org 86: [9][10][11][12][13] 2009 nerable (Matsui 2004), and Hynobius yangi is classified as endangered (Stuart 2008) MATERIALS AND METHODSFrogs were collected opportunistically from the northern part of South Korea between June 11 and December 13, 2007 (see Table 1). During nocturnal and occasional diurnal surveys, frogs were captured individually by hand, with a new pair of disposable latex gloves being used for each frog in order to avoid cross-contamination between individuals . The animals were sacrificed by soaking them in a bath of 0.05% aqueous tricaine methane sulfonate (MS-222). We aseptically cut small pieces of skin from the abdomen, inguinal region, and web between toes; we then divided this into 2, and placed 1 in 10% formalin for histological analysis and the other in a 1.5 ml microtube for DNA extraction. Instruments were flamed after dissection of each specimen to prevent cross-contamination. DNA was immediately extracted with Gene Releaser (Bio Ventures). The PCR assay used species-specific primers (Bd1a and Bd2a) located within internal transcribed spacer ITS1 and ITS2 to amplify the 5.8S region of nuclear rDNA (Annis et al. 2004). The PCR products were examined using 1.0% agarose gel and some of the positive bands were cut and sequenced to confirm the Bd sequence. Each sample was tested in triplicate, and was only recorded as positive if all 3 replicates indicated the presence of Bd. For PCR positive amphibians, histological sections of formalin fixed skin were stained using 2 techniques: haematoxylin and eosin (H&E) and Bd immunoperoxidase (IPX), the latter technique using polyclonal antibodies speci...

Mitochondrial DNA Data Unveil Highly Divergent Populations within the Genus Hynobius (Caudata: Hynobiidae) in South Korea

“…Aside from economic growth and development, emerging disease is another great cause threatening amphibian populations (Stauart et al, 2004). Recently, Yang et al (2009) revealed that chytridiomycosis, which can cause severe mortality rates in infected amphibians, was detected in wild populations in South Korea, indicating the need to pay more attention to amphibian conservation and protection. Despite the urgent need to understand these species better to efficiently protect and conserve them, very little information is available on the genus Hynobius thus far.…”

Two amphibian diseases, chytridiomycosis and ranaviral disease, are now globally notifiable to the World Organization for Animal Health (OIE): an assessment

“…The emergence of chytridiomycosis in the UK, for example, was associated with infected, feral, North American bullfrogs (Cunningham et al 2005). Invasive populations of the North American bullfrog have been documented across 4 continents, including Europe, North and South America, and Asia: of these, the majority of populations tested are positive for B. dendrobatidis (Hanselmann et al 2004, Garner et al 2006, Yang et al 2009). …”

Filling a gap in the distribution of Batrachochytrium dendrobatidis: evidence in amphibians from northern China

“…Bd infects at least 695 species of amphibians (anurans, salamanders, and caecilians) from all continents where they exist and has been implicated in the enigmatic disappearance of many frog species in Australia, Europe, and North, Central, and South America (Berger et al 1998, Lips et al 2006, Walker et al 2008, Vredenburg et al 2010, Cheng et al 2011, Soto-Azat et al 2013, Olson & Ronnenberg 2014. However, negative impacts at the population level on other regions where Bd exists, such as Asia and Africa, have not been recorded (Kusrini et al 2008, Goka et al 2009, Yang et al 2009, Bai et al 2010, Soto-Azat et al 2010, Savage et al 2011, Swei et al 2011. One possible explanation is that Bd is endemic in some areas but novel in others (Rosenblum et al 2013).…”

“…In Asia, Bd has been described infecting wild and captive amphibians from many countries, including China, Japan, South Korea, Indonesia, Malaysia, Laos, Philippines, Sri Lanka, Kyrgyzstan, Vietnam, and India (Kusrini et al 2008, Une et al 2008, Goka et al 2009, Yang et al 2009, Bai et al 2010, Savage et al 2011, Swei et al 2011, Bataille et al 2013, Dahanukar et al 2013, Zhu et al 2014a). Nevertheless, extensive areas of central, northern, and western mainland Asia still lack information on the presence and impacts of Bd (Olson et al 2013, Olson & Ronnenberg 2014.…”