Skip to main content

First detection of ranavirus in a wild population of Dybowski’s brown frog (Rana dybowskii) in South Korea

Abstract

Background

Ranavirus is an emerging infectious disease which has been linked to mass mortality events in various amphibian species. In this study, we document the first mass mortality event of an adult population of Dybowski’s brown frogs (Rana dybowskii), in 2017, within a mountain valley in South Korea.

Results

We confirmed the presence of ranavirus from all collected frogs (n = 22) via PCR and obtained the 500 bp major capsid protein (MCP) sequence from 13 individuals. The identified MCP sequence highly resembled Frog virus 3 (FV3) and was the same haplotype of a previously identified viral sequence collected from Huanren brown frog (R. huanrenensis) tadpoles in South Korea. Human habitat alteration, by recent erosion control works, may be partially responsible for this mass mortality event.

Conclusion

We document the first mass mortality event in a wild Korean population of R. dybowskii. We also suggest, to determine if ranavirus infection is a threat to amphibians, government officials and researchers should develop continuous, country-wide, ranavirus monitoring programs of Korean amphibian populations.

Background

Emerging infectious diseases are one of the key factors causing rapid global biodiversity declines in this century (Fey et al. 2015). Amphibians are particularly vulnerable to infectious diseases due to their permeable skin and metamorphic life cycle (Daszak et al. 1999). Fungal infections by Batrachochytrium dendrobatidis (Bd) and B. salamandriborans, causing chytridiomycosis, have been implicated as a primary cause of rapid amphibian population declines (Daszak et al. 1999; Scheele et al. 2019). In addition, ranavirus, a double-stranded DNA virus, has also been identified as a major emerging infectious disease and is associated with global amphibian declines (Green et al. 2002; Carey et al. 2003). In Northeast Asia, across China, Japan, and Korea, ranavirus infections have caused mortality in 17, native and invasive, amphibian species (4 urodelan and 13 anuran species) as well as amphibians in the pet trade (Zhang et al. 1996; Weng et al. 2002; Kim et al. 2009; Une et al. 2009a; Kolby et al. 2014; Une et al. 2014; Duffus et al. 2015; Kwon et al. 2017; Park et al. 2017). Out of the 21 reported ranavirus infection cases, 18 have been linked to mass mortality events (Table 1). Amphibian ranavirus susceptibility and mortality are often correlated with low environmental quality, such as habitat destruction and pollution (Carey et al. 1999; Gray et al. 2009; Warne et al. 2011). Additionally, distinct ranavirus strains may have varying virulence and infection capabilities (Miller et al. 2011). Thus, it is imperative to maintain and update infection cases, as well as develop country-level screening protocols, to successfully conserve amphibians at national and global scales (Gray et al. 2009; Miller et al. 2015; García-Díaz et al. 2017).

Table 1 List of amphibian ranavirus infection or mortality cases reported in Northeast Asia. Asterisks represent mass mortality event

Within Korea, studies on amphibian infectious diseases have focused on B. dendrobatidis, the fungal agent causing chytridiomycosis (Yang et al. 2009; O’hanlon et al. 2018). In contrast, studies of ranavirus infections are limited to a handful of reported mortality events between four anuran species including Gold-spotted pond frog (Pelophylax chosenicus) tadpoles (Kim et al. 2009), Huanren brown frog (R. huanrenensis) tadpoles (Kwon et al. 2017), adult Boreal digging frogs (Kaloula borealis), and Japanese tree frog (Hyla japonica) tadpoles (Park et al. 2017). To date, all reported ranavirus strains detected in Korean amphibians share the major capsid protein (MCP) DNA sequence, similar to frog virus 3 (FV3), originally identified from Lithobates pipiens (formerly R. pipiens; Granoff et al. 1965) and Atelognathus patagonicus samples (Fox et al. 2006). To understand the characteristics of ranavirus infections and spread in Korea and across Northeast Asia, it is necessary to determine which viral strains are involved in such mortalities.

In this study, we described a mass mortality event, which occurred in 2017, in a wild population of Dybowski’s brown frogs (R. dybowskii) in South Korea. All sampled R. dybowskii were PCR-positive for ranavirus. Additionally, we determined the strain of ranavirus collected from R. dybowskii samples. This is the first known case of a ranavirus-associated mortality event of adult R. dybowskii in South Korea.

Materials and methods

On March 16, 2017, we found 22 dead adult Dybowski’s brown frogs (R. dybowskii) during a field survey at the upper region of a stream in Moksang-dong, Gyeyang-gu, Incheon, South Korea (37° 33′ 34.05″ N, 126° 42′ 12.79″ E). We collected 22 less decayed dead frogs in individual bags, transported the specimens to the laboratory, and preserved them at −20 °C until future use. In 2014, the stream where the frogs were collected, was heavily modified for erosion control purposes. While collecting dead frog specimens in 2017, we documented that the collection site stream was heavily modified and was lined with stones and concrete along the banks and bottom, and was planted with trees on either side (Fig. 1). Some live adult R. dybowskii individuals were observed within the stream where we collected the dead individuals; however, we were unable to collect any live R. dybowskii due to permitting. At this site, we did not observe any distinctive external symptoms or erratic behaviors, such as loss of buoyancy that were described by previous study (Miller et al. 2015), from individual live frogs.

Fig. 1
figure1

The mountain valley (a) where the mass mortality of Rana dybowskii’s adults (b) occurred, associated with ranavirus-infection at Gyeyang-gu, Incheon, South Korea. Yellow arrowheads indicate dead frogs and in the insert, white arrowheads indicate red skins, a possible symptom of ranavirus infection

Fig. 2
figure2

Bayesian inference (BI) tree based on the 19 partial major capsid protein (MCP) DNA sequences (500 bp) of the ranavirus obtained from dead Rana dybowskii and from GenBank. We used short-finned eel ranavirus as the outgroup. The accession number of iridoviruses, which was obtained from GenBank, indicated after each species name. On each branch, analyzed values (ML bootstrap value/Bayesian posterior probabilities) are denoted

Ranavirus detection

Prior to analysis, samples were slowly defrosted in 10 °C water. Once thawed, we examined any external physical abnormalities under a dissecting microscope (Sunny Optical Technology, China). Liver tissues, which have often been used for ranavirus detection (St-Amour and Lesbarrères 2007), were collected from each frog individual. We extracted whole genomic DNA from 3 to 5 mg of liver tissue using the Qiagen DNeasy® Blood & Tissue Kit (Qiagen, Hilden, Germany). For ranavirus strain identification, the partial sequence of major capsid protein gene (MCP) was amplified using the specific primer pairs (MCP4 and MCP5; Mao et al. 1997). We ran polymerase chain reactions (PCR) following Mao et al. (1997) with a negative control using nuclease-free water and confirmed PCR products on 1% agarose gel by electrophoresis (Mao et al. 1997; Kwon et al. 2017). PCR was run on each sample at least twice to minimize viral false-positive detection.

Finally, PCR products were purified using an AccuPrep® PCR Purification Kit (Bioneer, Daejeon, Korea) and sequenced using the same primer set (Macrogen, Seoul, Korea). Sequences were edited and assembled using Geneious 9.1.8 (Biomatters Ltd., Auckland, New Zealand), and aligned using ClustalW (Thompson et al. 2003) for sequence comparison. For genetic relationship with other iridoviruses, we performed a custom nested BLAST using Geneious 9.1.8 and Bayesian Inference (BI) analysis with 18 ranavirus MCP genes, obtained from GenBank. The TIMef model was selected as the best Akaike information criterion (AIC) scored model after testing 56 nucleotide substitution models in MOTELTEST v3.7 (Posada and Crandall 1998). We analyzed the phylogenetic relationships among the iridoviruses by applying both maximum likelihood (ML) and Bayesian inference (BI) methods in PAUP v4.0 (Swofford 2001) and MrBayes v3.2.47 (Ronquist et al. 2012), respectively. For ML analysis and phylogenetic branching, we applied Bootstrap/Jackknife method, with 1000 bootstraps, and used the tree-bisection-reconnection (TBR) method. The BI analysis with Markov Chain Monte Carlo (MCMC) method was executed using the MrBayes (v3.2.47) software. With four random starting trees, we ran 1,000,000 generations, while sampling every 100 tree generations and discarding the first 5% of the sampled generations as burn-ins. Therefore, 500 of the 10,000 trees sampled were discarded.

Results

We found abdominal inflammation and erythema on the legs of seven collected frog specimens (Fig. 1). All 22 collected frogs were confirmed infected with ranavirus by PCR. Out of 22 PCR products, we obtained 13 partial MCP DNA sequences (> 500 bp) due to low sample quality. The obtained MCP DNA sequences (505 bp) were identical and had 100% sequence similarity to the haplotype (accession number KY264205) collected from a ranavirus-infected Huanren frog (R. huanrenensis; Kwon et al. 2017). In addition, the MCP sequence showed 99.8% similarity with ranavirus KRV-1 (HM133594) from South Korea, Rana catesbeiana virus (AB474588) from Japan, and 99.6% similarity for FV3 (FJ459783) a soft-shelled turtle iridovirus (DQ335253; Table 2). The Bayesian inference (BI) phylogenetic analysis revealed that our sequenced ranavirus grouped with a FV3-like virus including Rana grylio virus (RGV), FV3, and Bohle iridovirus. The nested BLAST analysis was consistent with our phylogenetic results.

Table 2 Results of custom BLAST using partial major capsid protein (MCP) DNA sequence (500 bp) of the ranavirus from Rana dybowskii in this study. The accession numbers of the sequences used for custom BLAST analysis are shown in Fig. 2

Discussion

Diagnosing the physical symptoms of ranavirus infection was often difficult due to the decomposition status of the collected frog samples. Nevertheless, we found abdominal inflammation and erythema on the legs of seven frog specimens. Considering that erythema and skin ulcerations on the legs and ventrum, in amphibians, are known external characteristics of ranavirus infection (Gray et al. 2009; Park et al. 2017), we suspected our specimens might be infected with ranavirus.

Results of our molecular analyses corroborated our suspicions, as all frog specimens were confirmed infected with ranavirus. These results suggest that FV3-like ranavirus infections may be correlated with mass mortality events in populations of adult R. dybowskii. This fact is nothing new, as recent amphibian mass mortality events have been correlated with ranavirus infections across several studies (Weng et al. 2002; Une et al. 2009a; Kwon et al. 2017; Yu et al. 2020). Within the pet trade, ranavirus has been detected in a large number of amphibians, including cases in Hong Kong and Japan (Kolby et al. 2014; Une et al. 2014). Ranavirus strains, detected in the pet trade, were similar to the common midwife toad virus (CMTV) and FV3-like viruses, like the virus detected here. FV3-like viruses have been documented worldwide including Northeast Asia (Kim et al. 2009; Xu et al. 2010), Europe (de Matos et al. 2011), North and South America (Granoff et al. 1965; Fox et al. 2006), and Australia (Hengstberger et al. 1993). To date, at least 11 mass mortality events have been documented in Northeast Asia and were caused by FV3-like viruses (Zhang et al. 1996; Zhou et al. 2012). To this regard, distribution patterns of specific ranavirus strains across Northeast Asia are still under investigation (Duffus et al. 2015). FV3-like viruses have also been found in other taxa such as turtles (Chen et al. 1999) and fish (Ahne et al. 1989), highlighting the importance of ranavirus screening across taxa.

Although the majority of ranavirus-associated amphibian mortalities have occurred in captivity (Table 1; Meng et al. 2014; Mu et al. 2018), there have also been confirmed cases in wild populations. Wild ranavirus-associated mortality events have occurred across Asia, including the Heilongjiang, Jiangxi, and Henan provinces in China (Xu et al. 2010; Chen et al. 2013; Zhu and Wang 2016), the western part of Japan (Une et al. 2009b), and in Gangwon-do, Gyeongsangnam-do, and Daejeon in South Korea (Kwon et al. 2017; Park et al. 2017). Various environmental factors are known to increase ranavirus susceptibility and virulence in amphibians and facilitate mortality (Brunner et al. 2015). In this study, two environmental factors may have contributed to the mortality of adult R. dybowskii. First, the discovery site was heavily altered with concrete for erosion protection 3 years prior, causing water stagnation. Stagnant water during early spring drought periods may contain high concentrations of various ions and pollutants (Kang et al. 2016), possibly increasing stress hormones and making R. dybowskii individuals more susceptible to infection (Gahl and Calhoun 2010; Leduc 2014). In a previous study, dead adult boreal digging frogs were discovered in concrete walled, low circulation waterways (Park et al. 2017), similar to the environment observed in this study. Future studies should determine if surface alterations may influence amphibian-ranavirus susceptibility.

Second, amphibian mortality due to ranavirus has often been correlated with elevated stress hormone levels. Distinct life-history stages, including metamorphosis and reproduction, may be periods where frogs have elevated stress, thus increasing their susceptibility to infection (Green et al. 2002; Duffus et al. 2008; Gray et al. 2009). For example, ranavirus-linked mortality events occurred during the metamorphosis stage of P. chosenicus and during the breeding season of K. borealis (Kim et al. 2009; Park et al. 2017). Rana dybowskii is an explosive breeding species that communally spawns (Yoo and Jang 2012), possibly resulting in elevated stress hormones (Norris and Jones 2012). Thus, highlighting a need to understand how amphibian life history patterns influence viral susceptibility and virulence.

Conclusion

Here, we document the first mass mortality event of R. dybowskii in the wild. All collected individuals were PCR positive for ranavirus, possibly indicating that these individuals died due to viral infection. Elevated stress levels by erosion control works and/or from natural life-history stages may have contributed to ranavirus infection and mortality. To understand if ranavirus infection is a threat to Korean amphibians there are three conservation strategies, which should be implemented. First, there is a need for continuous, country-wide, monitoring of amphibian populations. Second, ranavirus screening should be conducted across various taxa and not relegated to just amphibians. Third, government officials or researchers must identify which environmental factors may increase amphibian susceptibility to ranavirus. By implementing these three strategies, government officials and researchers may be able to successfully protect amphibians from ranavirus infections in Korea and perhaps globally.

Availability of data and materials

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

Abbreviations

AIC:

Akaike information criterion

Bd:

Batrachochytrium dendrobatidis

BI:

Bayesian inference

CMTV:

Common midwife toad virus

FV3:

Frog virus 3

MCP:

Major capsid protein

MCMC:

Markov Chain Monte Carlo

ML:

Maximum likelihood

PCR:

Polymerase chain reaction

RGV:

Rana grylio virus

TBR:

Tree-bisection-reconnection

References

  1. Ahne W, Schlotfeldt HJ, Thomsen I. Fish viruses: isolation of an icosahedral cytoplasmic deoxyribovirus from sheatfish (Silurus glanis). J Vet Med B. 1989;36(1-10):333–6.

    CAS  Article  Google Scholar 

  2. Brunner JL, Storfer A, Gray MJ, Hoverman JT. Ranavirus ecology and evolution: from epidemiology to extinction. In: Gray MJ, Chinchar VG, editors. Ranaviruses. Cham: Springer; 2015. p. 71–104.

    Google Scholar 

  3. Carey C, Bradford DF, Brunner JL, Collins JP, Davidson EW, Longcore JE, et al. Biotic factors in amphibian population declines. In: Linder G, Krest SK, Sparling DW, editors. Amphibian decline: an integrated analysis of multiple stressor effects. Pensacola: SETAC Press; 2003. p. 153–208.

    Google Scholar 

  4. Carey C, Cohen N, Rollins-Smith L. Amphibian declines: an immunological perspective. Dev Comp Immunol. 1999;23(6):459–72.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  5. Chen Z, Gui J, Gao X, Pei C, Hong Y, Zhang Q. Genome architecture changes and major gene variations of Andrias davidianus ranavirus (ADRV). Vet Res. 2013;44(1):1–13.

    Article  Google Scholar 

  6. Chen ZX, Zheng JC, Jiang YL. A new iridovirus isolated from soft-shelled turtle. Virus Res. 1999;63(1-2):147–51.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  7. Daszak P, Berger L, Cunningham AA, Hyatt AD, Green DE, Speare R. Emerging infectious diseases and amphibian population declines. Emerg Infect Dis. 1999;5(6):735.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  8. de Matos APA, da Silva Trabucho MFA, Papp T, Matos BADCA, Correia ACL, Marschang RE. New viruses from Lacerta monticola (Serra da Estrela, Portugal): further evidence for a new group of nucleo-cytoplasmic large deoxyriboviruses. Microsc Microanal. 2011;17(1):101–8.

    PubMed  Article  CAS  PubMed Central  Google Scholar 

  9. Duffus ALJ, Pauli BD, Wozney K, Brunetti CR, Berrill M. Frog virus 3-like infections in aquatic amphibian communities. J Wildlife Dis. 2008;44(1):109–20.

    CAS  Article  Google Scholar 

  10. Duffus ALJ, Waltzek TB, Stöhr AC, Allender MC, Gotesman M, Whittington RJ, et al. Distribution and host range of ranaviruses. In: Gray MJ, Chinchar VG, editors. Ranaviruses. Cham: Springer; 2015. p. 9–57.

    Google Scholar 

  11. Fey SB, Siepielski AM, Nusslé S, Cervantes-Yoshida K, Hwan JL, Huber ER, et al. Recent shifts in the occurrence, cause, and magnitude of animal mass mortality events. P Natl Acad Sci USA. 2015;112(4):1083–8.

    CAS  Article  Google Scholar 

  12. Fox SF, Greer AL, Torres-Cervantes R, Collins JP. First case of ranavirus-associated morbidity and mortality in natural populations of the South American frog Atelognathus patagonicus. Dis Aquat Organ. 2006;72(1):87–92.

    PubMed  Article  PubMed Central  Google Scholar 

  13. Gahl MK, Calhoun AJK. The role of multiple stressors in ranavirus-caused amphibian mortalities in Acadia National Park wetlands. Can J Zool. 2010;88(1):108–21.

    Article  Google Scholar 

  14. García-Díaz P, Ross JV, Woolnough AP, Cassey P. Managing the risk of wildlife disease introduction: pathway-level biosecurity for preventing the introduction of alien ranaviruses. J Appl Ecol. 2017;54(1):234–41.

    Article  Google Scholar 

  15. Geng Y, Wang K, Li C, Wang J, Liao Y, Huang J. Zhou Z. PCR detection and electron microscopic observation of bred Chinese giant salamander infected with ranavirus associated with mass mortality. Vet. Sci China. 2010;40(8):817–21.

    Google Scholar 

  16. Geng Y, Wang KY, Zhou ZY, Li CW, Wang J, He M, et al. First report of a ranavirus associated with morbidity and mortality in farmed Chinese giant salamanders (Andrias davidianus). J Comp Pathol. 2011;145(1):95–102.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  17. Granoff A, Came PE, Rafferty KA Jr. The isolation and properties of viruses from Rana pipiens: their possible relationship to the renal adenocarcinoma of the leopard frog. Ann NY Acad Sci. 1965;126(1):237–55.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  18. Gray MJ, Miller DL, Hoverman JT. Ecology and pathology of amphibian ranaviruses. Dis Aquat Organ. 2009;87(3):243–66.

    PubMed  Article  PubMed Central  Google Scholar 

  19. Green DE, Converse KA, Schrader AK. Epizootiology of sixty-four amphibian morbidity and mortality events in the USA, 1996-2001. Ann NY Acad Sci. 2002;969(1):323–39.

    PubMed  Article  PubMed Central  Google Scholar 

  20. He JG, Lü L, Deng M, He HH, Weng SP, Wang XH, et al. Sequence analysis of the complete genome of an iridovirus isolated from the tiger frog. Virology. 2002;292(2):185–97.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  21. Hengstberger SG, Hyatt AD, Speare R, Coupar BEH. Comparison of epizootic haematopoietic necrosis and Bohle iridoviruses, recently isolated Australian iridoviruses. Dis Aquat Organ. 1993;15(2):93–107.

    Article  Google Scholar 

  22. Kang MJ, Kim KD, Oh KS, Park JW, Park JH. Analysis of forest environmental factors on torrent erosion control work area in Gyeongsangnam-do: focus on erosion control dam and stream conservation. J Agric & Life Sci. 2016;50(5):111–20 (in Korean).

    Article  Google Scholar 

  23. Kim S, Sim MY, Eom AH, Park D, Ra NY. PCR detection of ranavirus in gold-spotted pond frogs (Rana plancyi chosenica) from Korea. Korean J Environ Biol. 2009;27(1):110–3.

    Google Scholar 

  24. Kolby JE, Smith KM, Berger L, Karesh WB, Preston A, Pessier AP, Skerratt LF. First evidence of amphibian chytrid fungus (Batrachochytrium dendrobatidis) and ranavirus in Hong Kong amphibian trade. PloS one. 2014;9(3):e90750.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  25. Kwon S, Park J, Choi WJ, Koo KS, Lee JG, Park D. First case of ranavirus-associated mass mortality in a natural population of the Huanren frog (Rana huanrenensis) tadpoles in South Korea. Anim Cells Syst. 2017;21(5):358–64.

    Article  Google Scholar 

  26. Leduc J. Life-history trade-offs in Northern leopard frog (Lithobates [Rana] pipiens) tadpoles: interactions of trace metals, temperature, and ranavirus. PhD Thesis, Laurentian University of Sudbury; 2014.

    Google Scholar 

  27. Mao J, Hedrick RP, Chinchar VG. Molecular characterization, sequence analysis, and taxonomic position of newly isolated fish iridoviruses. Virology. 1997;229(1):212–20.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  28. Meng Y, Ma J, Jiang N, Zeng LB, Xiao HB. Pathological and microbiological findings from mortality of the Chinese giant salamander (Andrias davidianus). Arch Virol. 2014;159(6):1403–12.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  29. Miller D, Gray M, Storfer A. Ecopathology of ranaviruses infecting amphibians. Viruses. 2011;3(11):2351–73.

    PubMed  PubMed Central  Article  Google Scholar 

  30. Miller D, Pessier A, Hick P, Whittington R. 2015. Comparative pathology of ranaviruses and diagnostic techniques. In: Gray MJ, Chinchar VG. Ranaviruses. Cham: Springer; 2015. p. 171-208.

    Google Scholar 

  31. Mu WH, Geng Y, Yu ZH, Wang KY, Huang XL, Ou YP, et al. FV3-like ranavirus infection outbreak in black-spotted pond frogs (Rana nigromaculata) in China. Microb Pathogenesis. 2018;123:111–4.

    CAS  Article  Google Scholar 

  32. Norris DO, Jones RE. Hormones and reproduction in fishes, amphibians, and reptiles. Berlin: Springer Science & Business Media; 2012.

    Google Scholar 

  33. O’hanlon SJ, Rieux A, Farrer RA, Rosa GM, Waldman B, Bataille A, et al. Recent Asian origin of chytrid fungi causing global amphibian declines. Science. 2018;360(6389):621–7.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  34. Park IK, Koo KS, Moon KY, Lee JG. Park D. PCR detection of ranavirus from dead Kaloula borealis and sick Hyla japonica tadpoles in the wild. Korean. J Herpetol. 2017;8:10–4.

    Google Scholar 

  35. Posada D, Crandall KA. Modeltest: testing the model of DNA substitution. Bioinformatics. 1998;14(9):817–8.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  36. Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Höhna S, et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol. 2012;61(3):539–42.

    PubMed  PubMed Central  Article  Google Scholar 

  37. Scheele BC, Pasmans F, Skerratt LF, Berger L, Martel A, Beukema W, et al. Amphibian fungal panzootic causes catastrophic and ongoing loss of biodiversity. Science. 2019;363(6434):1459–63.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. St-Amour V, Lesbarrères D. Genetic evidence of Ranavirus in toe clips: an alternative to lethal sampling methods. Conserv Genet. 2007;8(5):1247.

    Article  Google Scholar 

  39. Swofford DL. Paup*: Phylogenetic analysis using parsimony (and other methods). Sunderland: Sinauer Associates; 2001. Ver.4.0.b5.

  40. Thompson JD, Gibson TJ, Higgins DG. Multiple sequence alignment using ClustalW and ClustalX. Curr Protoc Bioinformatics. 2003;(1):2–3.

  41. Une Y, Kudo T, Tamukai KI, Murakami M. Epidemic ranaviral disease in imported captive frogs (Dendrobates and Phyllobates spp.), Japan, 2012: a first report. JMM Case Rep. 2014;1(3):e001198.

    Article  Google Scholar 

  42. Une Y, Nakajima K, Taharaguchi S, Ogihara K, Murakami M. Ranavirus infection outbreak in the salamander (Hynobius nebulosus) in Japan. J Comp Pathol. 2009a;4(141):310.

    Article  Google Scholar 

  43. Une Y, Sakuma A, Matsueda H, Nakai K, Murakami M. Ranavirus outbreak in North American bullfrogs (Rana catesbeiana), Japan, 2008. Emerg Infect Dis. 2009b;15(7):1146.

    PubMed  PubMed Central  Article  Google Scholar 

  44. Warne RW, Crespi EJ, Brunner JL. Escape from the pond: stress and developmental responses to ranavirus infection in wood frog tadpoles. Funct Ecol. 2011;25(1):139–46.

    Article  Google Scholar 

  45. Weng SP, He JG, Wang XH, Lü L, Deng M, Chan SM. Outbreaks of an iridovirus disease in cultured tiger frog, Rana tigrina rugulosa. in southern China. J Fish Dis. 2002;25(7):423–7.

    Article  Google Scholar 

  46. Xu K, Zhu DZ, Wei Y, Schloegel LM, Chen XF, Wang XL. Broad distribution of ranavirus in free-ranging Rana dybowskii in Heilongjiang, China. EcoHealth. 2010;7(1):18–23.

    PubMed  Article  PubMed Central  Google Scholar 

  47. Yang H, Baek H, Speare R, Webb R, Park S, Kim T, et al. First detection of the amphibian chytrid fungus Batrachochytrium dendrobatidis in free-ranging populations of amphibians on mainland Asia: survey in South Korea. Dis Aquat Organ. 2009;86(1):9–13.

    PubMed  Article  PubMed Central  Google Scholar 

  48. Yoo E, Jang Y. Abiotic effects on calling phenology of three frog species in Korea. Anim Cells Syst. 2012;16(3):260–7.

    Article  Google Scholar 

  49. Yu Z, Mou W, Geng Y, Wang K, Chen D, Huang X, et al. Characterization and genomic analysis of a ranavirus associated with cultured black-spotted pond frogs (Rana nigromaculata) tadpoles mortalities in China. Transbound Emerg Dis. 2020. https://doi.org/10.1111/tbed.13534.

  50. Zhang QY, Li ZQ, Jiang YL, Liang SC, Gui JF. Preliminary studies on virus isolation and cell infection from disease frog Rana grylio. Acta Hydrobiol Sin. 1996;20(4):390–2 (in Chinese).

    Google Scholar 

  51. Zhang QY, Xiao F, Li ZQ, Gui JF, Mao J, Chinchar VG. Characterization of an iridovirus from the cultured pig frog Rana grylio with lethal syndrome. Dis Aquat Organ. 2001;48(1):27–36.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  52. Zhou ZY, Geng Y, Liu XX, Ren SY, Zhou Y, Wang KY, Huang XL, et al. Characterization of a ranavirus isolated from the Chinese giant salamander (Andrias davidianus, Blanchard, 1871) in China. Aquaculture. 2013;384:66–73.

    Article  CAS  Google Scholar 

  53. Zhou ZY, Geng Y, Ren SY, Wang KY, Huang XL, Chen DF, et al. Ranavirus (family Iridoviridae) detection by polymerase chain reaction (PCR) in Chinese giant salamander (Andrias davidianus, Blanchard, 1871), China. Afr J Biotechnol. 2012;11(85):15130–4.

    CAS  Google Scholar 

  54. Zhu YQ, Wang XL. Genetic diversity of ranaviruses in amphibians in China: 10 new isolates and their implications. Pak J Zool. 2016;48:107–14.

    Google Scholar 

Download references

Acknowledgements

Not applicable

Funding

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (No. 2020R1A6A3A1306094911).

Author information

Affiliations

Authors

Contributions

JP analyzed experimental data and wrote the manuscript draft. IK performed the molecular experiment. NY performed the field survey. NH and AGP reviewed/edited the manuscript. DP designed the study and reviewed/edited the manuscript draft. The authors read and approved the final manuscript.

Corresponding author

Correspondence to Daesik Park.

Ethics declarations

Ethics approval and consent to participate

This study was approved by the Institutional Animal Care and Use Committee in Kangwon National University (KW-200618-3).

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Additional file 1

PCR detection of the MCP sequence of ranavirus from the liver tissues of dead Rana dybowskii’s adults. The numbers on the bands represent individual frogs. P, positive MCP sequence control of ranavirus from Rana huarenensis tadpoles and N, negative control, which used nuclease-free water instead of extracted DNA in PCR process.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Park, J., Grajal-Puche, A., Roh, NH. et al. First detection of ranavirus in a wild population of Dybowski’s brown frog (Rana dybowskii) in South Korea. j ecology environ 45, 2 (2021). https://doi.org/10.1186/s41610-020-00179-2

Download citation

Keywords

  • Ranavirus
  • Rana dybowskii
  • Mass mortality
  • Wild population
  • Major capsid protein