48 문성희ㆍ허준욱ㆍ차승주ㆍ황성돈ㆍ손맹현ㆍ권준영ㆍ권세련 SVC (spring viremia of carp) (Goodwin, 2002), 2004 Wisconsin SVCV (Audrey et al., 2004)., 227 (KHV, koi herpes virus),

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1 한수지 51(1), 47-53, 2018 Original Article Korean J Fish Aquat Sci 51(1),47-53, 년소양호에서채집한자연산잉어과어류와갑각류의법정전염병및기생충성질병모니터링 문성희 1 허준욱 2 차승주 3 황성돈 4 손맹현 4 권준영 1 권세련 1,5 * 1 선문대학교수산생명의학과, 2 생물모니터링센터, 3 국립수산과학원내수면양식연구센터, 4 국립수산과학원수산방역과, 5 유전체기반바이오 IT 융합연구소 Monitoring Pathogen Infection of Freshwater Cyprinid Fish and Crustacean in Soyang Lake in 2016 Seong Hee Mun 1, Jun Wook Hur 2, Seung Joo Cha 3, Seong Don Hwang 4, Maeng-Hyun Son 4, Joon Yeong Kwon 1 and Se Ryun Kwon 1,5 * 1 Department of Aquatic Life Medical Sciences, Sunmoon University, Asan 31460, Korea 2 Bio-Monitoring Center, Sejoung 30121, Korea 3 Inland Aquaculture Research Center, National Institute of Fisheries Sciences, Busan 51688, Korea 4 Aquatic Disease Control Division, National Institute of Fisheries Sciences, Busan 46083, Korea 5 Genome-based BioIT Convergence Institute, Asan 31460, Korea The presence of pathogens in wild fish is a potential threat to the fish being raised nearby fish farm. Surveillance of these pathogens in the wild is, thus, highly important to keep fish in the farm safe from serious communicable diseases. Fish and crustacean were sampled 4 times at Soyang Lake in Pathogens for five reportable communicable diseases including KHVD (koi herpesvirus disease), SVC (spring viraemia of carp), EUS (epizootic ulcerative syndrome) and WSD (white spot disease), and parasites were investigated. In Soyang Lake, pale chub Zacco platypus was the most abundant. Forty seven pale chub, 8 Korea piscivorous chub Opsarichthys uncirostris, 3 oily shiner Sarcocheilichthys variegatus wakiyae and 2 crucian carp Cyprinus carassius were investigated for detection of target diseases of fish. Seventeen Caridina denticulata denticulata, 12 Oriental river prawn Macrobrachium nipponense and 1 Chinese mitten crab Eriocheir sinensis were used for detection of WSD. As the result, reportable communicable diseases were not detected in all the freshwater animals. Dactylogyrus was detected in Korea piscivorous chub. Copepoda was also detected in pale chub and Korea piscivorous chub. Metacercaria was detected in the pale chub sampled in September and October. Those were not metacercaria of liver fluke Clonorchis sinensis. Key words: Soyang Lake, Cyprinid fish, wild crustacean, reportable communicable diseases, parasite 서론 (WSD, white spot disease) (WSSV, white spot syndrome virus) (Baumgartner et al., 2009) , (SVCV, spring viremia of carp virus) (Miller et al., 2007) North Carolina Korean J Fish Aquat Sci 51(1) 47-53, February 2018 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Licens ( which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Received 23 November 2017; Revised 21 December 2017; Accepted 10 January 2018 *Corresponding author: Tel: Fax: address: srkwon@sunmoon.ac.kr Copyright 2018 The Korean Society of Fisheries and Aquatic Science 47 pissn: , eissn:

2 48 문성희ㆍ허준욱ㆍ차승주ㆍ황성돈ㆍ손맹현ㆍ권준영ㆍ권세련 SVC (spring viremia of carp) (Goodwin, 2002), 2004 Wisconsin SVCV (Audrey et al., 2004)., 227 (KHV, koi herpes virus), (Fujioka et al., 2015). Fabian et al. (2013) KHVD (koi herpesvirus disease) real-time PCR KHV genome, KHV KHV SVC, ,, SVC.,, ,,,, metacercaria (Lee, 1968) metacercaria,, scalet carp, (Choi, 1976) (Park et al., 2004) , 13,,,, (Yu et al., 1994) % (Kim et al., 2016) , KHVD, SVC EUS (epizootic ulcerative syndrome) WSD.. 재료및방법 생물채집방법 (cast net,, 7 7 mm), (kick net,, 5 5 mm) (fyke, 5 5 mm). 200 m. 20, 60., 24. ( ),.. 채집생물의군집분석 (fish fauna), (dominant species) (subdominant species). (community analysis) (fish species) (DI, dominance index), (H, diversity index), (E, evenness) (R, richness).. (McNaughton, 1967). D = ni/n (D, ; N, ; ni, I ), Margalef (1958) Shannon-Weaver function (Pielou, 1975). H = - Pi Ln Pi (Pi = ni/n)

3 소양호잉어과어류와갑각류의질병모니터링 49, Pielou (1975). E = H'/ln(S) (E, ; H', ; S, ). (S). Margalef (1958). 법정전염병조사 R = (S-1)ln (N) (R, ; S, ; N, ),, ( ),.. PCR. SVC KHV 2, EUS, WSSV. PCR 3 pooling, 5, PCR. RNA TRIsure (Bioline, USA), DNA high pure PCR template preparation kit (ROCHE, Germany). protocol. PCR SVC BIONEER rocketscript RT-PCR kit hotstart PCR kit, KHV EUS Promega Go Taq green mastermix, WSSV Table 1. PCR primer sets and condition for 5 reportable communicable diseases Primer PCR condition Product size SVC KHVD EUS 1 st PCR 2 nd PCR SVCV F1 5 -TCTTGGAGCCAAATAGCTCARRTC-3 (95 1 min, 55 1 min, 72 1 SVCV R2 5 -AGATGGTATGGACCCCAATACATHACNCAY-3 min)x35 cycles min SVCV F1 5 -TCTTGGAGCCAAATAGCTCARRTC-3 (95 1 min, 55 1 min, 72 1 SVCV R4 5 -CTGGGGTTTCCNCCTCAAAGYTGY-3 min)x35 cycles min Bercovier TK TK F 5 -GGGTTACCTGTACGAG min - (95 1 min, 55 1 min, 72 1 min)x40 cycles - 72 TK R 5 -CACCCAGTAGATTATGC-3 10 min Gray Sph Method 1 Method 2 Method 3 GS F 5 -GACACCACATCTGCAAGGAG sec - (94 30 sec, 63 GS R 5 -GACACATGTTACAATGGTCGC-3 30 sec, sec)x40 cycles min Ainvad-2F 5 -TCATTGTGAGTGAAACGGTG min - (95 30 sec, sec, min)x35 cycles - Ainvad-ITSR1 5 -GGCTAAGGTTTCAGTATGTAG min ITS11 5 -GCCGAAGTTTCGCAAGAAAC min - (94 30 sec, ITS23 5 -CGTATAGACACAAGCACACCA-3 sec, 72 1 min)x25 cycles min BO73 5 -CTTGTGCTGAGCTCACACTC min - (96 1 min, 58 1 BO ACACCAGATTACACTATCTC-3 min, 72 1 min)x35 cycles min 714 bp 606 bp 409 bp 292 bp 234 bp 550 bp 564 bp WSD 1 st PCR 2 nd PCR 146F1 5 -ACTACTAACTTCAGCCTATCTAG-3 (94 4 min, 55 1 min, 72 2 min)- (94 1 min, 55 1 min, 1447 bp 146R1 5 -TAATGCGGGTGTAATGTTCTTACGA min)x39 cycles min 146F2 5 -GTAACTGCCCCTTCCATCTCCA-3 (94 4 min, 55 1 min, 72 2 min)- (94 1 min, 55 1 min, 941 bp 146R2 5 -TACGGCAGCTGCTGCACCTTGT min)x39 cycles min SVC, spring viremia of carp; KHVD, koi herpesvirus disease; EUS, epizootic ulcerative syndrome; WSD, white spot disease.

4 50 문성희ㆍ허준욱ㆍ차승주ㆍ황성돈ㆍ손맹현ㆍ권준영ㆍ권세련 Takara EX Taq. PCR mixture protocol. primer PCR Table 1. 기생충성질병조사,., cyst,.,,,,., 37. PS buffer (0.85% NaCl), metacercaria. Metacercaria PCR. primer PCR Cho et al. (2013) (Table 2). 결과및고찰 어류상및상대풍부도 (38.02 N, E) (family) (species) (Cyprinidae) 8, (Bagridae) (Centrarchidae) 2. (Anguillidae), (Siluridae), (Osmeridae), (Centropomidae) (Gobiidae) % %, 13.2% (75 ), 4.9% (28 ), 2.1%, 1.1%, Species Zacco platypus Lepomis macrochirus Hypomesus nipponensis Opsarichthys uncirostris Pseudobagrus fulvidraco Tridentiger brevispinis S. nigripinnis morii Micropterus salmoides Carassius auratus Silurus asotus Leiocassis ussuriensis Hemiculter eigenmanni siniperca scherzeri S.chankaensis tsuchigae Hemibarbus labeo Cyprinus carpio Anguilla japonica Relative abundance (%) Fig. 1. Relative richness index of fishes sampled from the Soyang Lake. Table 2. PCR primer sets and condition for detection of Clonorchis sinensis Primer PCR condition Product size Method 1 RT5-1 5'-ACTTCATCGAGTCATTGGTCGT-3' min - (95 30 sec, 54 1 min, RH3 5'-CGTACTGTAACGCGTTTGTGCA-3' 72 1 min)x30 cycles min 929 bp Method 2 Int5-2 5'-GGACATGCTAACTTCCCTCTCA-3' min - (95 30 sec, 54 1 min, Int3-1 5'-CGTGGTGAATATCGATCAGGTT-3' 72 1 min)x30 cycles min 1349 bp

5 소양호잉어과어류와갑각류의질병모니터링 %, 0.2%. (RA, relative abundance) (Fig. 1), (Zacco platypus) %, %. (Hypomesus nipponensis) 13.2% (75 ), (Opsarichthys uncirostris amurensis) 4.9% (28 ), (Pseudobagrus fulvidraco) 3.9% (22 ), (Tridentiger brevispinis) (Sarcocheilichthys nigripinnis morii) 2.1% (12 ), (Carassius auratus) 1.9% (11 ). (Byeon et al., 1997; Choi et al., 2003; Byeon, 2015),, %. Byeon (2015).. 3, Macrobrachium nipponense % Neocardina denticulata %. 법정전염병감염현황. Table SVC, KHVD, EUS WSD, specific primer PCR RT-PCR (data not shown). 기생충감염현황 Dactylogyrus sp. 1-2 (Fig. 2). 3 4 metacercaria. metacercaria PCR band (data not shown). metacercaria, metacercaria. Metagonimus sp., Echinostoma sp., Centrocestus armarus metacercaria PCR. Park et al. (2004) Table 3. Investigated fishes and crustaceans for detection of target diseases at Soyang Lake in 2016 Date of survey Species Number Mean total length (cm) Mean body weight (g) Pale chub (Zacco platypus) st Korea piscivorous chub (Opsarichthys uncirostris) Oily shiner (Sarcocheilichthys variegatus wakiyae) Pale chub (Z. platypus) nd Chinese mitten crab (Eriocheir sinensis) Oriental river prawn (Macrobrachium nipponense) Pale chub (Z. platypus) rd Crucian carp (Carassius auratus) Caridina denticulata denticulate Oriental river prawn (M. nipponense) Pale chub (Z. platypus) th Korea piscivorous chub (O. uncirostris) C. denticulata denticulate Total number 90

0 10 20 30 40 50 Relative abundance (%) 52

piscivorous chub Opsarichthys uncirostris (B)

Dactylogyrus (C) and copepoda (D) isolated

Copepoda (E) and metacercaia (F & G) isolated

피라미에서 흡충류 피낭유충 감염상태를 조사한 결과에서 빙 어 1마리 당 평균 0.

6 Relative abundance (%) 52 문성희ㆍ허준욱ㆍ차승주ㆍ황성돈ㆍ손맹현ㆍ권준영ㆍ권세련 (A) (B) (C) (E) (D) (F) (G) Fig. 2. The pale chub Zacco platypus (A) and Korea piscivorous chub Opsarichthys uncirostris (B) sampled from the Soyang Lake. Dactylogyrus (C) and copepoda (D) isolated from Korea piscivorous chub. Copepoda (E) and metacercaia (F & G) isolated from pale chub. 피라미에서 흡충류 피낭유충 감염상태를 조사한 결과에서 빙 어 1마리 당 평균 0.35개, 피라미 1마리 당 평균 0.43개의 간흡 충 metacercaria가 감염되어 있다고 보고하였다. 한편 2010년 에 강원도보건환경연구원에서 소양호에서 채집한 빙어를 대상 으로 간흡충 피낭유충을 검사한 결과 모든 빙어 시료에서 피낭 유충이 검출되지 않았다고 보고하였다(NewsWire, 2010). 아울 러 2014년에 소양호에서 포획한 빙어 200마리에서 간흡충 피 낭유충을 검사한 결과 모든 빙어 시료에서 피낭유충이 검출되

7 소양호잉어과어류와갑각류의질병모니터링 53 (Yonhap News, 2014) , KHVD, SVC, EUS WSD, 90. 사사 (R ). References Audrey LD, Craig R, Kathy TK, Susan M, Marty E, Andrew EG, Keith W, David MS and Clare L First report of spring viremia of carp virus (SVCV) in wild common carp in North America. J Aquat Anim Health 16, dx.doi.org/ /h Baumgartner WA, Hawke JP, Bowles K, Varner PW and Hasson KW Primary diagnosis and surveillance of white spot syndrome virus in wild and farmed crawfish (Procambarus clarkii, P. zonangulus) in Louisiana, USA. Dis Aquat Org 85, Byeon HK Fluctuation of introduced fish and characterisitics of the fish community in Lake Soyang. Kor J Environ Ecol 29, KJEE Byeon HK, Jeon SR and Kim DH Ichthyofauna and fish community in Lake Soyang. Kor J Limnol 30, Choi DW Clonorchis sinensis in Kyungpook province, Korea 2. Demonstration of metacercaria of Clonorchis sinensis from fresh-water fish. Kor J Parasitol 14, Choi JS, Lee KY, Jang YS, Ko MH, Kwon OK and Kim BC Study on the dynamics of the fish community in Lake Soyang. Kor J Ichtyol 15, Cho PY, Na BK, Choi KM, Kim JS, Cho SH, Lee WJ, Lim SB, Cha SH, Park YK, Pak JH, Lee HW, Hong SJ and Kim TS Development of a polymerase chain reaction applicable to rapid and sensitive detection of Clonorchis sinensis eggs in human stool samples. Pathog Glob Health 107, Fabian M, Baumer A and Steinhagen D Do wild fish species contribute to the transmission of koi herpesvirus to carp in hatchery ponds? J Fish Dis 36, org/ /jfd Fujioka H, Yamasaki K, Furusawa K, Tamura K, Oguro K, Kurihara S, Seki S, Oshima S and Imajoh M Prevalence and characteristics of Cyprinid herpesvirus 3 (CyHV- 3) infection in common carp (Cyprinus carpio L.) inhabiting three rivers in Kochi Prefecture, Japan. Vet Microbiol 175, Goodwin AE First report of spring viremia of carp virus (SVCV) in North America. J Aquat Anim Health 4, VO>2.0.CO;2. Kim JD, Cho SH, Sohn UM, Song NS, Park JJ, Bang EO, Cha YT and Park KS Infection status of freshwater fishes with metacercariae of Trematodes in the Geum River tributary system. J CNHE 26, Lee JT Studies on the metacercariae from fresh water fishes in the Kum-Ho River. Kor J Parasitol 6, dx.doi.org/ /kjp Margalef R Information theory in ecology. General Systematics 3, McNaughton SJ Relationship among functional properties of California Grassland. Nature 216, dx.doi.org/ /216168b0 Miller O, Fuller FJ, Gebreyes WA, Lewbart GA, Shchelkunov IS, Shivappa RB, Joiner C, Woolford G, Stone DM, Dixon PF, Raley ME and Levine JF Phylogenetic analysis of spring virema of carp virus reveals distinct subgroups with common origins for recent isolates in North America and the UK. Dis Aquat Org 76, dao NewsWire Pond smelt in Soyang Lake and Chuncheon Lake is free from parasites. Retrieved from newswire.co.kr/newsread.php?no= on Jan 21, Park JH, Guk SM, Kim TY, Shin EH, Lin A, Park JY, Kim JL, Hong ST and Chai JY Clonorchis sinensis metacercarial infection in the pond smelt Hypomesus olidus and the minnow Zacco platypus collected from the Soyang and Daechung Lakes. Kor J Parasitol 42, org/ /kjp Pielou EC Ecological Diversity. John Wiley, New York, U.S.A., Yonhap News Masou salmon and pond smelt are free from parasite. Retrieved from lletin/2014/01/09/ akr HTML on Oct 2, Yu JR, Kwon SO and Lee SH Clonorchiasis and metagonimiasis in the inhabitants along Talchongang (River), Chungwon-gun. Kor J Parasitol 32, org/ /kjp

Microsoft Word - 8-Infestation Status of Clonorchis sinensis Metacercariae in Cyprinid Fish from four Artificial Lakes in South

Microsoft Word - 8-Infestation Status of Clonorchis sinensis Metacercariae in Cyprinid Fish from four Artificial Lakes in South JMLS [Journal of Marine Life Science] December 2018; 3(2): 96-100 [ 단보 ] 소양호, 남양호, 안동호및춘천호에서식하는잉어과어류의간흡충 Clonorchis sinensis 감염조사 전상우 1 허준욱 2 송준영 3 지보영 4 이정호 3 권준영 1 권세련 1,5* 1 선문대학교수산생명의학과 2 생물모니터링센터