한수지 51(6), 673-681, 2018 Original Article Korean J Fish Aquat Sci 51(6),673-681,2018 용남광도해역의굴 (Crassostrea gigas) 및육상오염원에서분리한대장균 (Escherichia coli) 의항생제내성 권순재 정연중 윤현진 목종수 1 권지영 * 국립수산과학원남동해수산연구소, 1 국립수산과학원식품위생가공과 Antimicrobial Resistance in Escherichia coli Isolated from Oyster Crassostrea gigas and Inland Pollution Sources in the Yongnam-Gwangdo Area, Korea Soon Jae Kwon, Yeoun Joong Jung, Hyun Jin Yoon, Jong Soo Mok 1 and Ji Young Kwon* Southeast Sea Fisheries Research Institute, National Institute of Fisheries Science, Tongyeong 53085, Korea 1 Food Safety and Processing Research Division, National Institute of Fisheries Science, Busan 40683, Korea We isolated and characterized Escherichia coli (E. coli) from oyster Crassostrea gigas and inland pollution sources on Yongnam-Gwangdo island and neighboring areas on the southern coast of Korea in 2014-2015. A total of 222 strains of E. coli were isolated from 132 oysters and 88 samples from inland pollution sources. These 222 isolates were tested for their susceptibility to 24 antimicrobial agents, and 221 isolates were found resistant to the tested antibiotics. Of these 99.5% and 70.7% of the isolates showed very high resistance to rifampin and cephalothin respectively, followed by tobramycin (23.4%), streptomycin (20.2%), tetracycline (19.4%), cefepime (18.9%), ceftazidime (18.9%) and nalidixic acid (16.7%). The resistance rate of E. col isolated from oysters was higher than that from inland pollution sources. In addition, multiple resistance to at least four antibiotics were present in 73.2% and 26.5% of E. coli isolates from oysters and inland pollution source samples, respectively. Key words: Oyster, Antimicrobial resistance, Inland pollution source, Escherichia coli 서론 (Todd and Campbell, 2002). Escherichia coli, (Song et al., 2004). (Crassostrea gigas) (Sobsey and Jaykus, 1991).,., (Son et al., 2003). (Angulo et al., 2000; Donabedian et al., 2003).. *Corresponding author: Tel: +82. 55. 640. 4760 Fax: +82. 55. 641. 2036 E-mail address: kjy3t3@korea.kr This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Licens (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. https://doi.org/10.5657/kfas.2018.0673 Korean J Fish Aquat Sci 51(6), 673-681, December 2018 Received 20 November 2018 Revised 5 December 2018; Accepted 20 December 2018 저자직위 : 권순재 ( 인턴연구원 ), 정연중 ( 해양수산연구사 ), 윤현진 ( 인턴연구원 ), 목종수 ( 해양수산연구관 ), 권지영 ( 해양수산연구사 ) Copyright 2018 The Korean Society of Fisheries and Aquatic Science 673 pissn:0374-8111, eissn:2287-8815
674 권순재ㆍ정연중ㆍ윤현진ㆍ목종수ㆍ권지영,.., (Watts et al., 2017)., 70% (Kummerer, 2009). (Witte, 2000)., (Souli et al., 2008).,. 2015 91 49 54% 40 213,113 (NIFS, 2017).,.,. 재료및방법 시료채취및운반 2014 1 2015 12 1 5 (Fig. 1). 132 whirl-pak (Nasco International Inc., USA). 11, 2014 1 2015 12 1 8 88.. 10 Fig. 1. Sampling stations of oyster Crassostrea gigas ( ) and inland pollution sources ( ) in the Yongnam-Gwangdo area, Korea.. 대장균 (E. coli) 분리및동정 E. coli The recommended procedures for the examination of seawater and shellfish (APHA, 1970). (phosphate buffer solution),. 10 ml lauryl tryptose broth (LTB; Difco, Detroit, MI, USA) (35 0.5, 48 ), loop (10 L) 10 ml EC broth (EC; Difco, Detroit, MI, USA), (44.5, 24 ). EC broth tryptone bile X-glucuronide Table 1. Number of E. col isolated from oyster Crassostrea gigas and inland pollution sources in the Yongnam-Gwangdo area Samples No. of samples No. of isolated strains Oyster 132 56 Inland pollution source 88 166 Total 220 222
분리된대장균 (Escherichia coli) 의항생제내성 675 agar (TBX; Oxoid, Basingstoke, UK) streak (44 1 22 2 ), E. coli VITEK system (Biomerieux, Marcy, France),. 항생제내성시험 E. coli Acar and Goldstein (1991)., E. coli muller hinton broth (MHB; BBL, NJ, USA) 35 18-24, densicheck plus (Biomerieux, Marcy, France) 0.5. 4 mm muller hinton II agar (MHA; BBL, NJ, USA) 5 15 (Sensi-disc, BBL, NJ, USA). amikacin (30 g; AN), amoxicillin/ clavulanic acid (30 g; AmC), ampicillin (10 g; AM), aztreonam (30 g; ATM), cefamandole (30 g; MA), cefazolin (30 g; CZ), cefepime (30 g; FEP), cefotaxime (30 g; CTX), cefotetan (30 g; CTT), cefoxitin (30 g; FOX), ceftazidime (30 g; CAZ), cephalothin (30 g; CF), chloramphenicol (30 g; C), ciprofloxacin (5 g; CIP) gentamicin (10 g; GM), imipenem (10 g; IPM), nalidixic acid (30 g; NA), piperacillin (100 g; PIP), rifampin (5 g; RA), streptomycin (10 g; S), tetracycline (30 g; TE), tobramycin (10 g; NN), trimethoprim (5 g; TMP), trimethoprim/ sulfamethoxazole (1.25/ 23.75 g; SXT) 24. muller hinton II Agar 35 16-18, (inhibition zone). 결과및고찰 대장균의항생제내성 2014 1 2015 12 Table 2. Antimicrobial resistance of E. col isolated in the Yongnam-Gwangdo area Antimicrobial agents Concentration per disk (μg) Diffusion zone breakpoint (mm) No. of resistance isolates (%) Escherichia coli (n=222) Amikacin (AN) 30 <14 31 (13.9) Amoxicillin/Clavulanic acid (AMC) 20/10 <13 6 (2.7) Ampicillin (AM) 10 <13 10 (4.5) Aztreonam (ATM) 30 <10 5 (2.5) Cefamandole (MA) 30 <14 17 (7.7) Cefazolin (CZ) 30 <14 12 (5.4) Cefepime (FEP) 30 <14 42 (18.9) Cefotetan (CTT) 30 <12 1 (0.4) Cefotaxine (CTX) 30 <14 7 (3.2) Ceftazidime (CAZ) 30 <14 42 (18.9) Cephalothin (CF) 30 <14 157 (70.7) Cefoxitin (FOX) 30 <14 6 (2.7) Chloramphenico (C) 30 <12 15 (6.8) Ciprofloxacin (CIP) 5 <15 11 (5.0) Gentamicin (GM) 10 <12 14 (6.3) Imipenem (IPM) 10 <13 0 (0.0) Nalidixic acid (NA) 30 <13 37 (16.7) Piperacillin (PIP) 10 <17 32 (14.4) Rifampin (RA) 5 <17 221 (99.5) Streptomycin (S) 10 <11 45 (20.2) Tetracycline (TE) 19 <14 43 (19.4) Trimethoprim/ Sulfamethoxazole (SXT) 23.75/1.25 <10 16 (7.2) Tobramycin (NN) 10 <12 52 (23.4) Trimethoprim (TMP) 5 <10 18 (8.1)
676 권순재ㆍ정연중ㆍ윤현진ㆍ목종수ㆍ권지영 E. coli 56, ( 10, 1 ) 166 (Table 1), E. coli 222 24 (Table 2), rifampin (99.5%) cephalothin (70.7%), tobramycin (23.4%), streptomycin (20.2%), tetracycline (19.4%), cefepime (18.9%), ceftazidime (18.9%), nalidixic acid (16.7%), piperacillin (14.4%), amikacin (13.9%). trimethoprim, cefamandole, trimethoprim/ sulfamethoxazole, chloramphenicol, gentamicin, cefazolin. ciprofloxacin, ampicillin, cefoxitin, amoxicillin/ clavulanic acid, cefoxitim, aztreonam, cefotetan 10%, imipenem., 2011 2012 3 (,, ) E. coli tetracycline (29.9%), streptomycin (25.5%), ampicillin (18.6%) (Park et al., 2013), 2013 (,, ) E. coli tetracycline (10.6%) 5% (Park et al., 2018). 2013 2015 E. coli ampicillin (37.2%), cephalothin (21.7%), cefazolin (19.9%) (Jo et al., 2016) tetracycline 4% cephem. 2014 2015 E. coli ansamycin rifampin cephem cephalothin, tetracycline penicillin (MFDS, 2018) 2014 Table 3. Antimicrobial resistance of E. col isolated from oyster Crassostrea gigas and inland pollution sources in Yongnam-Gwangdo area Antimicrobial agents Concentration per disk (μg) Diffusion zone breakpoint (mm) No. of resistance isolates (%) Oyster Inland pollution source E. coli (n=56) E. coli (n=166) Amikacin (AN) 30 <14 2 (3.6) 8 (4.8) Amoxicillin/Clavulanic acid (AMC) 20/10 <13 1 (1.8) 5 (3.0) Ampicillin (AM) 10 <13 8 (14.3) 23 (13.9) Aztreonam (ATM) 30 <10 0 (0.0) 5 (3.0) Cefamandole (MA) 30 <14 0 (0.0) 17 (10.2) Cefazolin (CZ) 30 <14 0 (0.0) 5 (3.0) Cefepime (FEP) 30 <14 40 (71.4) 2 (1.2) Cefotetan (CTT) 30 <12 1 (1.8) 0 (0.0) Cefotaxine (CTX) 30 <14 0 (0.0) 7 (4.2) Ceftazidime (CAZ) 30 <14 40 (71.4) 2 (1.2) Cephalothin (CF) 30 <14 41 (73.2) 116 (69.9) Cefoxitin (FOX) 30 <14 0 (0.0) 6 (3.6) Chloramphenico (C) 30 <12 6 (10.7) 9 (5.4) Ciprofloxacin (CIP) 5 <15 1 (1.8) 10 (6.0) Gentamicin (GM) 10 <12 2 (3.6) 12 (7.2) Imipenem (IPM) 10 <13 0 (0.0) 0 (0.0) Nalidixic acid (NA) 30 <13 7 (12.5) 30 (18.1) Piperacillin (PIP) 10 <17 6 (10.7) 26 (15.7) Rifampin (RA) 5 <17 56 (100.0) 165 (99.4) Streptomycin (S) 10 <11 8 (14.3) 37 (22.3) Tetracycline (TE) 19 <14 12 (21.4) 31 (18.7) Trimethoprim/ Sulfamethoxazole (SXT) 23.75/1.25 <10 4 (7.1) 12 (7.2) Tobramycin (NN) 10 <12 40 (71.4) 12 (7.2) Trimethoprim (TMP) 5 <10 6 (10.7) 12 (7.2)
분리된대장균 (Escherichia coli) 의항생제내성 677. E. coli,, (Kim et al., 2008; Jang et al., 2017; Lee et al., 2017), (Ferreira et al., 2007; Peak et al., 2007; Reinthaler et al., 2010) (,,, )., 47%,,,,, 89.3% (NIFS, 2017). E. coli ( ) ansamycin rifampin (99.5%), cephem cephalothin (70.7%), cefepime (18.9%) ceftazidime (18.9%), aminoglyciside tobramycin (23.4%) streptomycin (20.2%) (Kim et al., 2016; Kim et al., 2017). E. coli tetracycline, E. coli quinolon nalidixic acid 90% nalidixic acid (16.7%) Table 4. Multiple antimicrobial resistance of E. col isolated from oyster Crassostrea gigas in Yongnam-Gwangdo Area No. of antimicrobials Resistance patterns No. of isolated strains Total (%) 1 RA 5 8.9 2 3 4 AM, RA 1 CF, RA 3 CTT, RA 1 RA, TE 1 TMP, RA 1 AMC, AN, RA 1 CF, AN, RA 1 CF, TMP, RA 1 FEP, NN, CAZ, RA 4 GM, S, TMP, RA 1 5 CF, FEP, NN, CAZ, RA 20 35.7 6 7 AM, CF, FEP, NN, CAZ, RA 3 CF, FEP, NN, CAZ, RA, C 1 CF, GM, FEP, NN, CAZ, RA 1 CF, FEP, NA, NN, CAZ, RA, TE 2 CF, S, FEP, NN, CAZ, RA, TE 2 8 CF, S, FEP, NN, CAZ, RA, C, TE 1 1.8 9 AM, CF, S, FEP, PIP, NN, CAZ, RA, TE 1 1.8 10 AM, CF, S, FEP, PIP, NN, CAZ, RA, C, TE 1 1.8 11 AM, CF, AN, FEP, NA, PIP, TMP, NN, CAZ, RA, SXT 1 AM, CF, FEP, NA, PIP, TMP, NN, CAZ, RA, TE, SXT 1 AM, CF, S, FEP, NA, PIP, NN, CAZ, RA, C, TE 1 12 AM, CF, S, FEP, NA, PIP, TMP, NN, CAZ, RA, TE, SXT 1 1.8 56 100 AN, Amikacin; AMC, Amoxicillin/Clavulanic acid; AM, Ampicillin; ATM, Aztreonam; MA, Cefamandole; CZ, Cefazolin; FEP, Cefepime; CTX, Cefotaxime; CTT, Cefotetan; FOX, Cefoxitin; CAZ, Ceftazidime; CF, Cephalothin; C, Chloramphenicol; CIP, Ciprofloxacin; GM, Gentamicin; IPM, Imipenem; NA, Nalidixic acid; PIP, Piperacillin; RA, Rifampin; S, Streptomycin; TE, Tetracycline; NN, Tobramycin; TMP, Trimethoprim; SXT, Trimethoprim/Sulfamethoxazole. 12.5 5.4 8.9 8.9 7.1 5.4
678 권순재ㆍ정연중ㆍ윤현진ㆍ목종수ㆍ권지영 Table 5. Multiple antimicrobial resistance of E. col isolated from inland pollution source in Yongnam-Gwangdo Area No. of antimicrobials Resistance patterns No. of isolated strains 0 1 0.6 1 RA 25 15.1 2 3 4 5 6 AN, RA 3 CF, RA 50 MM, RA 1 NA, RA 5 RA, C 2 RA, FOX 2 RA, TE 2 AN, NA, RA 1 CF, MA, RA 8 CF, NN, RA 1 CF, RA, TE 7 CF, S, RA 12 S, AN, RA 2 AM, CF, PIP, RA 2 AM, CF, S, RA 1 CF, GM, RA, FOX 1 CF, NA, RA, CIP 2 CF, NA, RA, TE 1 CF, PIP, RA, TE 1 CF, RA, C, TE 1 CF, S, NN, RA 1 CF, S, RA, TE 3 CF, RA, TE, CIP 1 GM, S, RA, TE 1 S, AN, NA, RA 1 AN, CF, GM, PIP, RA 1 AM, CF, NA, PIP, RA 1 AM, CF, PIP, RA, C 1 CF, S, AN, AMC, RA 1 CF, S, MA, RA, TE 1 AM, CF, NA, PIP, RA, TE 1 AM, CF, S, PIP, RA, FOX 1 CF, S, AN, PIP, RA, TE 1 CF, S, MA, RA, C, TE 1 CZ, CF, S, AM, RA, FOX 1 AN, Amikacin; AMC, Amoxicillin/Clavulanic acid; AM, Ampicillin; ATM, Aztreonam; MA, Cefamandole; CZ, Cefazolin; FEP, Cefepime; CTX, Cefotaxime; CTT, Cefotetan; FOX, Cefoxitin; CAZ, Ceftazidime; CF, Cephalothin; C, Chloramphenicol; CIP, Ciprofloxacin; GM, Gentamicin; IPM, Imipenem; NA, Nalidixic acid; PIP, Piperacillin; RA, Rifampin; S, Streptomycin; TE, Tetracycline; NN, Tobramycin; TMP, Trimethoprim; SXT, Trimethoprim/Sulfamethoxazole. Total (%) 39.2 18.7 9.6 3.0 3.0
분리된대장균 (Escherichia coli) 의항생제내성 679 Table 5. Continued No. of antimicrobials 7 8 Resistance patterns No. of isolated strains AM, GM, S, NA, RA, TE, CIP 1 AM, S, PIP, TMP, RA, TE, SXT 1 GM, PIP, TMP, MA, RA, TE, CIP 1 AM, CZ, CF, PIP, MA, RA, TE, CTX 1 AM, CZ, CF, PIP, TMP, RA, TE, SXT 1 9 AM, CZ, CF, NA, PIP, ATM, MA, RA, CTX 1 0.6 10 11 13 14 AM, CF, NA, PIP, TMP, NN, RA, C, TE, SXT 1 S, PIP, ATM, NN, MA, RA, TE, CTX, CIP, SXT 1 AM, CZ, CF, GM, NA, PIP, ATM, MA, RA, CTX, CIP 1 AM, CZ, CF, S, AMC, NA, PIP, TMP, MA, RA, SXT 3 CF, GM, S, NA, PIP, TMP, NN, MA, RA, C, STX 1 AM, CF, GM, S, NA, PIP, TMP, NN, RA, C, TE, CIP, SXT 1 AM, CZ, CF, GM, S, NA, PIP, NN, MA, RA, TE, CTX, CIP 1 AM, CZ, CF, GM, FEP, NA, PIP, TMP, ATM, NN, MA, RA, CTX, SXT 1 AM, CZ, CF, GM, S, PIP, TMP, NN, MA, CAZ, RA, TE, FOX, SXT 1 17 AM, CZ, CF, GM, FEP, NA, PIP, TMP, ATM, NN, MA, CAZ, RA, TE, CTX, CIP, SXT 1 0.6 Total 166 100 AN, Amikacin; AMC, Amoxicillin/Clavulanic acid; AM, Ampicillin; ATM, Aztreonam; MA, Cefamandole; CZ, Cefazolin; FEP, Cefepime; CTX, Cefotaxime; CTT, Cefotetan; FOX, Cefoxitin; CAZ, Ceftazidime; CF, Cephalothin; C, Chloramphenicol; CIP, Ciprofloxacin; GM, Gentamicin; IPM, Imipenem; NA, Nalidixic acid; PIP, Piperacillin; RA, Rifampin; S, Streptomycin; TE, Tetracycline; NN, Tobramycin; TMP, Trimethoprim; SXT, Trimethoprim/Sulfamethoxazole. Total (%) 1.8 1.2 1.2 3.0 1.2 1.2, E. coli. 대장균의항생제내성비교 E. coli Table 3. E. coli 24 18, rifampin (100%), cephalothin (73.2%), cefepime (71.4%), ceftazidime (71.4%), tobramycin (71.4%), 20%. 22 rifampin (99.4%), cephalothin (69.9%) 20%., (Grimes, 1991; Feldhusen, 2000). aztreonam (5%), cefamandole (17%), cefazolin (5%), cefotaxime (7%), cefoxitin (6%) E. coli, 대장균의다제내성양상 E. coli 56 Table 4. 24 1 12,. 56 4 (multiple antimicrobial resistance bacteria, MARB) 41 (73.2%), 5 (rifampin, cephalothin, cefepime, ceftazidime, tobramycin) (35.7%). E. coli E. coli (14.2%, 15.9%) (Park et al., 2013; Jo et al., 2016), (62.5%) (Son et al., 2009).,
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