돼지 폐렴병소에서 분리한 Actinobacillus pleuropneumoniae의 특성에 관한 연구

정병열,조길재,김봉환,조광현,Jung, Byeong-yeal,Cho, Gil-jae,Kim, Bong-hwan,Cho, Kwang-hyun 대한수의학회 1996 大韓獸醫學會誌 Vol.36 No.1

The present study was conducted to investigate the biochemical and serologic characteristic of Actinobacillus pleuropneumoniae isolated from pneumonic lungs of pigs during the period from January 1992 to April 1993. A pleuropneumoniae was isolated from 17(27.0%) of 63 growing pigs with respiratory signs and 21(6.4%) of 330 pneumonic lungs of slaughtered pigs. The seasonal isolation frequency of A pleuropneumoniae was higher in winter and spring than that in summer or fall. The biochemical and cultural properties of A pleluropneumoniae isolated from the pneumonic lungs of pigs were identical to those of the reference strains used. The isolates were highly susceptible to ampicillin, cephalothin, ceftiofur, ciprofloxacin(MIC : ${\leq}0.39{\mu}g/ml$) and moderately susceptible to amikacin, chloramphenicol, erythromycin, kanamycin, methicillin, penicillin-G, streptomycin(MIC : 0.78~25IU or ${\mu}g/ml$), respectively. Sulfadimethoxine, sulfamerazine, tylosine showed no response to the isolates(MIC : ${\geq}100{\mu}g/ml$). Among the 38 isolates, 21(55.3%) and 13(34.2%) were resistant to oxytetracycline aid lincomydn, respectively(MIC : ${\geq}50IU$ or ${\mu}g/ml$). The majority of 38 A pleuropneumoniae isolates were turned out as serotype 2(47.4%) or serotype 5(54.7%) and the remaining 3 isolates were evenly classified to serotype 7, 10 or 12. It was noted A pleuropneumonine serotype 5 isolates were more resistant to oxytetracycline than serotype 2 isolates.

Listeria monocytogenes의 신속검출을 위한 선택배지 및 multiplex PCR 기법 개발

정병열,임현숙,정석찬,Jung, Byeong-yeal,Lim, Hyun-sook,Jung, Suk-chan 대한수의학회 2003 大韓獸醫學會誌 Vol.43 No.2

Listeria (L.) monocytogenes in samples could not be detected occasioally by faster growth of other Listeria spp. especially L. innocua. The aim of this study was to develop the differential media and multiplex polymerase chain reaction (PCR) assays for the rapid detection of L. monocytogenes. L. monocytogenes colonies were characterized by their ${\beta}$-hemolysis with fluorescence under 366 nm UV light on the Listeria hemolysis agar (LHA). L. innocua, a species commonly present in foods, did not produce ${\beta}$-hemolysis on LHA. Therefore, one or more colonies of L. monocytogenes were easily distinguished from large populations of L. innocua. The multiplex PCR assays were developed to distinguish from L. monocytogenes and other Listeria spp. with two pairs of primers. The primers were designed in 16S rRNA and listeriolysin O gene for specific amplification of all members of the genus Listeria and L. monocytogenes, respectively. The multiplex PCR assays produced 560 and 938 bp products in L. monocytogenes; only 938 bp products in the genus Listeria. The multiplex PCR assays could detect as little as 50 pg of L monocytogenes DNA. These results indicated that the differential media and multiplex PCR assays might be useful diagnostic tools for the rapid detection of L. monocytogenes.

국내 돼지의 Streptococcus suis 감염율 조사 및 혈청형 동정

정병열,정석찬,김봉환,박용호,박정문,Jung, Byeong-yeal,Jung, Suk-chan,Kim, Bong-hwan,Park, Yong-ho,Park, Jeung-moon 대한수의학회 1997 大韓獸醫學會誌 Vol.37 No.3

To investigate the prevalence of Streptococcus suis in pig industry in Korea, we isolated S suis from pigs during two years. The isolates were identified by biochemical and coagglutination test. Serotypes and antimicrobial susceptibility of the isolates were determined. Fourteen strains(12.2%), 11 strains(27.5%), 58 strains(7.8%) and 4 strains(11.1%) of S suis were isolated from 115 nasal swabs, 40 tonsils of healthy pigs, 745 pneumonic lungs and 36 meningitis of diseased pigs, respectively. The isolates were highly susceptible to ampicillin, cephlothin and ofloxacin while they were resistant to oxytetracycline. Among the isolates, 63(75.9%) strains were to the S suis capsular type 1 to 10 and 11(13.3%) strains were untypable. Capsular type 2 was the most prevalent with 32(38.6%) strains of all isolates.

식육중 Escherichia coli O157 검출을 위한 enzyme immunoassay 기법 개발

정병열,정석찬,조동희,김종염,박용호,신쌍재,김성국,김봉환,Jung, Byeong-yeal,Jung, Suk-chan,Cho, Dong-hee,Kim, Jong-yeom,Park, Yong-ho,Shin, Sang-jae,Kim, Sung-guk,Kim, Bong-hwan 대한수의학회 1998 大韓獸醫學會誌 Vol.38 No.4

A sensitive and rapid enzyme immunoassay(EIA) to detect Escherichia coli O157 in ground beef was developed by using a sandwich type assay with polyclonal antibodies to E coli O157. E coli O157 in ground beef could be detected within 15hr, including incubation for 12hr in enrichment broth and 3hr in immunoassay. The EIA could detect $1.3{\times}10^5$ cells of E coli O157/g of ground beef without enrichment. The lowest limit of detection was 0.23 E coli O157 per g of meat after enrichment. Confirmation was required in the positive specimens in the EIA by culture method even though the negative specimens were not. These results suggested that the immunoassay could be a very efficient method for the screening E coli O157 in meat.

Streptococcus suis 신속동정을 위한 PCR 기법

정병열,정석찬,김종염,박용호,김봉환,Jung, Byeong-yeal,Jung, Suk-chan,Kim, Jong-yeom,Park, Yong-ho,Kim, Bong-hwan 대한수의학회 1998 大韓獸醫學會誌 Vol.38 No.4

Synthetic oligonucleotide primers of 20 and 21 bases, respectively, were used in the polymerase chain reaction (PCR) to amplify a sequence of the mrp gene, which encodes the muramidase released protein of Streptococcus suis. Amplification was not recorded when 5 other streptococcal species were tested or when 9 different nonstreptococcal species were tested. A DNA fragment of 517bp was amplified from the genomic DNA of S suis. The lower detection limit was 100pg of the genomic DNA. The primers recognized 34 serotypes of S suis reference strains and 9 isolates from pneumonic lung, brain, nasal discharge, tonsil. This results suggest that the amplification of the mrp gene by PCR method is potential for the identification of S suis isolates.

Characteristics of verotoxin non-producing Escherichia coli O157 and verotoxin-producing E coli isolated from healthy cattle

정병열,정석찬,박홍제,조길재,김봉환,Jung, Byeong-yeal,Jung, Suk-chan,Park, Hong-je,Cho, Gil-jae,Kim, Bong-hwan The Korean Society of Veterinary Science 2000 大韓獸醫學會誌 Vol.40 No.3

Verotoxin을 산생하지 않은 Escherichia coli O157과 verotoxin을 산생하는 E coli(VTEC)를 건강한 소의 분변에서 분리하여 생화학적 및 유전적인 특성에 대해서 비교하였다. E coli O157 : nonH7(운동성은 있으나 H혈청형이 7이 아님)의 sorbitol 분해능과 ${\beta}-glucurondase$ 활성은 E coli O157 : H7이 나타내는 것과는 차이가 있었다. 그리고 uidA 유전자는 verotoxin 산생능과 상관없이 sorbitol과 ${\beta}-glucuronidase$ 음성인 E coli O157 : H7에서 특이적으로 검출되었다. 한편 6개 목장에서 수거한 소 분변 45예에서 VTEC를 분리한 결과, 7주(15.6%)가 분리되었으며 이들은 모두 sorbitol을 분해하였으며 ${\beta}-glucurondase$ 활성이 있었으나 장벽 부착인자를 지배하는 eaeA 유전자가 없었다. 비록 소가 VTEC의 보균원으로 추정되나, 정상 소에서 분리한 VTEC는 eaeA 유전자가 결여된 균주가 많으므로 공중위생학상 eaeA 유전자를 보유한 E coli 보다 위해성이 낮으며, 이러한 결과는 왜 사람에서 유행하는 VTEC 혈청형과 소에서 유행하는 것과 차이가 있는지를 일부 설명해준다. Verotoxin non-producing E coli O157 strains have been isolated from cattle feces and compared in particular regard to biochemical properties and genotypes with verotoxin-producing E coli (VTEC). E coli O157 : nonH7 strains had different phenotypes in sorbitol fermentation and ${\beta}-glucuronidase$ activity from E coli O157 : H7. Regardless of verotoxin production ability of E coli O157 : H7, uidA gene was uniquely detected from sorbitol and ${\beta}-glucuronidase$ negative E coli O157 : H7. Forty five fecal samples from 6 dairy farms were obtained and VTEC was detected as 15.6% (7 strains) of the samples. Most VTEC isolates were positive for sorbitol fermentation and ${\beta}-glucuronidase$ activity but negative for eaeA gene. This study suggested that cattle could be a reservior for VTEC. However, absence of eaeA gene in VTEC isolates from most of healthy cattle suggested that they might be less virulent than eaeA-positive E coli against human health.

면역크로마토그라피 기법을 이용한 Salmonella 속균 신속 검출킷트 개발

정병열,정석찬,Jung, Byeong-yeal,Jung, Suk-chan 대한수의학회 2005 大韓獸醫學會誌 Vol.45 No.2

An immunochromatographic (IC) strip for the rapid detection of Salmonella spp. in the enriched sample was developed. Affinity purified Salmonella polyclonal antibody was conjugated with 40 nm colloidal gold particles which were prepared by citrate method in our laboratory. The antigen-antibody-gold complex was captured by Salmonella antibody attached to test line of nitrocellulose membrane during the capillary migration of sample. Specificity of the IC strip was calculated to be 100% (12/12) and sensitivity was 97.6% (41/42) in the test with pure cultured bacteria. Salmonella was artificially inoculated into raw pork macerated with enrichment broth. And then it was 10-fold diluted from $5.2{\times}10^{8}CFU/ml$ to 5.2 CFU/ml. The IC strip could detect $5.2{\times}10^{6}CFU/ml$ before enrichment. However, the lowest limit of detection was 5.2 CFU/ml after overnight incubation. The results indicated that the IC assay was a rapid, economical and simple method with high specificity and sensitivity for the detection of Salmonella spp. without using any equipment.

Listeria 속균 신속 검출을 위한 면역크로마토그라피 킷트 개발

정병열,정석찬,김종만,Jung, Byeong-yeal,Jung, Suk-chan,Kim, Jong-man 대한수의학회 2005 大韓獸醫學會誌 Vol.45 No.2

We developed the one-step strip based on an immunochromatographic (IC) assay for the rapid detection of Listeria spp. Genus-specific monoclonal antibody to flagella of L. monocytogenes was conjugated with 40 nm colloidal gold particles which were prepared in our laboratory. The specificity of the IC strip was tested with pure cultured bacteria. All strains of the genus of Listeria spp. yielded positive reactions and 12 strains of non-Listeria were negative, resulting in a specificity of 100%. L. monocytogenes was artificially inoculated in raw pork macerated with listeria enrichment broth. And then it was 10-fold diluted from $8.7{\times}10^6$ to 8.7 CFU/ml. L. monocytogenes could be detected at a minimum of $8.7{\times}10^5CFU/ml$ before enrichment, $8.7{\times}10^2CFU/ml$ after primary enrichment and 8.7 CFU/ml after secondary enrichment, respectively. These results indicated that the IC strip exhibited high specificity and sensitivity in the detection of Listeria spp.

PCR을 이용한 소 세균성 호흡기질병 원인체 신속동정

정병열,Jung, Byeong-yeal 대한수의학회 2004 大韓獸醫學會誌 Vol.44 No.3

Haemophilus somnus, Mycoplasma bovis and Pasteurella multocida were responsible for respiratory diseases in bovine. Methods for identifying these bacteria had poor sensitivity and specificity. In this paper, PCR assays were applied for rapid identification of H. somnus, M. bovis, P. multocida B:2 and P. multocida capsular types. The specific PCR products were amplified from H. somnus, but not from other bacteria. Ten-fold diluted H. somnus were mixed with P. multocida and then the mixed cultures were inoculated on agar plates. After incubation, PCR was performed with harvest from agar plates and could detect as few as 3.4 CFU/ml of H. somnus. The primers MboF and MboR produced an amplification product unique to M. bovis and sensitivity of PCR was as low as 100 pg of DNA. Only serotype B:2 of P. multocida, the causal agent of haemorrhagic septicemia in bovine, was specifically amplified in PCR among the 16 reference serotypes. The multiplex capsular PCR typing for P. multocida was produced the P. multocida specific product as well as the capsular serogroup-specific product. The present PCR assays should be useful for the rapid identification of bacterial pathogens from bovine respiratory diseases.

돈군의 Salmonella 모니터링을 위한 림프절 추출액 사용에 대한 평가

정병열,추지훈,김지훈,정재윤,Jung, Byeong-Yeal,Choo, Ji-Hoon,Kim, Ji-Hun,Jung, Jae-Yun 대한수의학회 2006 大韓獸醫學會誌 Vol.46 No.2

The objective of this study was to investigate the use of extract from mesenteric lymph nodes as an alternative to serum for ELISA to detect Salmonella antibodies in slaughter pigs. Among 324 slaughter pigs, 65 (20.1 %) were positive in the serum ELISA and 76 (23.5%) were positive in the ELISA with extract from lymph nodes. A total of 24 (7.4%) Salmonella representing 6 serotypes were isolated from mesenteric lymph nodes and 35 (10.8%) Salmonella belonging to 2 serotypes were also recovered from cecal contents of slaughter pig samples, respectively. The most prevalent serogroup was B (55.9% of isolates) and serotype was Typhimurium (52.5% of isolates). In the comparison of the results of between the serum ELISA and Salmonella isolation, kappa value was 0.28 with mesenteric lymph nodes and 0.37 with cecal contents, respectively. However, the extract ELISA had sensitivity of 98.5%, specificity of 95.4% and kappa value of 0.88 as compared with the serum ELISA. Because high degree of concordance between the serum ELISA and the extract ELISA was observed (P=0.24), extract from lymph nodes could be used as an alternative to serum for the detection of Salmonella antibodies in the ELISA.