• 제목/요약/키워드: spot on the lawn

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Screening of Bacteriocin-producing Enterococcus faecalis Strains for Antagonistic Activities against Clostridium perfringens

  • Han, Sun-Kyung;Shin, Myeong-Su;Park, Ho-Eun;Kim, So-Young;Lee, Wan-Kyu
    • 한국축산식품학회지
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    • 제34권5호
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    • pp.614-621
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    • 2014
  • This study was conducted to isolate and characterize bacteriocin-producing bacteria against Clostridium perfringens (C. perfringens) from domestic animals to determine their usefulness as probiotics. Bacteriocin-producing bacteria were isolated from pig feces by the spot-on-lawn method. A total of 1,370 bacterial stains were isolated, and six were tentatively selected after identifying the inhibitory activity against the pathogenic indicator C. perfringens KCTC 3269 and KCTC 5100. The selected strains were identified as Enterococcus faecalis (E. faecalis) by 16s rRNA sequencing. Most of the isolated bacterial strains were resistant to 0.5% bile salts for 48 h and remained viable after 2 h at pH 3.0. Some E. faecalis also showed strong inhibitory activity against Listeria monocytogenes KCTC 3569, KCTC 3586 and KCTC 3710. In the present study, we finally selected E. faecalis AP 216 and AP 45 strain based on probiotic selection criteria such as antimicrobial activity against C. perfringens and tolerance to acid and bile salts. The bacteriocins of E. faecalis AP 216 and AP 45 strains were highly thermostable, showing anticlostridial activities even after incubation at $121^{\circ}C$ for 15 min. These bacteriocin-producing bacteria and/or bacteriocins could be used in feed manufacturing as probiotics as an alternative to antibiotics in the livestock industry.

The Antimicrobial Activity of the Crude Extracts from Cichorium intybus L. (Chicory) against Bacillus cereus in Various Dairy Foods

  • Jeong, Dana;Kim, Dong-Hyeon;Chon, Jung-Whan;Kim, Hyunsook;Kim, Hong-Seok;Song, Kwang-Young;Kang, Il-Byung;Kim, Young-Ji;Park, Jin-Hyeong;Chang, Ho-Seok;Seo, Kun-Ho
    • Journal of Dairy Science and Biotechnology
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    • 제34권4호
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    • pp.239-244
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    • 2016
  • The roots of Cichorium intybus L. (chicory) have many nutritional qualities and also have been used for health promotion as traditional medicine. The antimicrobial activity of Cichorium intybus L.'s (chicory) ethanol extract on Bacillus cereus ATCC 10876 were investigated using spot on lawn method in this study. Ethanol extract exhibited antimicrobial activities against Bacillus cereus ATCC 10876 with significance. Also, the results produced the larger zones of inhibition against Bacillus cereus ATCC 10876 tested, with increasing concentration of Cichorium intybus L.'s (chicory) ethanol extracts,. Hence, Cichorium intybus L. (chicory) could be served for keeping the safety of various dairy products as natural antimicrobial agents.

Inhibition of Clostridium perfringens using Bacteriophages and Bacteriocin Producing Strains

  • Heo, Sunhak;Kim, Min Gon;Kwon, Mirae;Lee, Hee Soo;Kim, Geun-Bae
    • 한국축산식품학회지
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    • 제38권1호
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    • pp.88-98
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    • 2018
  • In this study, we isolated and characterized a bacteriocin-producing strain and two bacteriophages (P4, A3), showing antimicrobial effects against Clostridium perfringens, from chicken and swine feces by the spot-on-the lawn antagonism method. The selected strain was identified as Streptococcus hyointestinalis by 16S rRNA gene sequencing. The bacteriocin from the isolated strain exhibited strong inhibitory activity against four strains of C. perfringens and all the tested strains of Listeria monocytogenes, and the bacteriocin were highly heat- and pH-stable even at pH 2, pH 10 and $121^{\circ}C$ for 15 min. We also evaluated the combined effects of the isolated bacteriocin and phages. Combining the phage treatments and bacteriocin resulted in a synergetic effect compared with the phage or the bacteriocin alone. In addition, during the probiotic test, the bacteriocin-producing S. hyointestinalis B19 strain reduced the population of C. perfringens significantly. Treatment with S. hyointestinalis B19 and a cocktail of lytic bacteriophages eradicated the C. perfringens KCTC $3269^T$, completely. Consequently, the isolated bacteriocin and bacteriophages represent candidates for effective biocontrol of C. perfringens, and bacteriocin-producing S. hyointestinalis B19 is a potential probiotic candidate for use in domestic animals.

Selection and Characterization of Bacteriocin-Producing Lactobacillus sp. AP 116 from the Intestine of Pig for Potential Probiotics

  • Shin, Myeong-Su;Choi, Hyun-Jong;Jeong, Kyeong-Hyeon;Lim, Jong-Cheol;Kim, Kyeong-Su;Lee, Wan-Kyu
    • 한국축산식품학회지
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    • 제32권1호
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    • pp.31-39
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    • 2012
  • The purpose of this study was to isolate bacteriocin-producing bacteria with antagonistic activities against pathogens from the intestines of pigs for probiotic use. Lactobacillus sp. AP 116 possessing antimicrobial property was selected from a total of 500 isolates. The AP 116 strain showed a relatively broad spectrum of inhibitory activity against Listeria monocytogenes, Clostridium perfringens, Pediococcus dextrinicus, and Enterococcus strains using the spot-on-lawn method. Bacteriocin activity remained unchanged after 15 min of heat treatment at $121^{\circ}C$ and exposure to organic solvents; however, it diminished after treatment with proteolytic enzymes. Maximum production of bacteriocin occurred at $34^{\circ}C$ when a pH of 6.0 was maintained throughout the culture during fermentation. According to a tricine SDS-PAGE analysis, the molecular weight of the bacteriocin was approximately 5 kDa. The isolate tolerated bile salts and low pH, and also induced nitric oxide (NO) in mouse peritoneal macrophages. Bacteriocin and bacteriocin-producing bacteria, such as Lactobacillus sp. AP 116, could be potential candidates for use as probiotics as an alternative to antibiotics in the pig industry.

In vitro Characterization of Bacteriocin Produced by Lactic Acid Bacteria Isolated from Nem Chua, a Traditional Vietnamese Fermented Pork

  • Pilasombut, Komkhae;Rumjuankiat, Kittaporn;Ngamyeesoon, Nualphan;Duy, Le Nguyen Doan
    • 한국축산식품학회지
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    • 제35권4호
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    • pp.473-478
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    • 2015
  • The aim of this study was to screen and In vitro characterize the properties of bacteriocin produced by lactic acid bacteria isolated from Vietnamese fermented pork (Nem chua). One hundred and fifty LAB were isolated from ten samples of Nem chua and screened for bacteriocin-producing lactic acid bacteria. Antimicrobial activity of bacteriocin was carried out by spot on lawn method against both gram positive and gram negative bacteria. One isolate, assigned as KL-1, produced bacteriocin and showed inhibitory activity against Lactobacillus sakei, Leuconostoc mesenteroides and Enterococcus faecalis. To characterize the bacteriocin-producing strain, optimum temperature, incubation period for maximum bacteriocin production and identification of bacteriocin-producing strain were determined. It was found that the optimum cultivation temperature of the strain to produce the maximum bacteriocin activity (12,800 AU/mL) was obtained at 30℃. Meanwhile, bacteriocin production at 6,400 AU/mL was found when culturing the strain at 37℃ and 42℃. The isolate KL-1 was identified as L. plantarum. Antimicrobial activity of cell-free supernatant was completely inhibited by proteolytic enzyme of trypsin, alpha-chymotrypsin and proteinase K. Bacteriocin activity was stable at high temperature up to 100℃ for 10 min and at 4℃ storage for 2 d. However, the longer heating at 100℃ and 4℃ storage, its activity was reduced.

Antimicrobial Activity of Kefir against Various Food Pathogens and Spoilage Bacteria

  • Kim, Dong-Hyeon;Jeong, Dana;Kim, Hyunsook;Kang, Il-Byeong;Chon, Jung-Whan;Song, Kwang-Young;Seo, Kun-Ho
    • 한국축산식품학회지
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    • 제36권6호
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    • pp.787-790
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    • 2016
  • Kefir is a unique fermented dairy product produced by a mixture of lactic acid bacteria, acetic acid bacteria, and yeast. Here, we compared the antimicrobial spectra of four types of kefirs (A, L, M, and S) fermented for 24, 36, 48, or 72 h against eight food-borne pathogens. Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Enterococcus faecalis, Escherichia coli, Salmonella Enteritidis, Pseudomonas aeruginosa, and Cronobacter sakazakii were used as test strains, and antibacterial activity was investigated by the spot on lawn method. The spectra, potencies, and onsets of activity varied according to the type of kefir and the fermentation time. The broadest and strongest antimicrobial spectrum was obtained after at least 36-48 h of fermentation for all kefirs, although the traditional fermentation method of kefir is for 18-24 h at $25^{\circ}C$. For kefir A, B. cereus, E. coli, S. Enteritidis, P. aeruginosa, and C. sakazakii were inhibited, while B. cereus, S. aureus, E. coli, S. Enteritidis, P. aeruginosa, and C. sakazakii were inhibited to different extents by kefirs L, M, and S. Remarkably, S. aureus, S. Enteritidis, and C. sakazakii were only inhibited by kefirs L, M, and S, and L. monocytogenes by kefir M after fermentation for specific times, suggesting that the antimicrobial activity is attributable not only to a low pH but also to antimicrobial substances secreted during the fermentation.

김치에서 박테리오신을 분비하는 Lactobacillus sakei균주의 분리 (Isolation of a Bacteriocin - Producing Lactobacillus sakei Strain from Kimchi)

  • 김한택;박재용;이강권;김정환
    • 한국식품영양과학회지
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    • 제33권3호
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    • pp.560-565
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    • 2004
  • 배추김치로부터 식품유해균인 Listeria monocytogenes를 저해하는 박테리오신을 생산하는 유산균, Lactobacillus sakei P3-1이 분리되었다. 형태학적, 생화학적 특성조사와 최종적으로 PCR로 증폭하여 얻은 16S rDNA 염기서열 결정을 통해서 L. sakei로 동정되었다. L. sakei P3-1이 분비하는 박테리오신은 여러 그람 양성 및 음성균들 중에서 단지 L. monocytogenes만을 저해하는 그래서 저해범위가 매우 좁은 박테리오신으로 확인되었다. 이온교환 크로마토그래피에 의해서 박테리오신은 부분 정제되었으며 박테리오신의 열처리 안정성을 조사한 결과 121$^{\circ}C$에서 15분간 그리고10$0^{\circ}C$에서 10분간 열처리 후에도 각각 12.5%와 50%의 역가가 잔존하여 상당한 열안정성을 지니고 있음을 알 수 있었다. MRS배지에서 배양중 배양온도가 박테리오신 역가에 미치는 영향을 조사한 결과 3$0^{\circ}C$에서 배양할 때 그리고 18시간 이상 배양에서 가장 높은 1,000 AU/mL 역가를 보였다. 한편 SDS-PAGE 및 activity staining에 의해 측정된 박테리오신의 분자량은 4,000이었다. L. monocytogenes 생육 억제능, 작은 분자량 및 높은 열안정성 등의 성질들을 종합적으로 고려할 때 L. sakei P3-1이 생산하는 박테리오신은 박테리오신들 중에서 class II-a에 속하는 것으로 추정된다.

Antibacterial Activity of Crude Aronia melanocarpa (Black Chokeberry) Extracts against Bacillus cereus, Staphylococcus aureus, Cronobacter sakazakii, and Salmonella Enteritidis in Various Dairy Foods: Preliminary Study

  • Kim, Dong-Hyeon;Lim, Hyun-Woo;Kim, Se-Hyung;Lee, Jun-Man;Chon, Jung-Whan;Song, Kwang-Young;Bae, Dongryeoul;Kim, Jinhyun;Kim, Hyunsook;Seo, Kun-Ho
    • Journal of Dairy Science and Biotechnology
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    • 제36권3호
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    • pp.155-163
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    • 2018
  • In this study, the antibacterial activity of Aronia melanocarpa (black chokeberry) ethanol extract against Bacillus cereus, Staphylococcus aureus, Cronobacter sakazakii, and Salmonella Enteritidis was investigated using the spot-on-lawn assay. The results showed that this extract exhibited antibacterial activities against Bacillus cereus (complete inhibition) and Staphylococcus aureus (partial inhibition), but did not inhibit the growth of Cronobacter sakazakii and Salmonella Enteritidis. This study shows that the Aronia melanocarpa (black chokeberry) ethanol extract was more effective against Gram-positive bacteria than Gram-negative bacteria. Hence, it is suggested that Aronia melanocarpa could be a useful food supplement, and could be utilized as a naturally derived additive for maintaining the safety of various dairy products. Furthermore, future research should be conducted to examine the possibility of using such products as functional ingredients for improving the anti-oxidative and anti-inflammatory activities of food products.

Antibacterial activities of bark extracts from Fraxinus rhynchophylla Hance and Geranium koreanum Kom. against clinical strains of Clostridium perfringens in chickens

  • Oh, Jae-Young;Lee, Kwang-Jae;Wei, Bai;Roh, Jae-Hee;Kang, Min;Cha, Se-Yeoun;Jang, Hyung-Kwan
    • 대한수의학회지
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    • 제55권2호
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    • pp.117-123
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    • 2015
  • Necrotic enteritis (NE) caused by Clostridium (C.) perfringens commonly occurs in domestic broiler farms since antibiotic supplementation in poultry feed has been banned. We evaluated the antibacterial activities of medicinal plant extracts against C. perfringens isolates to select alternative compounds for preventing NE. We compared antibacterial activities using two methods and evaluated susceptibilities of the isolates based on minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Two (Fraxinus rhynchophylla Hance [FRH] and Geranium koreanum Kom. [GKK]) of the 30 plant extracts had potent antibacterial activities against C. perfringens ATCC 13124 in two assays. The MIC values for FRH and GKK against 20 C. perfringens isolates were $128{\sim}256{\mu}g/mL$ and $32{\sim}128{\mu}g/mL$, respectively. The geometric MIC mean values for the two extracts were $147.2{\mu}g/mL$ and $68.8{\mu}g/mL$, respectively. The MBCs for the two extracts against the same strains were $1,024{\sim}2,048{\mu}g/mL$ and $256{\sim}1,024{\mu}g/mL$, respectively. The geometric mean MIC and MBC for GKK were about two-fold lower than those of FRH. The modified spot-on-lawn assay may be useful for measuring primary antibacterial potential. FRH and GKK are expected to be used as feed additives to prevent or treat NE in veterinary practice.

돼지 장관으로부터 분리한 Lactobacillus sp. HN 235 균주가 생산하는 항균물질의 특성 (Characterization of Antimicrobial Substance Produced by Lactobacillus sp. HN 235 Isolated from Pig Intestine)

  • 신명수;한선경;최지현;지애란;김경수;이완규
    • 한국미생물·생명공학회지
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    • 제37권2호
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    • pp.125-132
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    • 2009
  • 항생제 대체물질로서 양돈용 생균제를 개발하기 위하여, 돼지의 소장 및 대장으로부터 500균주의 미생물을 분리하였으며, 항균물질인 박테리오신을 생산하는 5균주를 최종적으로 선발하였다. 선발균주들은 모두 Lactobacillus 속으로 동정되었으며, 생균제를 개발할 목적으로 상기 균주 중 Lactobacillus sp. HN 235 균주에 대한 균주 및 박테리오신 특성을 조사하였다. HN 235 균주의 성장시간에 따른 생균수 및 항균활성을 측정한 결과, 18시간째 최대 $3.5{\times}10^9\;cfu/mL$까지 성장하였으며, 항균활성은 유도기에 나타나기 시작하여 정체기에 6,400 AU/mL로 최고의 활성을 보이다가 급격하게 활성을 상실하였다. HN 235 균주가 생산하는 박테리오신은 병원성 균주로 알려진 L. monocytogenes와 Cl. perfringens에 대하여 강한 항균능력을 나타냈으며, 상기 박테리오신의 항균활성은 단백질 분해효소 처리시 활성을 상실하였다. 또한 $95^{\circ}C$에서도 활성을 유지하였으며, pH 2~10 범위 및 유기용매에 대해서도 비교적 안정한 항균활성을 나타내었다. SDS-PAGE를 실시하여 분자량을 측정한 결과, 대략 5 kDa으로 확인되었다.