• 한국미생물·생명공학회지
• /
• 제26권2호
• /
• pp.117-121
• /
• 1998

김치로부터 분리한 Lactobacillus plantarum bacteriophage SC 921은 M.O.I가 0.2일 경우 용균효과가 빠르게 진행됨을 알 수 있었고, SDS-PAGE를 실시하여 phage particle protein을 조사해 본 결과 4개의 major protein으로 구성되어 있는데 이들은 각각 48, 34, 32, 29 kDa으로 구성되어 있다. Exo III로 30분간 반응시킨 후 S1 nuclease를 처리하여 DNA의 형태를 조사해 본 결과 intact DNA는 linear form의 double strand를 유전전달 물질로 가지고 있었다. 제한효소에 대한 절단 효과를 조사한 결과, Sma I에 대해서 1개, Xba I, Cla I, Kpn I, EcoRI에 대해서 각각 2, 4, 5, 6개의 절단부위를 가지고 있으며, Hind III에 대해서는 절단부위가 매우 많은 것을 알 수 있었다. Hind III를 이용하여 intact DNA의 genome size를 측정해본 결과 약 66.5 kbp정도였다. 위의 실험결과와 restriction enzyme mapping을 통해 기존에 알려진 bacteriophage B2와 비교해본 결과 숙주 균주는 같으나 단백질적인 구조나 유전전달물질로 본 구조는 서로 다름을 알 수 있었다.

• 대한미생물학회지
• /
• 제20권1호
• /
• pp.1-11
• /
• 1985

A total of 211 strains of Staphylococcus aureus which included 118 strains isolated from various clinical specimens of admitted patients of University Hospital with systemic or severe cases of infection and 93 strains from infected skin diseases of out-patients of dermatology clinic located in rural area, were tested for the antimicrobial susceptibility to the 12 drugs of common use and the phage typing. An these were subjected to the study of plasmid profile analysis for the molecular epidemiology of nosocomial infections. University Hospital(UH) isolates showed higher frequency of resistance than local clinic(LC) isolates against 10 drugs excluding tetracycline(Tc), and trimethoprim(Tp). The MIC of UH isolates were above than $128{\mu}g/ml$ against 9 drugs except Tc, gentamicin(Gm), and Tp, but LC isolates did not show such a high level of MIC. There was difference of MIC needed to inhibit 90% of strains(MIC90) against each drugs tested between two groups of UH and LC isolates. UH isolates showed 2 to 4 times higher value of MIC90 by two-fold serial dilution of drug concentration than LC isolates. Tp was considered as an effective drug in treatment of staphylococcal infections whereas ampicillin and Gm were appeared to be ineffective. Seventy-three strains(61.9%) of UH isolates and 70(69.9%) of LC were typable with phages from Colindale Reference Laboratory. The prevailing phage type of UH isolates belonged to lytic group II were 27 strains(22.9%) and those of LC isolates belonged to lytic group II were 23 strains(24.7%). Thirteen strains(11.0%) of UH isolates were multiply resistant to more than 5 drugs to 10 drugs but none of LC isolates. Through the lysis method of Kado and Liu followed by agarose gel electrophoresis, none of 211 strains showed plasmid profile. These results were confirmed by re-examination through the method of Birnboim and Doly.

• Journal of Microbiology and Biotechnology
• /
• 제24권5호
• /
• pp.592-596
• /
• 2014

HMO-2011, a recently isolated lytic phage that infects the SAR116 bacterial clade, represents one of the most abundant phage types in the oceans. In this study, the HMO-2011 genome sequence was compared with virome sequences obtained from various depths of the Pacific Ocean regions using metagenome binning. HMO-2011 was confirmed to be one of the most highly assigned viruses, with a maximum of 7.6% of total reads assigned. The HMO-2011-type phages demonstrated a depth-specific distribution, showing more abundance in the euphotic zone of coastal, transition, and open ocean regions as compared with the dark ocean.

• 농업과학연구
• /
• 제14권2호
• /
• pp.286-294
• /
• 1987

오골계 난백 lysozyme의 용균특성을 조사하고, 이를 식품보존료로 이용하기 위한 자료를 얻고자 오골계 난백으로부터 추출 분리한 lysozyme의 각종 미생물에 대한 용균성을 조사하였다. 그 결과, 공시균중 Gram 양성균인 Staphylococcus aureus phage type 29와 Bacillus subtilis ATCC 6633에 대하여는 용균성이 인정되었으나, Gram 음성균인 E. coli를 비롯한 여타의 균종과 Staphylococcus aureus phage type 57은 lysozyme에 대한 용균 감수성이 인정되지 않았다. 분리된 lysozyme은 S. aureus phage type 29를 공시균주로 하여 이를 육즙배지에 접종하고, $37^{\circ}C$에서 24시간 정치배양하여 세포를 회수한 후 540nm에서 흡광도가 0.6이 되도록 0.05M 초산완충액(pH 4.5)에 현탁시켜, 여기에 0.05%의 lysozyme을 가하여 $30^{\circ}C$, 30분간의 조건으로 반응시킬 때 용균성이 가장 좋았다. 또한 0.05%의 lysozyme은 반응액 중에 glycine(1%)을 첨가하므로서 공시균에 대한 용균효과가 양자의 상승작용에 의하여 그 단용시보다 현저히 증대(<50%) 되었으나, 기타 다른 첨가제와 금속이온 및 lysozyme과의 혼용효과는 인정되지 않았다.

• KSBB Journal
• /
• 제10권5호
• /
• pp.533-539
• /
• 1995

재조합 대장균에 의해 생합성된 PHB를 분리정제하기 위하여 박테리오파아지의 세포파괴작용을 이용하는 방법의 가능성에 대해 알아 보았다. 먼저, $cI_{857}$ 유전자를 지난 박테리오파아지 A를 대장균에 감염시킨 후 lysogen, XLl-Blue($\lambda$HL1)를 선별하고, 이 균주에 PHA 생합성 플라스미드를 도입시켜 원하는 균주인 XLI-Blue($\lambda$HL1, pSYLl05)를 만들였다. 숙주인 XLl-Blue, 열적유도에 의해 세포파괴가 가능한 XLl-Blue($\lambda$HL1), 그리고 세포파괴와 PHB 생합성이 모두 가능한 XLl-Blue(AHL1, pSYL105) 에 대하여 여러 가지 조건에서의 실험결과를 비교.검토하였다. XLI-Blue($\lambda$IL1, pSYLl05)의 경우 대수기에서는 열적유도만으로 세포파괴를 효과적으로 야기할 수 있었으나 PHB가 축적되는 정지기에서는 열척유도만으로 세포파괴를 일으킬 수 없었다. 세포 파괴를 보다 용이하게 하기 위하여 열적유도 빛 2% (v/v)의 chloroform을 사용하는 화학적용균을 병행 하였는데, 이 경우 세포파괴가 효과적으로 일어남을 관찰할 수 있었다.

• Journal of Microbiology and Biotechnology
• /
• 제24권6호
• /
• pp.803-811
• /
• 2014

Concerns over drug-resistant bacteria have stimulated interest in developing alternative methods to control bacterial infections. Endolysin, a phage-encoded enzyme that breaks down bacterial peptidoglycan at the terminal stage of the phage reproduction cycle, is reported to be effective for the control of bacterial pathogenic bacteria. Bioinformatic analysis of the SPN9CC bacteriophage genome revealed a gene that encodes an endolysin with a domain structure similar to those of the endolysins produced by the P1 and P22 coliphages. The SPN9CC endolysin was purified with a C-terminal oligo-histidine tag. The endolysin was relatively stable and active over a broad temperature range (from $24^{\circ}C$ to $65^{\circ}C$). It showed maximal activity at $50^{\circ}C$, and its optimum pH range was from pH 7.5 to 8.5. The SPN9CC endolysin showed antimicrobial activity against only gram-negative bacteria and functioned by cutting the glycosidic bond of peptidoglycan. Interestingly, the SPN9CC endolysin could lyse intact gram-negative bacteria in the absence of EDTA as an outer membrane permeabilizer. The exogenous lytic activity of the SPN9CC endolysin makes it a potential therapeutic agent against gram-negative bacteria.

• Journal of Microbiology and Biotechnology
• /
• 제34권6호
• /
• pp.1189-1196
• /
• 2024

Bacterial resistance to commonly used antibiotics is one of the major challenges to be solved today. Bacteriophage endolysins (Lysins) have become a hot research topic as a new class of antibacterial agents. They have promising applications in bacterial infection prevention and control in multiple fields, such as livestock and poultry farming, food safety, clinical medicine and pathogen detection. However, many phage endolysins display low bactericidal activities, short half-life and narrow lytic spectrums. Therefore, some methods have been used to improve the enzyme properties (bactericidal activity, lysis spectrum, stability and targeting the substrate, etc) of bacteriophage endolysins, including deletion or addition of domains, DNA mutagenesis, chimerization of domains, fusion to the membrane-penetrating peptides, fusion with domains targeting outer membrane transport systems, encapsulation, the usage of outer membrane permeabilizers. In this review, research progress on the strategies for improving their enzyme properties are systematically presented, with a view to provide references for the development of lysins with excellent performances.

• 한국축산식품학회지
• /
• 제37권2호
• /
• pp.320-328
• /
• 2017

In this study, two Listeria bacteriophages, LMP1 and LMP7, were isolated from chicken feces as a means of biocontrol of L. monocytogenes. Both bacteriophages had a lytic effect on L. monocytogenes ATCC 7644, 15313, 19114, and 19115. Phages LMP1 and LMP7 were able to inhibit the growth of L. monocytogenes ATCC 7644 and 19114 in tryptic soy broth at $10^{\circ}C$ and $30^{\circ}C$. Nevertheless, LMP1 was more effective than LMP7 at inhibiting L. monocytogenes ATCC 19114. On the contrary, LMP7 was more effective than LMP1 at inhibiting L. monocytogenes ATCC 7644. The morphology of LMP1 and LMP7 resembled that of members of the Siphoviridae family. The growth of L. monocytogenes ATCC 7644 was inhibited by both LMP1 and LMP7 in milk; however, the growth of L. monocytogenes ATCC 19114 was only inhibited by LMP1 at $30^{\circ}C$. The lytic activity of bacteriophages was also evaluated at $4^{\circ}C$ in milk in order to investigate the potential use of these phages in refrigerated products. In conclusion, these two bacteriophages exhibit different host specificities and characteristics, suggesting that they can be used as a component of a phage cocktail to control L. monocytogenes in the food industry.

• Journal of Microbiology and Biotechnology
• /
• 제29권12호
• /
• pp.1916-1924
• /
• 2019

Outbreaks of staphylococcal food poisoning (SFP) causing serious human diseases and economic losses have been reported globally. Furthermore, the spread of Staphylococcus aureus with increased resistance to multiple antimicrobial agents has become a major concern in the food industries and medicine. Here, we isolated an endolysin LysSAP8, as one of the peptidoglycan hydrolases, derived from the bacteriophage SAP8 infecting S. aureus. This endolysin was tagged with a 6×His at the C-terminal of the target protein and purified using affinity chromatography. LysSAP8 demonstrated lytic activity against a broad spectrum of bacteria, which included a majority of the staphylococcal strains tested in this study as well as the methicillin-resistant S. aureus (MRSA); however, no such activity was observed against other gram-positive or gram-negative bacteria. Additionally, LysSAP8 could maintain bactericidal activity until 0.1 nM working concentration and after heat treatment at 37℃ for 30 min. The ability of LysSAP8 to lyse cells under varying conditions of temperature (4-43℃), pH (3-9), and NaCl concentrations (0-1,000 mM), and divalent metal ions (Ca²⁺, Co²⁺, Cu²⁺, Mg²⁺, Mn²⁺, Hg²⁺, and Zn²⁺) was examined. At the optimized condition, LysSAP8 could disrupt approximately 3.46 log CFU/ml of the planktonic cells in their exponential phase of growth within 30 min. In this study, we have suggested that LysSAP8 could be a potent alternative as a biocontrol agent that can be used to combat MRSA.

• 한국축산식품학회지
• /
• 제38권1호
• /
• pp.88-98
• /
• 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.