Microbes in insect gut significantly influence host physiology. While Lepidoptera is a diverse insect order, the relationship between microbial symbiosis and host development remains elusive, especially concerning role of gut-colonizing bacteria in metamorphosis. We investigated the gut microbial diversity in Galleria mellonella throughout its life cycle using 16S rRNA amplicon sequencing. Our findings revealed a predominance of Enterococcus spp. in larvae and Enterobacter spp. in pupae. Remarkably, removing Enterococcus spp. hastened the larval-to-pupal transition. Transcriptome analysis showed an upregulation of immune response genes in pupae and hormone genes in larvae. Notably, the production of antimicrobial peptides in the host gut varied with developmental stages. Some of these peptides suppressed the growth of Enterococcus innesii, a dominant gut bacterium in G. mellonella larvae. This research underscores the pivotal role of gut microbiota shifts in metamorphosis, driven by the secretion of antimicrobial peptides in the G. mellonella digestive system.

We investigated the epidemiological characteristics of the antimicrobial resistant Enterococcus isolates from the four major rivers of Korea in 2012. A total 316 surface water samples were collected from three distinct sites (nearby livestock farms, tributaries, and major rivers) at two different seasons (dry season: n = 76, wet season: n = 240). A total 654 bacterial cells were isolated from samples and their genus distribution were determined. We found that Gram-negative bacteria including various genera were prevalent (n = 522, 79.8%), and Enterococcus was the most common genus of Gram-positive bacteria (n = 119, 18.2%). The isolation rate of Gram-negative bacteria was higher in wet season, whereas that of Enterococcus isolates was higher in dry season. The prevalence of Enterococcus isolates was also higher nearby livestock farms than on tributaries and main rivers. Since Enterococcus isolate is a key indicator for animal fecal contamination, the following experiments focused on this microorganism. As compared to a previous report in 2006, the resistance rates in E. faecium to erythromycin (40.0% to 69.9%) and chloramphenicol (0% to 16.4%) were increased, whereas those to penicillin (56.0% to 4.1%) and teicoplanin (36.0% to 0%) were decreased. We also found that antimicrobial-resistant (AMR) E. faecium isolates from rivers and livestock samples shared similar pulsed-filed gel electrophoresis (PFGE) profiles, validating the transmission of AMR Enterococcus isolates from livestock to river. Taken together, this study provides us with detailed information about bacterial contamination status in four major rivers, and highlights the changes in AMR pattern of Enterococcus isolates, which are expected to have originated from livestock.

본 연구에서는 국내산 돌미나리에서 GABA생성능이 있 는 신규 유산균을 분리, 동정한 결과, Enteroccoccus casseliflavus로 확인되었다. 최근까지 Lactobacillus속과 같 은 GABA 생성 유산균에 대한 보고는 많이 되고 있고, 일 부 Enterococcs속 유산균도 보고되고 있으나 E. casseliflavus 종에 대한 보고는 없었다. 따라서 본 연구에서는 E. casseliflavus PL05 균주에 대한 GABA 생성 최적 조건을 찾기 위하여 배지의 유형, 생육 온도, 초기 pH 조건, 배양 시간, MGS 농도 및 탄소원을 포함한 다양한 조건을 테스 트하였다. PL05 균주는 MRS 혹은 TSB 배지보다 BHI 배 지에서 생육이 잘 되었으며, 배지의 초기 pH는 7-9 조건 에서 가장 생육이 왕성하였고, GABA 생성 조건 역시 유 사한 결과로 확인되었다. GABA의 기질에 해당하는 MSG 의 농도별 GABA 생성량을 조사한 결과, 7%에서 가장 높 은 생성량을 나타내었으나 5%에서도 유사한 수준으로 확 인되어 효율적인 측면에서 5%가 적합할 것으로 판단된다. 탄소원에 따른 생육 및 GABA 생성량은 말토오스를 사용 하였을 때 가장 높은 것으로 확인되었고, 이러한 최적의 조건들로 최종 테스트를 진행한 결과, 24시간째 140.06±0.71 mM의 GABA가 생성되었고, 전환율은 78.95%로 확인되 었다. 또한 반코마이신을 포함한 10개의 항생제에 대한 감 수성을 조사한 결과 내성이 없는 것을 확인하였다.

This study evaluated the antimicrobial activity of Endoseal TCS, an mineral trioxide aggregate-based root canal sealer, mixed with water-soluble mangostin derivatives (WsMD) of Garcinia mangostana L. (mangosteen) ethanol extract against Enterococcus faecalis and Staphylococcus aureus. The antibacterial activity of Endoseal TCS mixed with WsMD against three strains of E. faecalis and three strains of S. aureus was performed using agar diffusion test. The data showed that Endoseal TCS mixed with 0.115% WsMD had a zone of inhibition of 0.7 ± 0.2–2.4 ± 0.1 mm. The results suggest that Endoseal TCS mixed with WsMD of Garcinia mangostana L. ethanol extract is useful as a root canal sealer with antibacterial activity against E. faecalis and S. aureus .

This study was conducted to find an efficient and economical mixing ratio of deodorant and a isolated microorganism to reduce ammonia in livestock manure compost. In this study, a simple experimental device that can compare the degree of odor reduction by connecting the vial containing the odor generating source and the gas detection tube and leaving it to stand was used. This test method cannot accurately measure ammonia concentration according to the characteristics of the detector tube, but it is an easy method to compare various experimental conditions. The microorganism isolated from pig manure, “Enterococcus casseliflavus” was found to have an effect on ammonia reduction. Surfactant (sodium dodecyl sulfate), mineral A, mineral B, sulfur, persimmon leaves, and glycerin used as a deodorant were mixed with E. casseliflavus NO-2-L to find the optimum mixing ratio. When 20% of deodorants and E. casseliflavus NO-2-L were added to the source of odor alone, the ammonia reduction efficiency of NO-2-L was the highest (66.7%) compared with other deodorants. When a mixture of microorganisms and deodorants were used, the ammonia reduction efficiency was the best when the mixing ratio of the deodorant was 20%. When mineral B and sulfur were added, the ammonia concentration reduction efficiency was the highest at 83.3%.

In this study, a safety evaluation was conducted to confirm if the Enterococcus faecium CKDB003 strain obtained by selection from a mixed fermentation of fruit and milk is suitable for use as a probiotic. The MIC value for the 10 antibiotics specified in the EFSA guidance was below the acceptable cut-off value. The antibiotic resistance genes aac(6')-li, eatAv, and msr(C) exist by whole genome sequencing, but are in the chromosome and not in the plasmid, thus confirming that there is no possibility of transmission to other microorganisms. It was confirmed that cytolysin (cylA, cylB, cylI, cylL-l, cylL-s, cylM, cylR1, cylR2), aggregation substance (asa1, asp1), collagen adhesion (ace), enterococcal surface protein (esp), endocarditis antigen (efaA), hyaluronidase (hyl) and gelatinase (gelE) were not present in the genome by examining the genes of factors related to virulence. Also, the biochemical analysis showed no toxic enzyme activities, and no virulence genes were detected by the PCR method. Thus, the E. faecium CKDB003 strain can be safely used as a health functional food probiotic, based on the results of the safety assessment.

The creation of probiotic-containing fermented milk products for use by human is an important research topic and has high potential for development. Also, studies have shown that heat-killed probiotics are more stable and easier to use than live probiotics. However, as of this time, research has not been reported in Korea that has evaluated the product or functionality of fermented milk after the addition of heat-killed probiotics. This study was conducted to verify the physiological activity of heat-killed Enterococcus faecalis EF-2001 after its addition to Lactobacillus-fermented milk. Briefly, NFM normal fermented milk (NFM) was used as the control sample, whereas fermented milk with 100 μg/mL EF-2001 (EFM1) and fermented milk with 500 μg/mL EF-2001 (EFM2) were used as the treated samples. Among the samples, EFM2 had the highest acidity of 1.15, but no other factors significantly differed (p<0.05). Furthermore, EFM2 had the highest Lactobacillus count of 9.22 (p<0.05). ABTS, DPPH and FRAP were measured to determine the antioxidant activity of the samples. With respect to those parameters, EFM2 had the highest antioxidant measurements. Therefore, the study confirmed that the addition of E. faecalis EF-2001 to NFM is suitable with the standard and does not affect the quality of characteristics. In conclusion, the treatment sample had higher antioxidant activity than did NFM; this result may be used as a basic for further research and as a guideline for the manufacturing of heat-killed probiotic-containing NFM.

Various viral and bacterial pathogens interact with environmental factors to cause diarrhea in piglets. Enterococcus spp. are Gram-positive anaerobic bacteria that are commonly found in the gastrointestinal tract of several animal species, including pigs. Enterococcus spp. have been reported to infect several animal species as a pathogen. However, gastrointestinal infection by Enterococcus hirae is rare in pigs; only a few cases have been reported worldwide. Four piglets with diarrhea were examined in the diagnostic laboratory of Optipharm Inc. (Cheongju, Korea). During the initial post-mortem examination, no disease lesions were observed. Upon microscopic examination, we found numerous Gram-positive cocci that were adhered to epithelial villi in the jejunum and ileum. However, the villi did not exhibit significant structural damage. Cultured bacteria were identified as E. hirae using the VITEK 2 system and polymerase chain reaction (PCR). Using PCR, we also confirmed that viruses and protozoa that can potentially infect piglet intestines were absent. In antibiotic susceptibility test, the bacteria were resistant to most types of antibiotics. This study presents rare cases of E. hirae infection of the piglet small intestine, which can occur in association with diarrhea possibly by the continuous use of antibiotics.

박테리오신은 미생물이 생산하는 단백질성의 항균물질이다. 본 연구에서는 Enterococcus faecium CJNU 2008 균주로부터 생산되는 박테리오신에 대한 일부 특성을 규명 하였다. 부분 정제 박테리오신은 열처리(100℃ 30분, 121℃ 15분) 및 유기용매(메탄올, 에탄올, 아세톤, 아세토니트릴, 클로로포름)에 대한 안정성이 우수하였으며 효소처리의 경우 Lipase와 α-Amylase에 대해서는 안정하였으나 Protease 처리에서 활성이 소실되었다. 이는 E. faecium CJNU 2008 균주가 생산하는 항균물질이 단백질성의 박테리오신임을 추가적으로 증명하는 것이다. 병원성 세균인 Listeria monocytogenes 균주를 지시균으로 사용했을 때 박테리오신은 살균(bactericidal)의 작용양상을 보였다. Tricine-SDS-PAGE를 이용한 박테리오신의 분자량은 6.5 kDa 이하로 확인되었다. 부분 정제된 박테리오신을 이용하여 HPLC법을 활용한 정제를 수행하였으며 크로마토그램 상에서 단일 피크를 얻었을 수 있었다. 앞으로 정제된 박테리오신은 생화학적 분석 등에 활용할 계획이다.

Enterococcus faecalis is a major causative agent of endodontic treatment failure. The purpose of this study was to investigate bactericidal effects of ethanol extract of Garcinia mangostana L. (mangosteen extract) on five strains of E. faecalis that were isolated from human oral cavities. The bactericidal effects of mangosteen extract were assessed by measurement of minimum bactericidal concentration (MBC) value. The cytotoxicity of mangosteen extract on immortalized human gingival fibroblasts, hTERT-hNOF, was determined based on cell counting method. The data revealed the MBC value of mangosteen extract against the E. faecalis strains was 4 ㎍/ml. Additionally, the cell viability of mangosteen extract on hTERT-hNOF was 83.7-89.1% at the 1 to 16 ㎍/ml. These findings indicated that mangosteen extract could be used as a root canal cleaner during management of endodontic treatment failure caused by E. faecalis.

Probiotics are defined as advantageous microorganisms to human when they are ingested. However, without any protection, the viability of microbes and their adhesive ability to surface of colon decreases through acidic condition such as stomach and intestines. Therefore, many studies have been conducted to figure out to enhance not only the viability of probiotics, but also its adhesion for increasing effect of probiotics. In this study, extrusion method was conducted to encapsulate Enterococcus faecium. E. faecium-alginate solution was injected to CaCl2 solution with regular side air injection. To prevent coagulation of beads, stirring was conducted in CaCl2 solution and encapsulated alginate-Ca2+ microspheres were produced. For optimal encapsulation condition, air pressure was 100 mbar, flow rate of E. faecium solution was 0.02 ml/h and stirring rate was 200 rpm. For mucoadhesive ability, Monolayer of HT-29 cells used as a colon cell and encapsulated cells were inoculated and incubated in 37℃, 5% CO2/95% air atmosphere for 1 h. Encapsulation efficiency of the encapsulation method used in this study was 98.2%. For mucoadhesive test, the concentration of inoculated E. faecium was 9.9×108 CFU/ml and the concentration of adhered E. faecium was 1.6×106 CFU/ml. In conclusion, encapsulation efficiency of extrusion method was high enough to be accepted for this study, however, alginate-Ca2+ microspheres revealed lower adhesive ability compared to expectation. Therefore, it needs further studies to increase adhesive ability with other polymers.

본 연구는 서울시내에서 시판중인 식육에서 E. faecalis 를 분리하고 이 균들의 항생제 내성 패턴, 항생제 유출 펌 프 유전자 및 병독성 유전자의 분포를 분석하였다. 총 277 개의 식육시료에서 93균주의 E. faecalis 를 분리하였다. 이 균주들의 항생제 내성비율은 ampicillin에는 35.5%, chloramphenicol에 6.4%, ciprofloxacin에 4.3%, eryhtromycin 에 18.3%, quinupristin-dalfopristin에 76.3%, tetracycline에 45.2%의 내성이었으며 levofloxacin, teiconplanin 및 vancomycin에는 모든 균이 감수성이었다. 약물 유출펌프인 MFS 타입의 eme(A)와 ABC 타입의 efr(A)유전자는 모든 균주 (100%)에서 확인되었으며 efr(B)는 98.9%, lsa는 91.4%의 균주에서 확인되었다. 병독성 인자인 gel(E)는 68.8%, ace 는 90.3%, asa1는 47.3%, efaA는 91.4%, esp는 12.9%의 균주에서 확인되었다. 본 연구는 시판 식육에서 분리한 지 표 미생물의 하나인 E. faecalis의 항생제 내성, 약물유출 펌프 및 병독서 유전자의 분포를 분석한 연구로 지속적인 모니터링을 하여야 할 것으로 사료된다.

Characteristics and immuno-modulatory effects of Enterococcus (E.) faecium JS1-8 isolated from Kimchi were investigated for potential probiotic use. We measured their acid, bile, and heat tolerances, adhesion properties in Caco-2 cells, antimicrobial activity against pathogenic bacteria, and bacteriocin-like substance-producing activity. Immuno-modulatory effects of E. faecium JS1-8 were measured by determination of nitric oxide (NO), nuclear factor-κB (NF-κB), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) production in RAW 264.7 cells or RAW BLUE cells. JS1-8 survived at pH 2.0 for 2 hr and showed tolerance to 0.3% oxgall bile salt, and it survived after exposure for 5 min at 80°C. JS1-8 showed high antimicrobial inhibition zones to Staphylococcus aureus (460 mm), Listeria monocytogenes (310 mm), Salmonella enteritidis (280 mm), and E. coli (150 mm). Bacteriocin-like substance produced by JS1-8 showed a broad spectrum of activity against Listeria monocytogenes KCCM 40307 and Lactobacillus sake KCCM 40264. Low concentration (1 × 107 CFU/mL) of heat-killed E. faecium JS1-8 induced statistically higher production of NO than Lactobacillus rhamnosus GG (LGG), which is a well-known immuno-modulatory lactic acid bacteria. Low and high (5 × 107 CFU/mL) concentrations of JS1-8 induced statistically higher production of NF-κB than that produced by LGG. We also found that JS1-8 increased production of pro-inflammatory cytokines such as IL-1β and TNF-α in a concentration-dependent manner. As a result, E. faecium JSI-8 could be used as a useful probiotic for controlling pathogens and enhancing host immune responses.

본 연구는 P. aeruginosa와 E. hirae을 대상으로 orthophenylphenol 20%를 함유한 훈증소독제, Fumagari OPP®의 살균효과를 평가하기 위해 수행되었다. 예비 시험에서, P. aeruginosa와 E. hirae의 현탁액 균수는 각각 2.8 × 108 와 3.6 × 108CFU/mL이었으며, 모든 훈증소독제에 노출시킨 담체의 균수는 모두 평판배지법과 여과법으로 배양한 시험균주 현탁액의 균수의 50%보다 많았다. 또한, 대조 담체로부터 회복된 P. aeruginosa와 E. hirae 균수는 각각 2.9 × 106와 2.7 × 106CFU/mL이었다. 훈증소독제의 살균효과 시험에서는, 훈증소독제를 처리한 담체의 P. aeruginosa 와 E. hirae의 감소 균수는 각각 6.46와 5.19 logCFU/mL 로 나타났다. 이상의 결과로부터, 훈증소독제, Fumagari OPP®는 P. aeruginosa와 E. hirae에 대해 효과적인 살균력 을 갖는 것으로 확인되었으며, 병원성 세균에 오염된 식품재료 및 주방용품의 소독에 적용할 수 있을 것으로 사료된다.

The aim of this study was to investigate antimicrobial effect of oleanolic acid (OA), ursolic acid (UA), and sophoraflavanone G against Enterococcus faecalis and Propionibacterium acnes, which are the major causative bacteria of endodontic infections. The antimicrobial activity was evaluated by the minimal inhibitory concentration (MIC). The data showed that the OA, UA, and sophoraflavanone G had antimicrobial effect on all the strains use in the study with 16-64 µg/ml, 8-64 µg/ml, and 1-8 µg/ml of MIC values, respectively. These results indicate that OA, UA, and sophoraflavanone G could be useful in the development of antiseptic solution for washing the root canal in endodontic treatments.

This test was performed to evaluate the bactericidal efficacy of a fumigation disinfectant containing 20% ortho-phenylphenol against Pseudomonas aeruginosa (P. aeruginosa) and Enterococcus hirae (E. hirae). In preliminary tests, P. aeruginosa and E. hirae working culture suspension number (N value) were 2.8 × 108 and 4.0 × 108CFU/mL, respectively. And all the colony numbers on the carriers exposed to the fumigant (n1, n2, n3) were higher than 0.5N1 (the number of bacterial test suspentions by pour plate method), 0.5N2 (the number of bacterial test suspentions by filter membrane method) and 0.5N1, respectively. In addition, the mean number of P. aeruginosa and E. hirae recovered on the control-carriers (T value) was 2.8 × 108 and 3.4 × 106 CFU/mL, respectively. In the bactericidal effect of the fumigant, the reduction number of 2.8 × 108 (d value) was 6.46 and 5.19 logCFU/mL, respectively. According to the French standard for the fumigant, the d value for the effective bactericidal fumigant should be over than 5 logCFU/mL. With the results from this study, the fumigation disinfectant containing 20% ortho-phenylphenol has an effective bactericidal activity, then the fumigant can be applied to disinfect food materials and kitchen appliances contaminated with the pathogenic bacteria.

이 연구는 자돈의 분변에서 분리된 Enterococcus faecim 193이 생산한 박테리오신을 부분정제하고 그 특성을 파악하여 천연 식품보존제 또는 첨가제로서의 이용가능성을 알아보고자 실시되었다. Enterococcus faecium 193의 배양시간에 따른 항균활성을 측정한 결과 항균활성은 3시간째부터 나타나기 시작하여 12시간째에 1280 AU/ml로 최대 활성을 나타내었다. Enterocin 193은 단백질 분해효소(trypsin, pepsin, 및 proteinase K)의 처리에 의해 항균활성이 소실되었으나 넓은 범위의 pH (1-11)와 열처리(121℃, 15분)에 비교적 안정하였다. 또한 50% 농도의 acetone, acetonitrile, chloroform 및 ethanol 등 유기용매에 안정하였으나, 50% methanol을 처리할 경우 항균활성이 50% 감소하였다. 이 박테리오신은 황산암모늄 침전, 소수성 크로마토그래피와 젤 여과 크로마토그래피에 의해 정제되었고 specific activity는 14.8배 증가하였다. SDS-PAGE에 의해 측정한 이 단백질의 분자량은 약 5 kDa이었다.

본 연구는 하수 및 가축분뇨 처리장 방류수, 하천수를 이용하여 반코마이신 내성 장구균(Vancomycin Resistant Enterococci, VRE)의 검출여부를 확인하고 검출된 장구 균의 표현형을 분석함으로써 하천에 대한 공공처리시설 의 방류수에 포함되어있는 엔테로코커스의 영향을 파악 하여 항생제 내성균이 환경에 미치는 영향 및 항생제 내 성 연구에 기초 자료가 되고자 하였다. 엔테로코커스의 동정시험 결과 모두 32균주가 검출되었고 이는 모두 E. faecium으로 동정되었다. Multiplex PCR 시행한 결과 32 균주 모두 Van A 표현형에 해당하는 반코마이신 내성 장 구균으로 확인되었다. 조사지점별 세균의 E. faecium 집 락수의 측정 결과 하수처리장 지역에서 평균적으로 가장 높게 나타났고, 하천 지역의 경우 상류지점에서 하류 지 점으로 갈수록 세균의 colony 숫자도 늘어나는 것으로 조 사되었다. 하수처리장 방류수 시료 26균주에 대한 최소억 제농도 검사 결과, 19개의 항생제 중 내성, 감수성을 보인항생제 수는 각각 14, 5종으로 내성을 보이는 항생제의 수가 약 70% 이상으로 대부분을 차지하였다. 하천 시료 6균주에 대한 최소억제농도 검사 결과, 19개의 항생제 중 내성, 감수성을 보인 항생제 수는 각각 14, 5종으로 내성을 보이는 항생제의 수가 약 70% 이상으로 대부분 을 차지하여 하수처리장 방류수 시료의 검사 결과와 동 일하였다.