• Journal of Microbiology and Biotechnology
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• 제25권8호
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• pp.1205-1215
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• 2015

Arbuscular mycorrhizal fungi (AMF) have great potential for assisting heavy metal hyperaccumulators in the remediation of contaminated soils. However, little information is available about the symbiosis of AMF associated with an antimony (Sb) accumulator plant under natural conditions. Therefore, the objective of this study was to investigate the colonization and molecular diversity of AMF associated with the Sb accumulator ramie (Boehmeria nivea) growing in Sb-contaminated soils. Four Sb mine spoils and one adjacent reference area were selected from Xikuangshan in southern China. PCR-DGGE was used to analyze the AMF community composition in ramie roots. Morphological identification was also used to analyze the species in the rhizosphere soil of ramie. Results obtained showed that mycorrhizal symbiosis was established successfully even in the most heavily polluted sites. From the unpolluted site Ref to the highest polluted site T4, the spore numbers and AMF diversity increased at first and then decreased. Colonization increased consistently with the increasing Sb concentrations in the soil. A total of 14 species were identified by morphological analysis. From the total number of species, 4 (29%) belonged to Glomus, 2 (14%) belonged to Acaulospora, 2 (14%) belonged to Funneliformis, 1 (7%) belonged to Claroideoglomus, 1 (7%) belonged to Gigaspora, 1 (7%) belonged to Paraglomus, 1 (7%) belonging to Rhizophagus, 1 (7%) belonging to Sclervocystis, and 1 (7%) belonged to Scutellospora. Some AMF sequences were present even in the most polluted site. Morphological identification and phylogenetic analysis both revealed that most species were affiliated with Glomus, suggesting that Glomus was the dominant genus in this AMF community. This study demonstrated that ramie associated with AMF may have great potential for remediation of Sb-contaminated soils.

• Journal of Microbiology and Biotechnology
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• 제24권3호
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• pp.363-370
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• 2014

Campylobacter jejuni is a prevalent foodborne pathogen worldwide. Human infection by C. jejuni primarily arises from contaminated poultry meats. Genes expressed in vivo may play an important role in the pathogenicity of C. jejuni. We applied an immunoscreening method, in vivo-induced antigen technology (IVIAT), to identify in vivo-induced genes during human infection by C. jejuni. An inducible expression library of genomic proteins was constructed from sequenced C. jejuni NCTC 11168 and was then screened using adsorbed, pooled human sera obtained from clinical patients. We successfully identified 24 unique genes expressed in vivo. These genes were implicated in metabolism, molecular biosynthesis, genetic information processing, transport, and other processes. We selected six genes with different functions to compare their expression levels in vivo and in vitro using real-time RT-PCR. The results showed that the selected six genes were significantly upregulated in vivo but not in vitro. In short, these identified in vivo-induced genes may contribute to human infection of C. jejuni, some of which may be meaningful vaccine candidate antigens or diagnosis serologic markers for campylobacteriosis. IVIAT may present a significant and efficient method for understanding the pathogenicity mechanism of Campylobacter and for finding targets for its prevention and control.

• 대한환경공학회지
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• 제33권11호
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• pp.847-854
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• 2011

주요 신재생에너지인 바이오에너지의 일환으로 조류를 이용한 바이오에너지 및 자원화 기술에 대한 관심이 높아지고 있다. 조류는 영양염류 제거 능력을 활용해서 하수와 같은 오폐수 내 난분해성오염물질과 영양염류 제거의 고도처리도 가능하다. 조류와 박테리아 간의 생태적인 상호작용이 조류를 활용한 하수처리 및 하수자원화에 중요한 역할을 함에도 불구하고, 실지 하수 조건에서 조류와 박테리아간의 생태학적인 상호작용에 관한 과학적인 정보가 부족하다. 본 연구에서는 하수에서 배양이 잘 되고, 지질함량이 높다고 알려진 국내 조류 종인 Ankistrodesmus gracilis SAG 278-2의 하수오염물질 제거 특성과 조류 주입에 따른 하수 박테리아 군집의 반응을 실지 하수 조건에서 연구하였다. 하수 박테리아의 수가 증가는 조류의 성장 속도를 감소시켰으나, 반면 조류의 성장은 박테리아의 생존 및 내성호흡 생분해 속도에는 영향을 주지 않았다. 조류가 주입된 하수에서 난분해성 유기물질 및 총질소의 제거 향상이 관찰되었다. 박테리아 16S rRNA 유전자 T-RFLP 분석에 따르면 조류의 주입은 시간에 따라 박테리아 군집에 영향을 주었다. 박테리아 16S rRNA 유전자 PCR 증폭, clone 및 염기서열 분석 결과, 하수 내 조류의 성장은 박테리아 군집 구성을 변화시키며, 조류와 함께 공동 성장 가능한 박테리아는 Sediminibacterium, Sphingobacterium, Mucilaginibacter 속에 속하는 개체로 판명되었다.

• 한국가축번식학회지
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• 제26권2호
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• pp.87-94
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• 2002

본 연구는 섬유소분해효소 유전자(CelD)가 도입된 형질전환 돼지를 생산하기 위해 사계절동안 수행하였다. 약 8∼15개월령의 순종의 랜드레이스경산돈 및 미경산돈 126두는 유전자 미세주입을 위한 1세포기 단계의 수정란 채란 및 이식을 위해 사용하였으며, 발정동기화 및 과배란 방법은 PG600 주입 후 9일간 매일 20mg의 altrenogest를 사료에 첨가하여 급여하였다. Altrenogest를 9일간 급여 후 1000IU의 PMSG와 750IU의 hCG를 주입하므로서 과배란을 유도하였다. 미세주입을 위한 유전자는 Rat elasterase promoter에 CelD유전자를 연결하여 준비하였으며, 호르몬 처리후 91두의 공란돈으로부터 1,422개의 난자를 회수하였으며 이중 95.6% (1,359/1,422)는 DNA미세주입을 위해 전핵을 관찰 할 수 있는 1세포기의 수정란이었다. 이중 유전자가 미세주입된 725개의 난자를 35두의 수란돈에 이식하였으며 13두의 임신돈으로부터 65두의 자돈을 생산하였다. 한편 수란돈당 수정란 이식수가 임신율에 미치는 영향을 조사한 결과 약 21∼24개의 수정란을 이식한 구에서 임신율이 50%로 타구의 20.0%(20개 이하)와 33.3%(25개 이상)보다 높았으며, 꼬리조직으로부터 분리된 DNA의 PCR검정결과 65두중 5두가 형질전환 양성 반응을 나타내어 7.69%의 형질전환율을 나타내었다 따라서 본 연구는 생체반응기를 통한 형질전환 돼지생산을 위한 유용한 정보를 제공하게 될 것이다.

• 대한방사성의약품학회지
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• 제1권2호
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• pp.118-122
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• 2015

Suicidal gene therapy is based on the transduction of tumor cells with "suicide" genes encoding for prodrug-activating enzymes that render target cells susceptible to prodrug treatment. Suicidal gene therapy results in the death of tumor with the expression of gene encoding enzyme that converts non-toxic prodrug into cytotoxic product. Cytochrome P450 4B1 (CYP4B1) activates 4-ipomeanol (4-IPO) or 2-aminoanthracene (2-AA) to cytotoxic furane epoxide and unsaturated dialdehyde intermediate.In this study, therapeutic effects of suicidal gene therapy with rabbit CYP4B1/2-AA or 4-IPO system were evaluated in HT-29 (human colon cancer cell). pcDNA-CYP4B1 vector was transfected into HT-29 by lipofection and stable transfectant was selected by treatment of hygromycin ($500{\mu}g/mL$) for 3 weeks. Reverse transcription polymerase chain reaction (RT-PCR) analysis was performed for confirmation of CYP4B1 expression in CYP4B1 gene transduced cell. The cytotoxic effects of CYP4B1 transduced cell were determined using dye-exclusion assay after treatment of 2-AA or 4-IPO for 96 hrs. Dye-exclusion assay showed that $IC_{50}$ of HT-29 and CYP4B1 transduced HT-29 was 0.01 mM and 0.003 mM after 4-IPO or 2-AA treatment at 96 hrs exposure, respectively. In conclusion, CYP4B1 based prodrug gene therapy probably have the potential for treatment of colorectal adenocarcinoma.

• 한국잠사곤충학회지
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• 제49권2호
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• pp.37-42
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• 2007

락토페린 cDNA 유전자를 도입시킨 누에 형질전환 실험을 수행한 결과, 다음과 같은 결과를 얻을 수 있었다. 1. 사람 GI-101 세포주의 mRNA로부터 클로닝 된 락토페린 cDNA 유전자의 개시코돈 ATG와 종결코돈 TAA를 포함하는 open reading frame(2,136 bp) 영역을 확인하였다. 2. Sf9 배양세포의 조추출물 시료에 의한 Western blot 분석 결과, 락토페린으로 추정되는 약 80kDa의 단백질 발현을 확인하였다. 3. 누에 형질전환에 높은 전이효율과 활성을 나타내는 트랜스포존을 이용한 전이벡터 pPIGA3GFP를 개조하여 락토페린 cDNA를 삽입시킨 전이벡터 pPT-HLf를 구축하였다. 4. DNA 미량 주사법에 의한 누에 형질전환 개체의 발현 비율은 약 6.7% 정도를 나타냈다. 5. 형질전환 누에(G0) 동일한 세대간 교배 및 처리하지 않은 성충간의 역교배에 의한 차세대(G1) 개체로부터 락토페린 유전자와 동일한 크기의 2.1 kb DNA 단편을 확인 할 수 있었으며, 형질전환 G1 세대의 조추출물 시료에 의한 Western blot 분석 결과, 표준 락토페린 항체와 반응하는 약 80 kDa의 단백질 발현을 확인할 수 있었다.

• 한국임상수의학회지
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• 제32권4호
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• pp.324-329
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• 2015

The primary aim of this study was to investigate biosecurity practices in pig farms and to determine the major risk factors associated with PCV2 infection for a sampled swine population in Korea. To this end, we analyzed data from a cross-sectional study of 296 farrow-to-finish farms, which was conducted between March and September 2014 to explore the prevalence of swine disease at farm level. Face-to-face interviews by on-site visit of trained veterinarians were conducted with the farm owners or managers using a standardized questionnaires with information about basic demographical data and management practices. Farms were classified as negative or positive through the use of infection profiles that combined data on serological testing including PCR antigen test result, antibody titer and sero-conversion pattern at each age category taking into account vaccination status. Data were analyzed using multivariate ordinal logistic regression. Results from this study indicated that biosecurity level of the farms was considered not good given low compliance of the biosecurity programs and facilities in the farm: off-site removal of dead stocks (7%), off-site location of storage facility for incoming feeds (12.6%), off-site pick-up location for finishers (19.3%), restrictions on feed supplier vehicles for farm entrance (19.6%), restriction of finisher trucks entering the farm (22.4%), and restriction on manure disposal trucks entering the farm (26.4%). In the final model (n = 255), allowance of finisher truck driver to the pig unit had increased risk of infection (OR = 2.4, 95% CI 1.22-4.67) whereas farms with a sign forbidding the entrance had decreased risk of infection (OR = 0.19, 95% CI 0.10-0.58). Further comprehensive research with larger sample size is required to better understand the multifactorial characteristic that some predisposing risk factors that were not available in this study. To the best knowledge of the authors, this was the first study to use empirical data to report risk factors associated with PCV2 infection in the Korean pig farms. Results from the current study could be used to decide optimal biosecurity measures to reduce the impact of PCV2 infection to farmers and policy makers.

• Journal of Microbiology
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• 제43권4호
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• pp.345-353
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• 2005

The complex ecosystem of intestinal micro flora is estimated to harbor approximately 400 different microbial species, mostly bacteria. However, studies on bacterial colonization have mostly been based on culturing methods, which only detect a small fraction of the whole microbiotic ecosystem of the gut. To clarify the initial acquisition and subsequent colonization of bacteria in an infant within the few days after birth, phylogenetic analysis was performed using 16S rDNA sequences from the DNA iso-lated from feces on the 1st, 3rd, and 6th day. 16S rDNA libraries were constructed with the amplicons of PCR conditions at 30 cycles and $50^{\circ}C$ annealing temperature. Nine independent libraries were produced by the application of three sets of primers (set A, set B, and set C) combined with three fecal samples for day 1, day 3, and day 6 of life. Approximately 220 clones ($76.7\%$) of all 325 isolated clones were characterized as known species, while other 105 clones ($32.3\%$) were characterized as unknown species. The library clone with set A universal primers amplifying 350 bp displayed increased diversity by days. Thus, set A primers were better suited for this type of molecular ecological analysis. On the first day of the life of the infant, Enterobacter, Lactococcus lactis, Leuconostoc citreum, and Streptococcus mitis were present. The largest taxonomic group was L. lactis. On the third day of the life of the infant, Enterobacter, Enterococcus faecalis, Escherichia coli, S. mitis, and Streptococcus salivarius were present. On the sixth day of the life of the infant, Citrobacter, Clostridium difficile, Enterobacter sp., Enterobacter cloacae, and E. coli were present. The largest taxonomic group was E. coli. These results showed that microbiotic diversity changes very rapidly in the few days after birth, and the acquisition of unculturable bacteria expanded rapidly after the third day.

• 동의생리병리학회지
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• 제20권1호
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• pp.93-97
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• 2006

To investigate whether GyeongshinhaeGihwan 1(GGT1), an anti-obesity herbal medicine widely used in oriental medicine, regulates the expression of obesity-related genes, we measured the changes in mRNA levels of these genes by GGT1 in human growth hormone transgenic (hGHTg) obese male rats, and these effects by GGT1 were compared with those of reductil (RD), an anti-obesity drug approved by FDA. Rats received once daily oral administrations of autoclaved water, RD, or GGT1 for 8 weeks. At the end of study, rats were sacrificed and tissues were harvested. Total RNA from adipose tissue, liver and kidney was prepared and the mRNA levels for LPL (lipoprotein lipase), PPAR $\gamma$ (peroxisome proliferator activated receptor-gamma), PPAR$\delta$ (peroxisome proliferator activated receptor-delta), leptin, TNF$\alpha$ (tumor necrosis factor-alpha), and internal standard G3PDH (glyceraldehyde-3- phosphate dehydrogenase) were analyzed by RT-PCR. PPAR$\gamma$ mRNA levels of liver and kidney were decreased in drug-treated groups compared with control group and the decrease of PPAR$\gamma$ expression was more prominent in GGT1 group than in RD group, suggesting that GGT1 is effective in the inhibition of adipogenesis and lipid storage by decreasing the PPAR$\gamma$ expression. In contrast, PPAR$\delta$ mRNA levels of adipose tissue and kidney were increased by RD and GGT1 , and the magnitudes of increase were higher in GGT1 group than in RD group, indicating that GGT1 stimulates fatty acid oxidation and energy metabolism by activating PPAR$\delta$ expression, Compared with control and RD groups, GGT1 group had higher concentrations of serum leptin, a well-known inhibitor of appetite. However, The mRNA levels of leptin, LPL, and TNF$\alpha$ were not changed by GGT1 and RD, compared with DW. These results demonstrate that GGT1 not only decreases PPAR$\gamma$ expression of liver and kidney, but also increases PPAR$\delta$ expression of adipose tissue and kidney, leading to the regulation of obesity and that these effects were more pronounced in GGT1 group compared with RD group. In addition, GGT1 seems to prevent obesity by increasing the serum leptin levels.

• Parasites, Hosts and Diseases
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• 제53권2호
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• pp.155-162
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• 2015

Toxoplasma gondii is a protozoan parasite with a broad range of intermediate hosts. Chickens as important food-producing animals can also serve as intermediate hosts. To date, experimental studies on the pathogenicity of T. gondii in broiler chickens were rarely reported. The objective of the present study was to compare the pathogenicity of 5 different T. gondii strains (RH, CN, JS, CAT2, and CAT3) from various host species origin in 10-day-old chickens. Each group of chickens was infected intraperitoneally with $5{\times}10^8$, $1{\times}10^8$, $1{\times}10^7$, and $1{\times}10^6$ tachyzoites of the 5 strains, respectively. The negative control group was mockly inoculated with PBS alone. After infection, clinical symptoms and rectal temperatures of all the chickens were checked daily. Dead chickens during acute phage of the infection were checked for T. gondii tachyzoites by microscope, while living cases were checked for T. gondii infection at day 53 post-inoculation (PI) by PCR method. Histopathological sections were used to observe the pathological changes in the dead chickens and the living animals at day 53 PI. No significant differences were found in survival periods, histopathological findings, and clinical symptoms among the chickens infected with the RH, CN, CAT2, and CAT3 strains. Histopathological findings and clinical symptoms of the JS (chicken origin) group were similar to the others. However, average survival times of infected chickens of the JS group inoculated with $5{\times}10^8$ and $1{\times}10^8$ tachyzoites were 30.0 and 188.4 hr, respectively, significantly shorter than those of the other 4 mammalian isolates. Chickens exposed to $10^8$ of T. gondii tachyzoites and higher showed acute signs of toxoplasmosis, and the lesions were relatively more severe than those exposed to lower doses. The results indicated that the pathogenicity of JS strain was comparatively stronger to the chicken, and the pathogenicity was dose-dependent.