• Journal of Microbiology and Biotechnology
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• 제28권11호
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• pp.1823-1833
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• 2018

Fluorescence in-situ hybridization (FISH) is a common and popular method used to investigate microbial communities in natural and engineered environments. In this study, two specific 16S rRNA-targeted oligonucleotide probes, CLZ and KCLZ, were designed and verified to quantify the genus Clostridium and the species Clostridium kluyveri. The optimal concentration of hybridization buffer solution for both probes was 30% (w/v). The specificity of the designed probes was high due to the use of pellets from pure reference strains. Feasibility was tested using samples of Chinese liquor from the famed Luzhou manufacturing cellar. The effectiveness of detecting target cells appears to vary widely in different environments. In pit mud, the detection effectiveness of the target cell by probes CLZ and KCLZ was 49.11% and 32.14%, respectively. Quantitative analysis by FISH technique of microbes in pit mud and fermented grains showed consistency with the results detected by qPCR and PCR-DGGE techniques, which showed that the probes CLZ and KCLZ were suitable to analyze the biomass of Clostridium spp. and C. kluyveri during liquor fermentation. Therefore, this study provides a method for quantitative analysis of Clostridium spp. and C. kluyveri and monitoring their community dynamics in microecosystems.

• Asian-Australasian Journal of Animal Sciences
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• 제28권4호
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• pp.584-591
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• 2015

This study characterized the fecal bacterial community structure and inter-individual variation in 30-week-old Duroc pigs, which are known for their excellent meat quality. Pyrosequencing of the V1-V3 hypervariable regions of the 16S rRNA genes generated 108,254 valid reads and 508 operational taxonomic units at a 95% identity cut-off (genus level). Bacterial diversity and species richness as measured by the Shannon diversity index were significantly greater than those reported previously using denaturation gradient gel electrophoresis; thus, this study provides substantial information related to both known bacteria and the untapped portion of unclassified bacteria in the population. The bacterial composition of Duroc pig fecal samples was investigated at the phylum, class, family, and genus levels. Firmicutes and Bacteroidetes predominated at the phylum level, while Clostridia and Bacteroidia were most abundant at the class level. This study also detected prominent inter-individual variation starting at the family level. Among the core microbiome, which was observed at the genus level, Prevotella was consistently dominant, as well as a bacterial phylotype related to Oscillibacter valericigenes, a valerate producer. This study found high bacterial diversity and compositional variation among individuals of the same breed line, as well as high abundance of unclassified bacterial phylotypes that may have important functions in the growth performance of Duroc pigs.

• Asian-Australasian Journal of Animal Sciences
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• 제28권4호
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• pp.568-572
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• 2015

Acetes chinensis is an economically important shrimp that belongs to the Sergestidae family; following fermentation, A. chinensis' economic value, however, is low in China, and much of the catch in China is exported to Korea at a low price, thus leading to potential false labeling. For this reason, we developed a simple method to identify A. chinensis' origin using allele-specific polymerase chain reaction (PCR). Ten single nucleotide polymorphisms (SNPs) were identified from partial (i.e., 570 bp) DNA sequence analysis of the mitochondrial 16s rRNA gene in 96 Korean and 96 Chinese individual shrimp. Among 10 SNP sites, four sites were observed in populations from both countries, and two sites located in the middle with SNP sites at their 3'-ends were used to design allele-specific primers. Among the eight internal primers, the C220F primer specific to the Chinese A. chinensis population amplified a DNA fragment of 364 bp only from that population. We were able to identify the A. chinensis population origin with 100% accuracy using multiplex PCR performed with two external primers and C220F primers. These results show that the 16S rRNA gene that is generally used for the identification of species can be used for the identification of the origin within species of A. chinensis, which is an important finding for the fair trade of the species between Korea and China.

• Journal of Microbiology and Biotechnology
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• 제28권8호
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• pp.1318-1331
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• 2018

Lettuce (Lactuca sativa L.) is a major ingredient used in many food recipes in South Korea. Lettuce samples were collected during their maximum production period between April and July in order to investigate the microbiota of lettuce during different seasons. 16S rRNA gene-based sequencing was conducted using Illumina MiSeq, and real-time PCR was performed for quantification. The number of total bacterial was greater in lettuce collected in July than in that collected in April, albeit with reduced diversity. The bacterial compositions varied according to the site and season of sample collection. Potential pathogenic species such as Bacillus spp., Enterococcus casseliflavus, Klebsiella pneumoniae, and Pseudomonas aeruginosa showed season-specific differences. Results of the network co-occurrence analysis with core genera correlations showed characteristics of bacterial species in lettuce, and provided clues regarding the role of different microbes, including potential pathogens, in this microbiota. Although further studies are needed to determine the specific effects of regional and seasonal characteristics on the lettuce microbiota, our results imply that the 16S rRNA gene-based sequencing approach can be used to detect pathogenic bacteria in lettuce.

• 한국환경보건학회지
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• 제35권3호
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• pp.162-168
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• 2009

The aim of this study is to investigate microbial sanitary condition of public baths in Seoul, Korea. A total of 28 water samples were collected from 14 different public baths and sudatoriums. The prevalence of fecal indicator microorganisms such as total coliform, fecal coliform, and Escherichia coli was characterized. In addition, bacteria in water was membrane filtered by 0.45um nitrocellulose membrane, and the filter was analyzed by both cultivation and PCR amplification of partial 16S rRNA gene. The levels of chlorine were measured for each of water samples. More than 40% of 14 collected water samples, the concentrations of total coliform bacteria exceeded the water quality for bath water guideline. There was no significant correlation between chlorine residue and the presence of total coliform. Various microorganisms including pathogenic microorganisms were identified from cultivation and subsequent analysis of 16s rRNA gene sequences. Our results suggest that appropriate hygiene practice and continuous monitoring is needed for reducing health risk associated with public bathhouses.

• Animal Systematics, Evolution and Diversity
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• 제36권2호
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• pp.113-122
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• 2020

From a moss sample, we isolated and identified Notohymena gangwonensis Kim et al., 2019 based on morphological and molecular data. The moss and type population has completely identical 18S rRNA (nuclear small subunit ribosomal RNA) gene sequences and both are highly similar in morphological and morphometric attributes, except for the diameter and arrangement of the cortical granules. Thus, we reexamined the type materials(i.e., micrographs and gDNA) and resulted in finding mistakes made by the authors of the species. Based on these data and supporting materials newly obtained (i.e., internal transcribed spacer [ITS] 1, ITS2, 5.8S, and partial 28S rDNA sequences, and scanning electron micrographs), we provide improved diagnosis of the species to clarify its identity. In addition, a key for Notohymena species is provided.

• 한국미생물·생명공학회지
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• 제33권1호
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• pp.24-28
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• 2005

전통 된장으로부터 단백질 분해능이 있으면서 혈전용해활성이 우수한 D-1 균주를 분리하였다. 분리된 D-1 균주는 표준물질인 plasmin 2.51 $\mu}g$과 같은 혈전용해효소활성을 보였으며 이 균주를 동정하기 위하여 세포막 지방산 조성 분석과 탄수화물 이용성을 조사한 결과 김 등[12], 허 등[7] 청국장에서 혈전용해효소활성을 보이는 균주로 보고된 Bacillus subtilis로 유사하게 나타났다. 탄수화물 이용성, 세포막 지방산 분석 그리고 16S rRNA sequence를 통하여 Bacillus속으로 동정 할 수 있었고, 이에 된장으로부터 분리한 혈전용해효소를 생산하는 D-1균주를 최종적으로 Bacillus sp. D-1으로 명명하였으며 앞으로 이 균주를 이용하여 혈전용해효소를 함유하는 기능성 된장과 청국장 제조에 큰 효과를 보일 것으로 기대된다.

• 한국토양비료학회지
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• 제39권4호
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• pp.185-194
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• 2006

몇 년동안 게껍질이 풍부하게 있었던 밭토양에서 강한 키틴분해능력을 가진 Trichoderma 곰팡이를 분리하였다. 분리된 곰팡이의 5.8S rRNA, partial 18S, 28S rRNA genes, ITS1, ITS2 sequence 분석과 형태학적 특징을 살펴본 결과 Trichoderma asperellum으로 동정되었고, 이를 Trichoderma asperellum T-5 (TaT-5)로 명명하였다. 이 곰팡이는 chitianse와 ${\beta}$-1,3-glucanase같은 lytic enzyme을 분비하며, 키틴배지 상에서 6가지의 항 곰팡이성 물질을 생산했다. R. solani가 원인인 오이의 모잘록병에 대해 TaT 5의 방제효과를 보기 위해서 TaT-5 배양액(TA), chitin medium(CM), 증류수(DW)를 씨를 심은 후 10일 째에 각 pot에 관주했다. 그리고 관주 1주일 후에 R. solani의 균사를 갈아서 각 pot에 주었다. 실험기간 동안에 오이의 지상부와 지하부 생체중은 다른 처리구에 비하여 TA 처리구가 더 많이 증가하였다. 오이 잎에서 PR-protein (chitianse, ${\beta}$-1,3-glucanase) 활성은 R. solani 감염 후 CM과 DW에서 증가를 보였고, TA 처리구에서는 증가하다가 감소하는 경향을 보였다. 뿌리에서는 모든 처리구가 감소하는 경향을 보였지만 TA 처리구가 CM과 DW 처리구보다 감소하는 정도가 적었다. 오이의 잎과 뿌리에서 lignification related enzyme(POD, PPO, PAL)활성은 R. solani 감염 초기에는 증가하다가 점점 감소하는 경향을 보였다. 이러한 결과들은 TaT-5에 의하여 생산된 lytic enzymes와 항 곰팡이성 물질들이 오이에 R. solani의 공격을 줄여준다고 생각된다.

• 한국식품위생안전성학회지
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• 제28권2호
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• pp.188-193
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• 2013

Aphanizomenon flos-aquae는 시아노박테리아 일종이며 anatoxin-a, saxitoxin, neosaxitoxin 등의 독소를 생산할 수 있어 국내에서는 식품원료로 사용이 금지되어있다. 전통적으로 시아노박테리아는 사상체 넓이, 세포 크기, 분열방법, 세포형태, 가스주머니의 존재유무 등의 형태학적 특징에 의한 분류가 가능하다. 그러나 가스주머니 혹은 무성포자와 같은 특징은 주변 환경 또는 생장조건에 따라 차이가 있으며 경우에 따라 소실되기도 한다. 따라서 PCR에 의한 Aph. flos-aquae를 함유하는 기능식품을 검출할 수 있는 분석법을 개발하였다. 프라이머를 설계하기 위하여 유전자은행(www.ncbi.nlm.nih.gov)에 등록되어있는 Aph. flos-aquae, 스피루리나의 16S rRNA 염기서열을 이용하였으며, 비교 및 분석에는 BioEdit ver. 7.0.9.0 프로그램을 사용하였다. 최종적으로 클로렐라, 스피루리나, 녹차, 시금치로부터 Aph. flos-aquae를 검출할 수 있는 AFA-F1/AFA-R1(363 bp) 프라이머를 최종 선정하였다. 그리고 상기 프라이머는 Aph. flos-aquae가 각각 1% 함유 되도록 제조된 클로렐라, 스피루리나 제품에서 모두 혼입여부의 확인이 가능함을 확인하였다.

• 생명과학회지
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• 제28권2호
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• pp.257-264
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• 2018

YrdC 수퍼 패밀리는 지금까지 유전 서열이 알려진 거의 모든 생명체에서 매우 잘 보존 된 단백질 중 하나이다. Escherichia coli의 YrdC는 리보솜 생합성, 번역 종결, 저온 적응, tRNA에서 threonylcarbamoyl adenosine의 형성에 관여하는 것으로 제안되었다. 이 연구에서, yrdC 유전자가 대장균에서 필수적이라는 것을 명확하게 증명하기 위해, 대장균에서 두 개의 yrdC 결손 돌연변이 균주를 만들고 그 표현형을 조사하였다. 특히 온도에 민감한 yrdC 돌연변이 균주는 $42^{\circ}C$ 온도 조건 하에서 거의 즉시 세포 성장을 멈추었으며 30S 리보솜 단위체의 상당한 축적없이 16S rRNA 전구체를 축적하는 것으로 나타났다. 또한 효모와 인간의 yrdC 유전자를 클로닝하여 이들이 대장균 yrdC 결손 균주의 성장억제를 회복 할 수 있다는 것을 입증하였다. 이밖에도 여러 돌연변이 연구에 의해, 우리는 YrdC 단백질의 중간에 위치한 오목한 표면이 대장균, 효모 및 인간의 YrdC 단백질에서 중요한 역할을 한다는 것을 보여 주었다. 따라서, 두 개의 yrdC 결손 균주를 비교하여, yrdC 유전자가 대장균에서 생존력에 필수적이며, 효모 및 인간 동족체의 기능이 대장균 YrdC의 기능과 중복된다는 것을 규명하였고, 이 균주를 이용하여 아직까지 밝혀지지 않은 대장균 YrdC 단백질이 tRNA 형성에 관여한다는 것을 증명할 수 있는 토대를 제공한다는데 의의가 있다.