• Genomics & Informatics
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• 제21권3호
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• pp.40.1-40.11
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• 2023

Microbial community profiling using 16S rRNA amplicon sequencing allows for taxonomic characterization of diverse microorganisms. While amplicon sequence variant (ASV) methods are increasingly favored for their fine-grained resolution of sequence variants, they often discard substantial portions of sequencing reads during quality control, particularly in datasets with large number samples. We present a streamlined pipeline that integrates FastP for read trimming, HmmUFOtu for operational taxonomic units (OTU) clustering, Vsearch for chimera checking, and Kraken2 for taxonomic assignment. To assess the pipeline's performance, we reprocessed two published stool datasets of normal Korean populations: one with 890 and the other with 1,462 independent samples. In the first dataset, HmmUFOtu retained 93.2% of over 104 million read pairs after quality trimming, discarding chimeric or unclassifiable reads, while DADA2, a commonly used ASV method, retained only 44.6% of the reads. Nonetheless, both methods yielded qualitatively similar β-diversity plots. For the second dataset, HmmUFOtu retained 89.2% of read pairs, while DADA2 retained a mere 18.4% of the reads. HmmUFOtu, being a closed-reference clustering method, facilitates merging separately processed datasets, with shared OTUs between the two datasets exhibiting a correlation coefficient of 0.92 in total abundance (log scale). While the first two dimensions of the β-diversity plot exhibited a cohesive mixture of the two datasets, the third dimension revealed the presence of a batch effect. Our comparative evaluation of ASV and OTU methods within this streamlined pipeline provides valuable insights into their performance when processing large-scale microbial 16S rRNA amplicon sequencing data. The strengths of HmmUFOtu and its potential for dataset merging are highlighted.

• ALGAE
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• 제38권4호
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• pp.265-281
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• 2023

Hypoxia can indeed impact the survival of protists, which play a crucial role in marine ecosystems. To better understand the protistan community structure and species that can thrive in hypoxic waters, we collected samples from both the surface and bottom waters during the hypoxic period in Jinhae and Masan Bays and the non-hypoxic period in Jinhae Bay. Subsequently, we utilized metabarcoding techniques to identify the protistan species. During hypoxia, with dissolved oxygen concentrations of 0.8 mg L^-1 in Jinhae Bay and 1.8 mg L^-1 in Masan Bay within the bottom waters, the phylum Dinoflagellata exhibited the highest amplicon sequence variants richness among the identified protist phyla. Following the Dinoflagellata, Ochrophyta and Ciliophora also displayed notable presence. In hypoxic waters of Jinhae and Masan Bays, we identified a total of 36 dinoflagellate species that exhibited various trophic modes. These included one autotrophic species, 14 mixotrophic species, 9 phototrophic species with undetermined trophic modes (either autotrophic or mixotrophic), 2 kleptoplastidic species, and 10 heterotrophic species. Furthermore, the hypoxic bottom water exhibited a greater number of heterotrophic dinoflagellate species compared to the non-hypoxic surface water within the same water column or the non-hypoxic bottom water. Therefore, feeding by mixotrophic and heterotrophic dinoflagellates may be partially responsible for their dominance in terms of the number of species surviving in hypoxic waters. This study not only introduces the initial documentation of 26 dinoflagellate species surviving in hypoxic conditions but also establishes a foundation for a more comprehensive understanding of the ecophysiology of dinoflagellates in hypoxic marine environments.

• Asian Pacific Journal of Cancer Prevention
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• 제16권17호
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• pp.7935-7941
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• 2015

The aim of this study was to implement massive parallel sequencing (MPS) technology in clinical genetics testing. We developed and tested an amplicon-based method for resequencing the BRCA1 and BRCA2 genes on an Illumina MiSeq to identify disease-causing mutations in patients with hereditary breast or ovarian cancer (HBOC). The coding regions of BRCA1 and BRCA2 were resequenced in 96 HBOC patient DNA samples obtained from different sample types: peripheral blood leukocytes, whole blood drops dried on paper, and buccal wash epithelia. A total of 16 random DNA samples were characterized using standard Sanger sequencing and applied to optimize the variant calling process and evaluate the accuracy of the MPS-method. The best bioinformatics workflow included the filtration of variants using GATK with the following cut-offs: variant frequency >14%, coverage ($>25{\times}$) and presence in both the forward and reverse reads. The MPS method had 100% sensitivity and 94.4% specificity. Similar accuracy levels were achieved for DNA obtained from the different sample types. The workflow presented herein requires low amounts of DNA samples (170 ng) and is cost-effective due to the elimination of DNA and PCR product normalization steps.

• 한국자원식물학회지
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• 제25권3호
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• pp.308-316
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• 2012

The purpose of this study was to identify the variant of Platanus occidentalis, whose bark looks white, also can be classified as P. occidentalis and to examine its genetic difference from the general P. occidentalis. For the variant identification of P. occidentalis, SNP and ISSR analysis were used in this study. Thirteen samples of P. occidentalis white variant were collected in Cheongju and 24 samples of normal P. occidentalis obtained in Cheongju, Pyongtaek, Ansan, Suwon, Osan and Jincheon area. ITS 1 and ITS 2 sequences of white variants were identical with those of P. occidentalis. We could not find any sequence difference between normal and white P. occidentalis. So we concluded that the white variant belongs to normal P. occidentalis even their bark is white and peeled easily. By ISSR test, 98 amplicons were acquired using 10 primers. P. occidentalis and white P. occidentalis showed different band patterns from the UBC #834. According to the result of Nei (1979)'s genetic distance analysis, the members of white P. occidentalis were grouped more tightly than the members of normal P. occidentalis. The UPGMA dendrogram shows that the variant and P. occidentalis divided widely into two groups. These results show that the phenotype of P. occidentalis white variant is caused by genetic factors rather than by environmental factors.

• The Plant Pathology Journal
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• 제39권3호
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• pp.245-254
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• 2023

The plant microbiota plays a crucial role in promoting plant health by facilitating the nutrient acquisition, abiotic stress tolerance, biotic stress resilience, and host immune regulation. Despite decades of research efforts, the precise relationship and function between plants and microorganisms remain unclear. Kiwifruit (Actinidia spp.) is a widely cultivated horticultural crop known for its high vitamin C, potassium, and phytochemical content. In this study, we investigated the microbial communities of kiwifruit across different cultivars (cvs. Deliwoong and Sweetgold) and tissues at various developmental stages. Our results showed that the microbiota community similarity was confirmed between the cultivars using principal coordinates analysis. Network analysis using both degree and eigenvector centrality indicated similar network forms between the cultivars. Furthermore, Streptomycetaceae was identified in the endosphere of cv. Deliwoong by analyzing amplicon sequence variants corresponding to tissues with an eigenvector centrality value of 0.6 or higher. Our findings provide a foundation for maintaining kiwifruit health through the analysis of its microbial community.

• Parasites, Hosts and Diseases
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• 제61권1호
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• pp.42-52
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• 2023

The genus Babesia includes parasites that can induce human and animal babesiosis, which are common in tropical and subtropical regions of the world. The gut microbiota has not been examined in hamsters infected by Babesia duncani. Red blood cells infected with B. duncani were injected into hamsters through intraperitoneal route. To evaluate the changes in gut microbiota, DNAs were extracted from small intestinal contents, acquired from hamsters during disease development. Then, the V4 region of the 16S rRNA gene of bacteria was sequenced using the Illumina sequencing platform. Gut microbiota alternation and composition were assessed according to the sequencing data, which were clustered with >97.0% sequence similarity to create amplicon sequence variants (ASVs). Bacteroidetes and Firmicutes were made up of the major components of the gut microbiota in all samples. The abundance of Bacteroidetes elevated after B. duncani infection than the B. duncani-free group, while Firmicutes and Desulfobacterota declined. Alpha diversity analysis demonstrated that the shown ASVs were substantially decreased in the highest parasitemia group than B. duncani-free and lower parasitemia groups. Potential biomarkers were discovered by Linear discriminant analysis Effect Size (LEfSe) analysis, which demonstrated that several bacterial families (including Muribaculaceae, Desulfovibrionaceae, Oscillospiraceae, Helicobacteraceae, Clostridia UGG014, Desulfovibrionaceae, and Lachnospiraceae) were potential biomarkers in B. duncani-infected hamsters. This research demonstrated that B. duncani infectious can modify the gut microbiota of hamsters.

• Animal Bioscience
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• 제36권9호
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• pp.1453-1464
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• 2023

Objective: This study investigated the changes in bacterial communities within decomposing swine microcosms, comparing soil with or without intact microbial communities, and under aerobic and anaerobic conditions. Methods: The experimental microcosms consisted of four conditions: UA, unsterilized soil-aerobic condition; SA, sterilized soil-aerobic condition; UAn, unsterilized soil-anaerobic condition; and San, sterilized soil-anaerobic condition. The microcosms were prepared by mixing 112.5 g of soil and 37.5 g of ground carcass, which were then placed in sterile containers. The carcass-soil mixture was sampled at day 0, 5, 10, 30, and 60 of decomposition, and the bacterial communities that formed during carcass decomposition were assessed using Illumina MiSeq sequencing of the 16S rRNA gene. Results: A total of 1,687 amplicon sequence variants representing 22 phyla and 805 genera were identified in the microcosms. The Chao1 and Shannon diversity indices varied in between microcosms at each period (p<0.05). Metagenomic analysis showed variation in the taxa composition across the burial microcosms during decomposition, with Firmicutes being the dominant phylum, followed by Proteobacteria. At the genus level, Bacillus and Clostridium were the main genera within Firmicutes. Functional prediction revealed that the most abundant Kyoto encyclopedia of genes and genomes metabolic functions were carbohydrate and amino acid metabolisms. Conclusion: This study demonstrated a higher bacteria diversity in UA and UAn microcosms than in SA and SAn microcosms. In addition, the taxonomic composition of the microbial community also exhibited changes, highlighting the impact of soil sterilization and oxygen on carcass decomposition. Furthermore, this study provided insights into the microbial communities associated with decomposing swine carcasses in microcosm.

• Journal of Ecology and Environment
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• 제47권4호
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• pp.168-176
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• 2023

Background: Many insectivorous bats have flexible diets, and the difference in prey item consumption among species is one of the key mechanisms that allows for the avoidance of interspecies competition and promotes coexistence within a microhabitat. In Korea, of the 24 bat species that are known to be distributed, eight insectivorous bats use forest areas as both roosting and foraging sites. Here, we aimed to understand the resource partitioning and coexistence strategies between two bat species, Myotis ikonnikovi and Plecotus ognevi, cohabiting the Mt. Jumbong forests, by comparing the differences in dietary consumption based on habitat utilization. Results: Upon examining their dietary composition using the DNA meta-barcoding approach, we identified 403 prey items (amplicon sequence variants). A greater prey diversity including Lepidoptera, Diptera, Coleoptera, and Ephemeroptera, was detected from M. ikonnikovi, whereas most prey items identified from P. ognevi belonged to Lepidoptera. The diversity index of prey items was higher for M. ikonnikovi (H': 5.67, D: 0.995) than that for P. ognevi (H': 4.31, D: 0.985). Pianka's index value was 0.207, indicating little overlap in the dietary composition of these bat species. Our results suggest that M. ikonnikovi has a wider diet composition than P. ognevi. Conclusions: Based on the dietary analysis results, our results suggests the possibility of differences in foraging site preferences or microhabitat utilization between two bat species cohabiting the Mt. Jumbong. In addition, these differences may represent one of the important mechanism in reducing interspecific competition and enabling coexistence between the two bat species. We expected that our results will be valuable for understanding resource partitioning and the coexistence of bats inhabiting the Korean forests.

• 한국산림과학회지
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• 제90권1호
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• pp.155-160
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• 2001

경기도 화성군 반월면 속달리 소재의 백합나무 종자 산지 시험림을 대상으로 모수림 작업 전후의 유전다양성의 변화를 예측하기 위해서 총 305개체에 대한 I-SSR 표지자 분석을 수행하였다. 9개의 I-SSR primer를 이용하여 PCR을 수행한 결과 총 89개의 증폭산물 변이 체를 확인했으며, 속달리 시험림에 식재되어 있는 전 개체에 대한 다양도는 0.4532로 비교적 높은 수치를 보였다. 종자산지별로 구분하여 분석한 결과 미국 채종원산 종자를 양묘하여 식재한 개체군에서 가장 높은 다양도(0.4530)를 보였으며, 그 다음이 안양 산지로 0.4152, 전북 산지의 경우 0.3929로 가장 낮은 다양도를 보였다. 식재목의 형태적 특성과 식재 간격을 고려하여 두 번에 걸친 간벌을 수반하는 모수림 작업 결과 남겨질 37개 개체목에 대한 유전다양도 및 유전적 거리를 모의 통계분석한 결과, 종자 산지내 다양도는 전북이 28.3%로 가장 크게 감소했으며, 그 다음이 안양으로 16.3%, 미국이 8.0%로 가장 적게 감소한 것으로 나타났다. 개체목 감소율의 차이가 적었음에도 불구하고(87.5~88.2%) 다양도의 변화에 큰 차이가 나는 이유는 미국산 채종원 종자는 비교적 다수의 모수로부터 유래되었으나 안양 및 전북에서 생산된 종자들은 상대적으로 소수의 모수로부터 유래되었을 가능성이 크기 때문인 것으로 추정된다. 비록 2차에 걸친 간벌에 의해서 각 종자 산지내 개체간 다양도는 평균 17.5%가 감소했으나, 전체 37개체에 대한 다양도의 감소는 불과 6.1%에 지나지 않기 때문에 속달리 시험림에 식재되어 있는 305개체 중에서 종자생산을 목적으로 본 연구에서 선정한 기준에 따라 37개체를 잔존시키는 모수림 작업을 수행하더라도 이들로부터 생산될 차대들에 있어서 전체적인 유전다양성의 현격한 감소를 초래하지는 않을 것으로 생각된다.

• 한국산림과학회지
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• 제89권2호
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• pp.167-172
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• 2000

국내 5개 지역에서 채집한 비자나무(Torreya nucifera Siev. et Zucc.) 95개체를 대상으로 I-SSR 표지자를 분석하였다. 총 62개의 I-SSR 증폭산물(增幅産物)이 관찰되었으며, 그 중 7개의 증폭산물(增幅産物)은 분석된 95개 개체에서 단형성(單形性)이었다. 관찰된 전체 I-SSR 증폭산물(增幅産物)을 통합(統合)하여 분석한 결과 개체목에 대한 DNA지방판별(指放判別)이 가능하였다. 대부분의 유전다양성(遺傳多樣性)이 임분(林分)내의 개체목 간에 존재하는 것으로 나타났고(90.65%), 전체 5개 임분(林分)에서 유사한 수준의 유전다양성(遺傳多樣性)을 보였다. 집단간의 유전적(遺傳的) 분화(分化)정도는 심하지 않았다(${\phi}_{ST}=9.35%$). UPGMA법에 의한 유집분석(類集分析) 결과 각 집단의 유전적(遺傳的) 유연관계(類緣關係)는 임분(林分)의 지리적(地理的) 분포양상(分布樣相)과 일치(一致)하지 않았으며, 각 교점(交點)의 형성(形成)에 있어서 통계적 유의성이 없었고 따라서 전체 집단들이 유전적(遺傳的)으로 크게 분화(分化)되지 않았음을 알 수 있었다.