Combining an innovative non-invasive sampling method and high-throughput sequencing to characterize fungal communities on a canvas painting

2019 ◽  
Vol 145 ◽  
pp. 104816 ◽  
Author(s):  
Hugo Paiva de Carvalho ◽  
Sílvia Oliveira Sequeira ◽  
Diogo Pinho ◽  
João Trovão ◽  
Ricardo Manuel Fernandes da Costa ◽  
...  
Keyword(s):  
High Throughput ◽  
Sampling Method ◽  
High Throughput Sequencing ◽  
Fungal Communities ◽  
Non Invasive

Characterizing fungal communities in medicinal and edible Cassiae Semen using high-throughput sequencing

2020 ◽  
Vol 319 ◽  
pp. 108496 ◽  
Author(s):  
Mengyue Guo ◽  
Wenjun Jiang ◽  
Meihua Yang ◽  
Xiaowen Dou ◽  
Xiaohui Pang
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Fungal Communities

scholarly journals Different Amplicon Targets for Sequencing-Based Studies of Fungal Diversity

2017 ◽  
Vol 83 (17) ◽  
Author(s):  
Francesca De Filippis ◽  
Manolo Laiola ◽  
Giuseppe Blaiotta ◽  
Danilo Ercolini
Keyword(s):  
Microbial Ecology ◽  
High Throughput ◽  
Biological Samples ◽  
Fungal Diversity ◽  
High Throughput Sequencing ◽  
Amplicon Sequencing ◽  
Fungal Species ◽  
Fungal Communities ◽  
Its2 Region ◽  
Mock Community

ABSTRACT Target-gene amplicon sequencing is the most exploited high-throughput sequencing application in microbial ecology. The targets are taxonomically relevant genes, with 16S rRNA being the gold standard for bacteria. As for fungi, the most commonly used target is the internal transcribed spacer (ITS). However, the uneven ITS length among species may promote preferential amplification and sequencing and incorrect estimation of their abundance. Therefore, the use of different targets is desirable. We evaluated the use of three different target amplicons for the characterization of fungal diversity. After an in silico primer evaluation, we compared three amplicons (the ITS1-ITS2 region [ITS1-2], 18S ribosomal small subunit RNA, and the D1/D2 domain of the 26S ribosomal large subunit RNA), using biological samples and a mock community of common fungal species. All three targets allowed for accurate identification of the species present. Nevertheless, high heterogeneity in ITS1-2 length was found, and this caused an overestimation of the abundance of species with a shorter ITS, while both 18S and 26S amplicons allowed for more reliable quantification. We demonstrated that ITS1-2 amplicon sequencing, although widely used, may lead to an incorrect evaluation of fungal communities, and efforts should be made to promote the use of different targets in sequencing-based microbial ecology studies. IMPORTANCE Amplicon-sequencing approaches for fungi may rely on different targets affecting the diversity and abundance of the fungal species. An increasing number of studies will address fungal diversity by high-throughput amplicon sequencing. The description of the communities must be accurate and reliable in order to draw useful insights and to address both ecological and biological questions. By analyzing a mock community and several biological samples, we demonstrate that using different amplicon targets may change the results of fungal microbiota analysis, and we highlight how a careful choice of the target is fundamental for a thorough description of the fungal communities.

scholarly journals Uncovering fungal community composition in natural habitat of Ophiocordyceps sinensis using high-throughput sequencing and culture-dependent approaches

2020 ◽  
Author(s):  
Chuanbo Zhang ◽  
Chao-Hui Ren ◽  
Yan-Li Wang ◽  
Qi-Qi Wang ◽  
Yun-Sheng Wang ◽  
...  
Keyword(s):  
Community Structure ◽  
High Throughput ◽  
Fungal Community ◽  
High Throughput Sequencing ◽  
Fungal Communities ◽  
Bioactive Metabolites ◽  
Fungal Community Structure ◽  
Ophiocordyceps Sinensis ◽  
Culturable Fungi ◽  
Culture Dependent

Abstract Background The fungal communities inhabiting natural Ophiocordyceps sinensis play critical ecological roles in alpine meadow ecosystem, contribute to infect host insect, influence the occurrence of O. sinensis, and are repertoire of potential novel metabolites discovery. However, a comprehensive understanding of fungal communities of O. sinensis remain elusive. Therefore, the present study aimed to unravel fungal communities of natural O. sinensis using combination of high-throughput sequencing and culture-dependent approach. Results A total of 280,519 high-quality sequences, belonging to 5 fungal phyla, 15 classes, 41 orders, 79 families, 112 genera, and 352 putative operational taxonomic units (OTUs) were obtained from natural O. sinensis using high-throughput sequencing. Among of which, 43 genera were identified in external mycelial cortices (EMC), Ophiocordyceps, Sebacinia, Archaeorhizomyces were predominant genera with the abundance of 95.86%, 1.14%, 0.85%, respectively. Total 66 genera were identified from soil microhabitat, Inocybe, Archaeorhizomyces, Unclassified Thelephoraceae, Tomentella, Thelephora, Sebacina, Unclassified Ascomycota, Unclassified Fungi were predominant genera with an average abundance of 53.32%, 8.69%, 8.12%, 8.12%, 7.21%, 4.6%, 3.08% and 3.05%, respectively. The fungal communities in external mycelial cortices (EMC) were significantly distinct from the soil microhabitat (Soil). Meanwhile, seven culture media that benefit for the growth of O. sinensis were used to isolate culturable fungi at 16 °C, resulted in 77 fungal strains isolated for rDNA ITS sequence analysis, belonging to 33 genera, including Ophiocordyceps, Trichoderma, Cytospora, Truncatella, Dactylonectria, Isaria, Cephalosporium, Fusarium, Cosmospora, Paecilomyces, etc.. Among all culturable fungi, Mortierella and Trichoderma were predominant genera of total isolates. Conclusions The significantly distinction and overlap in fungal community structure between two approaches highlight that integration of approaches would generate more information than either of them. Our finding is the first investigation of fungal community structure of natural O. sinensis by two approachs, provide new insight into O. sinensis associated fungi, and support that microbiota of O. sinensis is an untapped source for novel bioactive metabolites discovery.

Non-invasive prenatal DNA screening using high-throughput sequencing in pregnant women with recurrent miscarriage

2018 ◽  
Vol 8_2018 ◽  
pp. 48-55
Author(s):  
Sukhikh G.T. Sukhikh ◽  
Tetruashvili N.K. Tetruashvili ◽  
Kim L.V. Kim ◽  
Trofimov D.Yu. Trofimov ◽  
Barkov I.Yu. Barkov I ◽  
...  
Keyword(s):  
Pregnant Women ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Recurrent Miscarriage ◽  
Non Invasive

scholarly journals Antigenic and genetic characterization of Streptococcus pneumoniae strains isolated from patients with invasive and non-invasive pneumococcal infections by using high-throughput sequencing

2021 ◽  
Vol 98 (5) ◽  
pp. 512-518
Author(s):  
K. O. Mironov ◽  
I. I. Gaponova ◽  
V. I. Korchagin ◽  
Yu. V. Mikhailova ◽  
A. A. Shelenkov ◽  
...  
Keyword(s):  
Data Analysis ◽  
Streptococcus Pneumoniae ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Whole Genome ◽  
Pneumococcal Infections ◽  
Genome Sequences ◽  
Illumina Platform ◽  
Genetic Characteristics ◽  
Non Invasive

The objective of this study was to characterize and compare antigenic and genetic characteristics of Streptococcus pneumoniae strains isolated from patients with invasive and non-invasive pneumococcal infections (PIs) by using the data of high-throughput sequencing.Materials and methods. A total of 158 S. pneumoniae strains were studied. All of them were isolated during different stages of the PEHASus multicenter study performed in 2015-2020. The data analysis was based on the information about whole-genome sequences of 46 strains isolated during the above study. Real-time PCR methods and high-throughput sequencing (the Illumina platform) were used for identification of serotypes. The SeroBA, PneumoCaT software and PubMLST.org website resources were used in the data processing.Results and discussion. The serotypes of all the studied strains were identified. A number of discrepancies among serotypes in serogroup 6 and one discordant result were revealed by the analysis of whole-genome sequences using 2 programs. The PCR methods were effectively used to characterize serotypes in 87% and 69% of the pathogens of invasive and non-invasive PIs, respectively. The serotypes contained in PCV13 accounted for 59% and 37%, while PPV23 serotypes accounted for 78% and 53% of the strains isolated from patients with invasive and non-invasive PIs, respectively. The data analysis was unable to identify either the dominant sequence type (a total of 81 sequence types have been identified) or clonal complexes, except for serotype 3 strains, thus demonstrating consistency with the data from previous studies suggesting the absence of a well-represented clonal structure of S. pneumoniae associated with pneumococcal meningitis in Russia.Conclusion. The obtained data made it possible to identify the distribution of the circulating serotypes and genetic characteristics of the strains isolated from PI patients, thus being instrumental for assessment of the effectiveness of the existing polyvalent vaccines and providing information for improvement of the PCR-based methods of serotyping.

scholarly journals Uncovering fungal community composition in natural habitat of Ophiocordyceps sinensis using high-throughput sequencing and culture-dependent approaches

2020 ◽  
Author(s):  
Chuanbo Zhang ◽  
Chao-Hui Ren ◽  
Yan-Li Wang ◽  
Qi-Qi Wang ◽  
Yun-Sheng Wang ◽  
...  
Keyword(s):  
Community Structure ◽  
High Throughput ◽  
Fungal Community ◽  
High Throughput Sequencing ◽  
Fungal Communities ◽  
Comprehensive Understanding ◽  
Fungal Community Structure ◽  
Ophiocordyceps Sinensis ◽  
Culturable Fungi ◽  
Culture Dependent

Abstract Background: The fungal communities inhabiting natural Ophiocordyceps sinensis play critical ecological roles in alpine meadow ecosystem, contribute to infect host insect, influence the occurrence of O. sinensis, and are repertoire of potential novel metabolites discovery. However, a comprehensive understanding of fungal communities of O. sinensis remain elusive. Therefore, the present study aimed to unravel fungal communities of natural O. sinensis using combination of high-throughput sequencing and culture-dependent approaches. Results: A total of 280,519 high-quality sequences, belonging to 5 fungal phyla, 15 classes, 41 orders, 79 families, 112 genera, and 352 putative operational taxonomic units (OTUs) were obtained from natural O. sinensis using high-throughput sequencing. Among of which, 43 genera were identified in external mycelial cortices, Ophiocordyceps, Sebacinia, Archaeorhizomyces were predominant genera with the abundance of 95.86%, 1.14%, 0.85%, respectively. A total of 66 genera were identified from soil microhabitat, Inocybe, Archaeorhizomyces, unclassified Thelephoraceae, Tomentella, Thelephora, Sebacina, unclassified Ascomycota, unclassified Fungi were predominant genera with an average abundance of 53.32%, 8.69%, 8.12%, 8.12%, 7.21%, 4.6%, 3.08% and 3.05%, respectively. The fungal communities in external mycelial cortices were significantly distinct from the soil microhabitat. Meanwhile, seven types of culture media were used to isolate culturable fungi at 16°C, resulted in 77 fungal strains isolated by rDNA ITS sequence analysis, belonging to 33 genera, including Ophiocordyceps, Trichoderma, Cytospora, Truncatella, Dactylonectria, Isaria, Cephalosporium, Fusarium, Cosmospora and Paecilomyces, etc.. Among all culturable fungi, Mortierella and Trichoderma were predominant genera. Conclusions: The significantly differences and overlap in fungal community structure between two approaches highlight that the integration of high-throughput sequencing and culture-dependent approaches would generate more information. Our result reveal a comprehensive understanding of fungal community structure of natural O. sinensis, provide new insight into O. sinensis associated fungi, and support that microbiota of natural O. sinensis is an untapped source for novel bioactive metabolites discovery.

scholarly journals Uncovering fungal community composition in natural habitat of Ophiocordyceps sinensis using high-throughput sequencing and culture-dependent approaches

2020 ◽  
Author(s):  
Chuanbo Zhang ◽  
Chao-Hui Ren ◽  
Yan-Li Wang ◽  
Qi-Qi Wang ◽  
Yun-Sheng Wang ◽  
...  
Keyword(s):  
Community Structure ◽  
High Throughput ◽  
Fungal Community ◽  
High Throughput Sequencing ◽  
Fungal Communities ◽  
Comprehensive Understanding ◽  
Fungal Community Structure ◽  
Ophiocordyceps Sinensis ◽  
Culturable Fungi ◽  
Culture Dependent

Abstract Background: The fungal communities inhabiting natural Ophiocordyceps sinensis play critical ecological roles in alpine meadow ecosystem, contribute to infect host insect, influence the occurrence of O. sinensis, and are repertoire of potential novel metabolites discovery. However, a comprehensive understanding of fungal communities of O. sinensis remain elusive. Therefore, the present study aimed to unravel fungal communities of natural O. sinensis using combination of high-throughput sequencing and culture-dependent approaches. Results: A total of 280,519 high-quality sequences, belonging to 5 fungal phyla, 15 classes, 41 orders, 79 families, 112 genera, and 352 putative operational taxonomic units (OTUs) were obtained from natural O. sinensis using high-throughput sequencing. Among of which, 43 genera were identified in external mycelial cortices (EMC), Ophiocordyceps, Sebacinia, Archaeorhizomyces were predominant genera with the abundance of 95.86%, 1.14%, 0.85%, respectively. A total of 66 genera were identified from soil microhabitat (Soil), Inocybe, Archaeorhizomyces, unclassified Thelephoraceae, Tomentella, Thelephora, Sebacina, unclassified Ascomycota, unclassified Fungi were predominant genera with an average abundance of 53.32%, 8.69%, 8.12%, 8.12%, 7.21%, 4.6%, 3.08% and 3.05%, respectively. The fungal communities in external mycelial cortices were significantly distinct from the soil microhabitat. Meanwhile, seven types of culture media were used to isolate culturable fungi at 16°C, resulted in 77 fungal strains isolated by rDNA ITS sequence analysis, belonging to 33 genera, including Ophiocordyceps, Trichoderma, Cytospora, Truncatella, Dactylonectria, Isaria, Cephalosporium, Fusarium, Cosmospora and Paecilomyces, etc.. Among all culturable fungi, Mortierella and Trichoderma were predominant genera. Conclusions: The significantly differences and overlap in fungal community structure between two approaches highlight that the integration of high-throughput sequencing and culture-dependent approaches would generate more information. Our result reveal a comprehensive understanding of fungal community structure of natural O. sinensis, provide new insight into O. sinensis associated fungi, and support that microbiota of natural O. sinensis is an untapped source for novel bioactive metabolites discovery.

Fungal communities in Nelumbinis semen characterized by high-throughput sequencing

2021 ◽  
pp. 109428
Author(s):  
Nan Long ◽  
Jinxin Liu ◽  
Xiaofang Liao ◽  
Boyu Jia ◽  
Jiali Liu ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Fungal Communities ◽  
Nelumbinis Semen

scholarly journals Multi-element fingerprinting and high throughput sequencing identify multiple elements that affect fungal communities inQuercus macrocarpafoliage

2010 ◽  
Vol 5 (9) ◽  
pp. 1157-1161 ◽  
Author(s):  
Ari Jumpponen ◽  
Karen Keating ◽  
Gary Gadbury ◽  
Kenneth L. Jones ◽  
J. David Mattox
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Fungal Communities

scholarly journals Improved Selection of Internal Transcribed Spacer-Specific Primers Enables Quantitative, Ultra-High-Throughput Profiling of Fungal Communities

2013 ◽  
Vol 79 (8) ◽  
pp. 2519-2526 ◽  
Author(s):  
Nicholas A. Bokulich ◽  
David A. Mills
Keyword(s):  
High Throughput ◽  
Internal Transcribed Spacer ◽  
High Throughput Sequencing ◽  
Work Flow ◽  
Fungal Communities ◽  
Amplification Bias ◽  
Sequencing Technologies ◽  
Yeast Community ◽  
Primer Sets ◽  
Sequencing Platforms

ABSTRACTUltra-high-throughput sequencing (HTS) of fungal communities has been restricted by short read lengths and primer amplification bias, slowing the adoption of newer sequencing technologies to fungal community profiling. To address these issues, we evaluated the performance of several common internal transcribed spacer (ITS) primers and designed a novel primer set and work flow for simultaneous quantification and species-level interrogation of fungal consortia. Primer comparison and validation were predictedin silicoand by sequencing a “mock community” of mixed yeast species to explore the challenges of amplicon length and amplification bias for reconstructing defined yeast community structures. The amplicon size and distribution of this primer set are smaller than for all preexisting ITS primer sets, maximizing sequencing coverage of hypervariable ITS domains by very-short-amplicon, high-throughput sequencing platforms. This feature also enables the optional integration of quantitative PCR (qPCR) directly into the HTS preparatory work flow by substituting qPCR with these primers for standard PCR, yielding quantification of individual community members. The complete work flow described here, utilizing any of the qualified primer sets evaluated, can rapidly profile mixed fungal communities and capably reconstructed well-characterized beer and wine fermentation fungal communities.

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