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Impact of Endosymbionts on Antimicrobial Properties of Medicinal Plants
Published in Mahendra Rai, Chistiane M. Feitosa, Eco-Friendly Biobased Products Used in Microbial Diseases, 2022
Flávia Figueira Aburjaile, José Ribamar Costa Ferreira-Neto, Thamara de Medeiros Azevedo, Juan Carlos Ariute, Jéssica Barboza da Silva, Roberta Lane de Oliveira Silva, Valesca Pandolfi, Ana Maria Benko-Iseppon
High-throughput shotgun metagenomic sequencing has been estimated as a complementary approach to amplicon sequencing due to its ability to provide a deeper understanding of the structure and function of the microbiome associated with medicinal plants (Hao et al. 2018). As a result, publications in recent years have adopted HTS technology followed by bioinformatics analysis as an independent and successful approach to characterize and elucidate the diversity and composition of endophytic bacterial and fungal communities of medicinal plants (Li et al. 2017; Lu et al. 2020; Chen et al. 2020). Some recent publications have also adopted a combined methodology between culture-dependent methods or molecular approaches with metagenomics or metabarcoding approaches, to integrate and compare information provided in different ways. This is the case of studies on the endophytic microbiomes of medicinal plants Passiflora incarnata(Goulart et al. 2019), Astragalus membranaceus, A. mongholicus (Sun et al. 2017), and Paris polyphylla(Liu et al. 2016) (Table 14.1).
Liquid Biopsies for Pancreatic Cancer: A Step Towards Early Detection
Published in Surinder K. Batra, Moorthy P. Ponnusamy, Gene Regulation and Therapeutics for Cancer, 2021
Joseph Carmicheal, Rahat Jahan, Koelina Ganguly, Ashu Shah, Sukhwinder Kaur
Other targeted approaches are being developed based on Deep sequencing/Next Generation Sequencing (NGS) using site-specific primers to amplify a particular genomic region. Some examples of these regions include “Safe-SeqS”, Tagged Amplicon Sequencing (TAm-Seq), AmpliSeq and Personalized Analysis of Rearranged Ends (PARE). Though they require further standardization and experimental study, these are some of the advanced technologies that will prove invaluable in the identification of specific hotspot mutations found in ctDNA [15, 16].
Applications of Circulating DNA Analysis in Personalized Medicine
Published in II-Jin Kim, Cancer Genetics and Genomics for Personalized Medicine, 2017
Dana W.Y. Tsui, Muhammed Murtaza
Greater depth of coverage (~1000s-fold) can be achieved by focusing on a few genes of interest using custom enrichment by hybridization (Newman et al., 2014) or multiplexed PCR-based amplicon sequencing (Forshew et al., 2012). Both methods are accurate for identification of mutations with >1–2% allele fraction and quantification of known mutations with >0.1% allele fraction. Enrichment by hybridization allows several megabases of the genomic coverage compared with several kilobases of coverage achievable with current approaches for amplicon sequencing. However, ligation-based library preparation methods are required for enrichment by hybridization and these have limited efficiency with <10 nanograms of input DNA and can suffer from allelic dropout. In contrast, amplicon-based methods such as tagged amplicon deep dequencing (TAm-Seq) have been shown to prepare sequencing libraries from as little as 1–2 genome equivalents of input DNA (Forshew et al., 2012).
Deoxycholic acid induces gastric intestinal metaplasia by activating STAT3 signaling and disturbing gastric bile acids metabolism and microbiota
Published in Gut Microbes, 2022
Duochen Jin, Keting Huang, Miao Xu, Hongjin Hua, Feng Ye, Jin Yan, Guoxin Zhang, Yun Wang
Statistical analyses were performed by using IBM SPSS Statistics 23.0 and GraphPad Prism 8.0 software. Student’s t test or the chi-square test was applied to determine significant differences between two groups, while ANOVA with Tukey’s test was used to analyze significant differences among multiple groups. Pearson’s test was performed to evaluate correlations between two groups. The cell experiments were performed in at least triplicate, and normally distributed data are presented as the mean ± SEM of at least three independent experiments. A 2-sided P < .05 was considered significant. The amplicon sequencing statistical analyses were performed with R software. Microbial α-diversity indices were calculated using the Wilcoxon test, while microbial β-diversity was compared between two groups using the ANOSIM nonparametric test.
Faster infection diagnostics for intensive care unit (ICU) patients
Published in Expert Review of Molecular Diagnostics, 2022
Almudena Burillo, Emilio Bouza
Targeted amplicon sequencing involves amplification and sequencing of one or more genes (e.g. 16S ribosomal RNA gene or 18S ribosomal RNA gene in fungi) directly on clinical specimens. In work by Salipante et al., in a collection of sputum from patients with cystic fibrosis and brain abscesses, this technique demonstrated its usefulness in the diagnosis of polymicrobial infections in complex biological samples almost ten years ago [38]. This is an expensive technique, but it also provides all the information on the microorganism, so its applicability reside in situations in which other faster or cheaper techniques will not provide the necessary information (e.g. a rare resistance mechanism, a mutation, the study of an outbreak, etc.), and also when a microorganism does not grow or takes a long time to grow, as in the case of mycobacteria. The price at present is around 70–100 euros per sample.
The rectal mucosal but not fecal microbiota detects subclinical ulcerative colitis
Published in Gut Microbes, 2021
Yu-Fei Lin, Chang Mu Sung, Huei-Mien Ke, Chia-Jung Kuo, Wei-an Liu, Wen-Sy Tsai, Cheng-Yu Lin, Hao-Tsai Cheng, Meiyeh J Lu, Isheng. J. Tsai, Sen-Yung Hsieh
Amplicon sequencing libraries were prepared as previously described.37 Amplicons were visualized by running 2 µl of the product on 2.0% (w/v) agarose gel to confirm that a product was generated. Sample normalization was performed using SequalPrep Normalization Plate Kit, 96-well (Cat #: A1051001, ThermoFisher, Waltham, Massachusetts, U.S.). Normalized amplicon products were pooled at equal volumes. The pooled DNA library was concentrated using an equal volume of Agencourt AMPure XP beads (Cat #: A63880, Beckman Coulter, Pasadena, California, U.S.). Sequencing was performed by the NGS High Throughput Genomics Core in Biodiversity Research Center, Academia Sinica, Taiwan. Sequencing of the 16S amplicon was carried out using Illumina MiSeq with paired-end 2 × 250 bp chemistry.