Publications by authors named "Christopher J Fields"

39 Publications

A non-canonical microRNA derived from the snaR-A non-coding RNA targets a metastasis inhibitor.

RNA 2021 Apr 1. Epub 2021 Apr 1.

University of Florida

MicroRNAs (miRNAs) are small noncoding RNAs that function as critical post-transcriptional regulators in various biological processes. While most miRNAs are generated from processing of long primary transcripts via sequential Drosha and Dicer cleavage, some miRNAs that bypass Drosha cleavage can be transcribed as part of another small non-coding RNA. Here, we develop the Target-Oriented miRNA Discovery (TOMiD) bioinformatic analysis method to identify Drosha-independent miRNAs from Argonaute crosslinking and sequencing of hybrids (Ago-CLASH) datasets. Using this technique, we discovered a novel miRNA derived from a primate specific non-coding RNA, the small NF90 associated RNA A (snaR-A). The miRNA derived from snaR-A (miR-snaR) arises independent of Drosha processing, but requires Exportin-5 and Dicer for biogenesis. We identify that miR-snaR is concurrently upregulated with the full snaR-A transcript in cancer cells. Functionally, miR-snaR associates with Ago proteins and targets NME1, a key metastasis inhibitor, contributing to snaR-A's role in promoting cancer cell migration. Our findings suggest a functional link between a novel miRNA and its precursor non-coding RNA.
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http://dx.doi.org/10.1261/rna.078694.121DOI Listing
April 2021

Competitions between Fibrobacter succinogenes, Ruminococcus flavefaciens, and Ruminoccus albus in a Gnotobiotic Sheep Model Revealed by Multi-Omic Analyses.

mBio 2021 03 3;12(2). Epub 2021 Mar 3.

Université Clermont Auvergne, INRAE, UMR 454 MEDIS, Clermont-Ferrand, France

, , and are the three predominant cellulolytic bacterial species found in the rumen. studies have shown that these species compete for adherence to, and growth upon, cellulosic biomass. Yet their molecular interactions have not heretofore been examined. Gnotobiotically raised lambs harboring a 17-h-old immature microbiota devoid of culturable cellulolytic bacteria and methanogens were inoculated first with S85 and sp. strain 87.7, and 5 months later, the lambs were inoculated with 8 and FD-1. Longitudinal samples were collected and profiled for population dynamics, gene expression, fibrolytic enzyme activity, fibrolysis, and metabolite profiling. Quantitative PCR, metagenome and metatranscriptome data show that establishes at high levels initially but is gradually outcompeted following the introduction of the ruminococci. This shift resulted in an increase in carboxymethyl cellulase (CMCase) and xylanase activities but not in greater fibrolysis, suggesting that and ruminococci deploy different but equally effective means to degrade plant cell walls. Expression profiles showed that relied upon outer membrane vesicles and a diverse repertoire of CAZymes, while and preferred type IV pili and either CBM37-harboring or cellulosomal carbohydrate-active enzymes (CAZymes), respectively. The changes in cellulolytics also affected the rumen metabolome, including an increase in acetate and butyrate at the expense of propionate. In conclusion, this study provides the first demonstration of competition between the three predominant cellulolytic bacteria and provides insight on the influence of these ecological interactions on rumen fibrolytic function and metabolomic response. Ruminant animals, including cattle and sheep, depend on their rumen microbiota to digest plant biomass and convert it into absorbable energy. Considering that the extent of meat and milk production depends on the efficiency of the microbiota to deconstruct plant cell walls, the functionality of predominant rumen cellulolytic bacteria, , , and , has been extensively studied to obtain a better knowledge of how they operate to hydrolyze polysaccharides and ultimately find ways to enhance animal production. This study provides the first evidence of competitions between and the two species. It shows that a simple disequilibrium within the cellulolytic community has repercussions on the rumen metabolome and fermentation end products. This finding will have to be considered in the future when determining strategies aiming at directing rumen fermentations for animal production.
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http://dx.doi.org/10.1128/mBio.03533-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092306PMC
March 2021

Using a multiple-delivery-mode training approach to develop local capacity and infrastructure for advanced bioinformatics in Africa.

PLoS Comput Biol 2021 Feb 25;17(2):e1008640. Epub 2021 Feb 25.

Computational Biology Division, Department of Integrative Biomedical Sciences, IDM, CIDRI Africa Wellcome Trust Centre, University of Cape Town, Cape Town, South Africa.

With more microbiome studies being conducted by African-based research groups, there is an increasing demand for knowledge and skills in the design and analysis of microbiome studies and data. However, high-quality bioinformatics courses are often impeded by differences in computational environments, complicated software stacks, numerous dependencies, and versions of bioinformatics tools along with a lack of local computational infrastructure and expertise. To address this, H3ABioNet developed a 16S rRNA Microbiome Intermediate Bioinformatics Training course, extending its remote classroom model. The course was developed alongside experienced microbiome researchers, bioinformaticians, and systems administrators, who identified key topics to address. Development of containerised workflows has previously been undertaken by H3ABioNet, and Singularity containers were used here to enable the deployment of a standard replicable software stack across different hosting sites. The pilot ran successfully in 2019 across 23 sites registered in 11 African countries, with more than 200 participants formally enrolled and 106 volunteer staff for onsite support. The pulling, running, and testing of the containers, software, and analyses on various clusters were performed prior to the start of the course by hosting classrooms. The containers allowed the replication of analyses and results across all participating classrooms running a cluster and remained available posttraining ensuring analyses could be repeated on real data. Participants thus received the opportunity to analyse their own data, while local staff were trained and supported by experienced experts, increasing local capacity for ongoing research support. This provides a model for delivering topic-specific bioinformatics courses across Africa and other remote/low-resourced regions which overcomes barriers such as inadequate infrastructures, geographical distance, and access to expertise and educational materials.
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http://dx.doi.org/10.1371/journal.pcbi.1008640DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7906323PMC
February 2021

The 21st annual Bioinformatics Open Source Conference (BOSC 2020, part of BCC2020).

F1000Res 2020 21;9. Epub 2020 Sep 21.

Wellcome Trust, London, NW1 2BE, UK.

Launched in 2000 and held every year since, the Bioinformatics Open Source Conference (BOSC) is a volunteer-run meeting coordinated by the Open Bioinformatics Foundation (OBF) that covers open source software development and open science in bioinformatics. Most years, BOSC has been part of the Intelligent Systems for Molecular Biology (ISMB) conference, but in 2018, and again in 2020, BOSC partnered with the Galaxy Community Conference (GCC). This year's combined BOSC + GCC conference was called the Bioinformatics Community Conference (BCC2020, bcc2020.github.io). Originally slated to take place in Toronto, Canada, BCC2020 was moved online due to COVID-19. The meeting started with a wide array of training sessions; continued with a main program of keynote presentations, talks, posters, Birds of a Feather, and more; and ended with four days of collaboration (CoFest). Efforts to make the meeting accessible and inclusive included very low registration fees, talks presented twice a day, and closed captioning for all videos. More than 800 people from 61 countries registered for at least one part of the meeting, which was held mostly in the Remo.co video-conferencing platform.
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http://dx.doi.org/10.12688/f1000research.26498.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7506190PMC
October 2020

DNA punch cards for storing data on native DNA sequences via enzymatic nicking.

Nat Commun 2020 04 8;11(1):1742. Epub 2020 Apr 8.

Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA.

Synthetic DNA-based data storage systems have received significant attention due to the promise of ultrahigh storage density and long-term stability. However, all known platforms suffer from high cost, read-write latency and error-rates that render them noncompetitive with modern storage devices. One means to avoid the above problems is using readily available native DNA. As the sequence content of native DNA is fixed, one can modify the topology instead to encode information. Here, we introduce DNA punch cards, a macromolecular storage mechanism in which data is written in the form of nicks at predetermined positions on the backbone of native double-stranded DNA. The platform accommodates parallel nicking on orthogonal DNA fragments and enzymatic toehold creation that enables single-bit random-access and in-memory computations. We use Pyrococcus furiosus Argonaute to punch files into the PCR products of Escherichia coli genomic DNA and accurately reconstruct the encoded data through high-throughput sequencing and read alignment.
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http://dx.doi.org/10.1038/s41467-020-15588-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7142088PMC
April 2020

A Staphylococcus pro-apoptotic peptide induces acute exacerbation of pulmonary fibrosis.

Nat Commun 2020 03 24;11(1):1539. Epub 2020 Mar 24.

Research Institute of Green Science and Technology, Graduate School of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal disease of unknown etiology; however, apoptosis of lung alveolar epithelial cells plays a role in disease progression. This intractable disease is associated with increased abundance of Staphylococcus and Streptococcus in the lungs, yet their roles in disease pathogenesis remain elusive. Here, we report that Staphylococcus nepalensis releases corisin, a peptide conserved in diverse staphylococci, to induce apoptosis of lung epithelial cells. The disease in mice exhibits acute exacerbation after intrapulmonary instillation of corisin or after lung infection with corisin-harboring S. nepalensis compared to untreated mice or mice infected with bacteria lacking corisin. Correspondingly, the lung corisin levels are significantly increased in human IPF patients with acute exacerbation compared to patients without disease exacerbation. Our results suggest that bacteria shedding corisin are involved in acute exacerbation of IPF, yielding insights to the molecular basis for the elevation of staphylococci in pulmonary fibrosis.
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http://dx.doi.org/10.1038/s41467-020-15344-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7093394PMC
March 2020

Soybean aphid biotype 1 genome: Insights into the invasive biology and adaptive evolution of a major agricultural pest.

Insect Biochem Mol Biol 2020 05 25;120:103334. Epub 2020 Feb 25.

USDA-ARS and Department of Crop Sciences, University of Illinois, Urbana, IL, USA.

The soybean aphid, Aphis glycines Matsumura (Hemiptera: Aphididae) is a serious pest of the soybean plant, Glycine max, a major world-wide agricultural crop. We assembled a de novo genome sequence of Ap. glycines Biotype 1, from a culture established shortly after this species invaded North America. 20.4% of the Ap. glycines proteome is duplicated. These in-paralogs are enriched with Gene Ontology (GO) categories mostly related to apoptosis, a possible adaptation to plant chemistry and other environmental stressors. Approximately one-third of these genes show parallel duplication in other aphids. But Ap. gossypii, its closest related species, has the lowest number of these duplicated genes. An Illumina GoldenGate assay of 2380 SNPs was used to determine the world-wide population structure of Ap. Glycines. China and South Korean aphids are the closest to those in North America. China is the likely origin of other Asian aphid populations. The most distantly related aphids to those in North America are from Australia. The diversity of Ap. glycines in North America has decreased over time since its arrival. The genetic diversity of Ap. glycines North American population sampled shortly after its first detection in 2001 up to 2012 does not appear to correlate with geography. However, aphids collected on soybean Rag experimental varieties in Minnesota (MN), Iowa (IA), and Wisconsin (WI), closer to high density Rhamnus cathartica stands, appear to have higher capacity to colonize resistant soybean plants than aphids sampled in Ohio (OH), North Dakota (ND), and South Dakota (SD). Samples from the former states have SNP alleles with high F values and frequencies, that overlap with genes involved in iron metabolism, a crucial metabolic pathway that may be affected by the Rag-associated soybean plant response. The Ap. glycines Biotype 1 genome will provide needed information for future analyses of mechanisms of aphid virulence and pesticide resistance as well as facilitate comparative analyses between aphids with differing natural history and host plant range.
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http://dx.doi.org/10.1016/j.ibmb.2020.103334DOI Listing
May 2020

BOSC 2019, the 20th annual Bioinformatics Open Source Conference.

F1000Res 2019 20;8. Epub 2019 Dec 20.

Department of Genetics, University of Cambridge, Cambridge, CB2 3EH, UK.

The Bioinformatics Open Source Conference is a volunteer-organized meeting that covers open source software development and open science in bioinformatics. Launched in 2000, BOSC has been held every year since. BOSC 2019, the 20th annual BOSC, took place as one of the Communities of Special Interest (COSIs) at the Intelligent Systems for Molecular Biology meeting (ISMB/ECCB 2019). The two-day meeting included a total of 46 talks and 55 posters, as well as eight Birds of a Feather interest groups. The keynote speaker was University of Cape Town professor Dr. Nicola Mulder, who spoke on "Building infrastructure for responsible open science in Africa". Immediately after BOSC 2019, about 50 people participated in the two-day CollaborationFest (CoFest for short), an open and free community-driven event at which participants work together to contribute to bioinformatics software, documentation, training materials, and use cases.
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http://dx.doi.org/10.12688/f1000research.21568.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6971845PMC
May 2020

The ' lifestyle' of bile acid 7α-dehydroxylating bacteria: comparative genomics, metatranscriptomic, and bile acid metabolomics analysis of a defined microbial community in gnotobiotic mice.

Gut Microbes 2020 05 9;11(3):381-404. Epub 2019 Jun 9.

Department of Animal Sciences, University of Illinois at Urbana-Champaign , Urbana, IL, USA.

The formation of secondary bile acids by gut microbes is a current topic of considerable biomedical interest. However, a detailed understanding of the biology of anaerobic bacteria in the genus that are capable of generating secondary bile acids is lacking. We therefore sought to determine the transcriptional responses of two prominent secondary bile acid producing bacteria, and to bile salts () and the cecal environment of gnotobiotic mice. The genomes of DSM 15053 and DSM 13275 were closed, and found to encode 3,647 genes (3,584 protein-coding) and 2,363 predicted genes (of which 2,239 are protein-coding), respectively, and 1,035 orthologs were shared between and . RNA-Seq analysis was performed in growth medium alone, and in the presence of cholic acid (CA) and deoxycholic acid (DCA). Growth with CA resulted in differential expression (>0.58 logFC; FDR < 0.05) of 197 genes in and 118 genes in . The bile acid-inducible operons () from each organism were highly upregulated in the presence of CA but not DCA. We then colonized germ-free mice with human gut bacterial isolates capable of metabolizing taurine-conjugated bile acids. This consortium included bile salt hydrolase-expressing ATCC 8492, ATCC 8482, DSM 20701, as well as taurine-respiring DSM 11045, and deoxycholic/lithocholic acid generating DSM 15053 and DSM 13275. Butyrate and iso-bile acid-forming ATCC 27340 was also included. The Bacteroidetes made up 84.71% of 16S rDNA cecal reads, , constituted 14.7%, and the clostridia made up <.75% of 16S rDNA cecal reads. Bile acid metabolomics of the cecum, serum, and liver indicate that the synthetic community were capable of functional bile salt deconjugation, oxidation/isomerization, and 7α-dehydroxylation of bile acids. Cecal metatranscriptome analysis revealed expression of genes involved in metabolism of taurine-conjugated bile acids. The transcriptomes of and suggest fermentation of simple sugars and utilization of amino acids glycine and proline as electron acceptors. Genes predicted to be involved in trimethylamine (TMA) formation were also expressed.
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http://dx.doi.org/10.1080/19490976.2019.1618173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7524365PMC
May 2020

Host species and site of collection shape the microbiota of Rift Valley fever vectors in Kenya.

PLoS Negl Trop Dis 2019 06 7;13(6):e0007361. Epub 2019 Jun 7.

International Centre of Insect Physiology and Ecology (icipe), Nairobi, Kenya.

The composition and structure of microbial communities associated with mosquitoes remain poorly understood despite their important role in host biology and potential to be harnessed as novel strategies for mosquito-borne disease control. We employed MiSeq sequencing of the 16S rRNA gene amplicons to characterize the bacterial flora of field-collected populations of Aedes mcintoshi and Aedes ochraceus, the primary vectors of Rift Valley fever (RVF) virus in Kenya. Proteobacteria (53.5%), Firmicutes (22.0%) and Actinobacteria (10.0%) were the most abundant bacterial phyla accounting for 85.5% of the total sequences. Non-metric multi-dimensional scaling plots based on Bray-Curtis dissimilarities revealed a clear grouping of the samples by mosquito species, indicating that the two mosquito species harbored distinct microbial communities. Microbial diversity, richness and composition was strongly influenced by the site of mosquito collection and overall, Ae. ochraceus had significantly higher microbial diversity and richness than Ae. mcintoshi. Our findings suggest that host species and site of collection are important determinants of bacterial community composition and diversity in RVF virus vectors and these differences likely contribute to the spatio-temporal transmission dynamics of RVF virus.
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http://dx.doi.org/10.1371/journal.pntd.0007361DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6584011PMC
June 2019

Physiology, Metabolism, and Fossilization of Hot-Spring Filamentous Microbial Mats.

Astrobiology 2019 12 30;19(12):1442-1458. Epub 2019 Apr 30.

Carl R. Woese Institute for Genomic Biology, University of Illinois Urbana-Champaign, Urbana, Illinois, USA.

The evolutionarily ancient Aquificales bacterium spp. dominates filamentous microbial mat communities in shallow, fast-flowing, and dysoxic hot-spring drainage systems around the world. In the present study, field observations of these fettuccini-like microbial mats at Mammoth Hot Springs in Yellowstone National Park are integrated with geology, geochemistry, hydrology, microscopy, and multi-omic molecular biology analyses. Strategic sampling of living filamentous mats along with the hot-spring CaCO () in which they are actively being entombed and fossilized has permitted the first direct linkage of spp. physiology and metabolism with the formation of distinct travertine streamer microbial biomarkers. Results indicate that, during chemoautotrophy and CO carbon fixation, the 87-98% -dominated mats utilize chaperons to facilitate enzyme stability and function. High-abundance transcripts and proteins for type IV pili and extracellular polymeric substances (EPSs) are consistent with their strong mucus-rich filaments tens of centimeters long that withstand hydrodynamic shear as they become encrusted by more than 5 mm of travertine per day. Their primary energy source is the oxidation of reduced sulfur ( sulfide, sulfur, or thiosulfate) and the simultaneous uptake of extremely low concentrations of dissolved O facilitated by bd-type cytochromes. The formation of elevated travertine ridges permits the -dominated mats to create a shallow platform from which to access low levels of dissolved oxygen at the virtual exclusion of other microorganisms. These ridged travertine streamer microbial biomarkers are well preserved and create a robust fossil record of microbial physiological and metabolic activities in modern and ancient hot-spring ecosystems.
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http://dx.doi.org/10.1089/ast.2018.1965DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6918859PMC
December 2019

Exercise and the Gut Microbiome: A Review of the Evidence, Potential Mechanisms, and Implications for Human Health.

Exerc Sport Sci Rev 2019 04;47(2):75-85

Division of Nutritional Sciences, University of Illinois Urbana-Champaign, Champaign, IL.

The gastrointestinal tract contains trillions of microbes (collectively known as the gut microbiota) that play essential roles in host physiology and health. Studies from our group and others have demonstrated that exercise independently alters the composition and functional capacity of the gut microbiota. Here, we review what is known about the gut microbiota, how it is studied, and how it is influenced by exercise training and discuss the potential mechanisms and implications for human health and disease.
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http://dx.doi.org/10.1249/JES.0000000000000183DOI Listing
April 2019

Sequencing Framework for the Sensitive Detection and Precise Mapping of Defective Interfering Particle-Associated Deletions across Influenza A and B Viruses.

J Virol 2019 06 15;93(11). Epub 2019 May 15.

Department of Microbiology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA

The mechanisms and consequences of defective interfering particle (DIP) formation during influenza virus infection remain poorly understood. The development of next-generation sequencing (NGS) technologies has made it possible to identify large numbers of DIP-associated sequences, providing a powerful tool to better understand their biological relevance. However, NGS approaches pose numerous technical challenges, including the precise identification and mapping of deletion junctions in the presence of frequent mutation and base-calling errors, and the potential for numerous experimental and computational artifacts. Here, we detail an Illumina-based sequencing framework and bioinformatics pipeline capable of generating highly accurate and reproducible profiles of DIP-associated junction sequences. We use a combination of simulated and experimental control data sets to optimize pipeline performance and demonstrate the absence of significant artifacts. Finally, we use this optimized pipeline to reveal how the patterns of DIP-associated junction formation differ between different strains and subtypes of influenza A and B viruses and to demonstrate how these data can provide insight into mechanisms of DIP formation. Overall, this work provides a detailed roadmap for high-resolution profiling and analysis of DIP-associated sequences within influenza virus populations. Influenza virus defective interfering particles (DIPs) that harbor internal deletions within their genomes occur naturally during infection in humans and during cell culture. They have been hypothesized to influence the pathogenicity of the virus; however, their specific function remains elusive. The accurate detection of DIP-associated deletion junctions is crucial for understanding DIP biology but is complicated by an array of technical issues that can bias or confound results. Here, we demonstrate a combined experimental and computational framework for detecting DIP-associated deletion junctions using next-generation sequencing (NGS). We detail how to validate pipeline performance and provide the bioinformatics pipeline for groups interested in using it. Using this optimized pipeline, we detect hundreds of distinct deletion junctions generated during infection with a diverse panel of influenza viruses and use these data to test a long-standing hypothesis concerning the molecular details of DIP formation.
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http://dx.doi.org/10.1128/JVI.00354-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6532088PMC
June 2019

Clostridium scindens ATCC 35704: Integration of Nutritional Requirements, the Complete Genome Sequence, and Global Transcriptional Responses to Bile Acids.

Appl Environ Microbiol 2019 04 22;85(7). Epub 2019 Mar 22.

Microbiome Metabolic Engineering Theme, Carl R. Woese Institute for Genomic Biology, Urbana, Illinois, USA

In the human gut, ATCC 35704 is a predominant bacterium and one of the major bile acid 7α-dehydroxylating anaerobes. While this organism is well-studied relative to bile acid metabolism, little is known about the basic nutrition and physiology of ATCC 35704. To determine the amino acid and vitamin requirements of , the leave-one-out (one amino acid group or vitamin) technique was used to eliminate the nonessential amino acids and vitamins. With this approach, the amino acid tryptophan and three vitamins (riboflavin, pantothenate, and pyridoxal) were found to be required for the growth of In the newly developed defined medium, fermented glucose mainly to ethanol, acetate, formate, and H The genome of ATCC 35704 was completed through PacBio sequencing. Pathway analysis of the genome sequence coupled with transcriptome sequencing (RNA-Seq) under defined culture conditions revealed consistency with the growth requirements and end products of glucose metabolism. Induction with bile acids revealed complex and differential responses to cholic acid and deoxycholic acid, including the expression of potentially novel bile acid-inducible genes involved in cholic acid metabolism. Responses to toxic deoxycholic acid included expression of genes predicted to be involved in DNA repair, oxidative stress, cell wall maintenance/metabolism, chaperone synthesis, and downregulation of one-third of the genome. These analyses provide valuable insight into the overall biology of which may be important in treatment of disease associated with increased colonic secondary bile acids. is one of a few identified gut bacterial species capable of converting host cholic acid into disease-associated secondary bile acids such as deoxycholic acid. The current work represents an important advance in understanding the nutritional requirements and response to bile acids of the medically important human gut bacterium, ATCC 35704. A defined medium has been developed which will further the understanding of bile acid metabolism in the context of growth substrates, cofactors, and other metabolites in the vertebrate gut. Analysis of the complete genome supports the nutritional requirements reported here. Genome-wide transcriptomic analysis of gene expression in the presence of cholic acid and deoxycholic acid provides a unique insight into the complex response of ATCC 35704 to primary and secondary bile acids. Also revealed are genes with the potential to function in bile acid transport and metabolism.
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http://dx.doi.org/10.1128/AEM.00052-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6585500PMC
April 2019

Near-infrared fluorescent northern blot.

RNA 2018 12 10;24(12):1871-1877. Epub 2018 Sep 10.

Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, Florida 32610, USA.

Northern blot analysis detects RNA molecules immobilized on nylon membranes through hybridization with radioactive P-labeled DNA or RNA oligonucleotide probes. Alternatively, nonradioactive northern blot relies on chemiluminescent reactions triggered by horseradish peroxidase (HRP) conjugated probes. The use of regulated radioactive material and the complexity of chemiluminescent reactions and detection have hampered the adoption of northern blot techniques by the wider biomedical research community. Here, we describe a sensitive and straightforward nonradioactive northern blot method, which utilizes near-infrared (IR) fluorescent dye-labeled probes (irNorthern). We found that irNorthern has a detection limit of ∼0.05 femtomoles (fmol), which is slightly less sensitive than P-Northern. However, we found that the IR dye-labeled probe maintains the sensitivity after multiple usages as well as long-term storage. We also present alternative irNorthern methods using a biotinylated DNA probe, a DNA probe labeled by terminal transferase, or an RNA probe labeled during in vitro transcription. Furthermore, utilization of different IR dyes allows multiplex detection of different RNA species. Therefore, irNorthern represents a more convenient and versatile tool for RNA detection compared to traditional northern blot analysis.
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http://dx.doi.org/10.1261/rna.068213.118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6239192PMC
December 2018

Mosquito microbiota cluster by host sampling location.

Parasit Vectors 2018 Aug 14;11(1):468. Epub 2018 Aug 14.

Illinois Natural History Survey, University of Illinois at Urbana-Champaign, 1816 S. Oak St., Champaign, IL, 61820, USA.

Background: Microbial communities that inhabit the mosquito body play an import role in host biology and may have potential for mosquito control. However, the forces that shape these microbial communities are poorly understood.

Methods: To gain a better understanding of how host location influences the composition and diversity of mosquito microbiota, we performed a survey of microbial communities in mosquito samples collected from six USA states using HiSeq sequencing of the 16S rRNA gene.

Results: A total of 284 bacterial operational taxonomic units (OTUs) belonging to 14 phyla were detected in nine mosquito species, with Proteobacteria, Firmicutes and Actinobacteria accounting for 95% of total sequences. OTU richness varied markedly within and between mosquito species. The microbial composition and diversity was heavily influenced by the site of mosquito collection, suggesting that host location plays an important role in shaping the mosquito microbiota.

Conclusions: Variation in microbial composition and diversity between mosquitoes from different locations may have important implications on vector competence and transmission dynamics of mosquito-borne pathogens. Future studies should investigate the environmental factors responsible for these variations and the role of key bacteria characterized in this study on mosquito biology and their potential application in symbiotic control of mosquito-borne diseases.
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http://dx.doi.org/10.1186/s13071-018-3036-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6092830PMC
August 2018

Genome Sequence of the Soybean Cyst Nematode (Heterodera glycines) Endosymbiont "Candidatus Cardinium hertigii" Strain cHgTN10.

Genome Announc 2018 Jun 28;6(26). Epub 2018 Jun 28.

Department of Crop Science, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.

In this study, we present the genome sequence of the " Cardinium hertigii" strain cHgTN10, an endosymbiotic bacterium of the plant-parasitic nematode This is the first genome assembly reported for an endosymbiont directly sequenced from a tylenchid nematode.
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http://dx.doi.org/10.1128/genomeA.00624-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025951PMC
June 2018

Bile acid oxidation by Eggerthella lenta strains C592 and DSM 2243.

Gut Microbes 2018 11 24;9(6):523-539. Epub 2018 May 24.

a Department of Microbiology and Immunology , Virginia Commonwealth University , Richmond , VA , USA.

Strains of Eggerthella lenta are capable of oxidation-reduction reactions capable of oxidizing and epimerizing bile acid hydroxyl groups. Several genes encoding these enzymes, known as hydroxysteroid dehydrogenases (HSDH) have yet to be identified. It is also uncertain whether the products of E. lenta bile acid metabolism are further metabolized by other members of the gut microbiota. We characterized a novel human fecal isolate identified as E. lenta strain C592. The complete genome of E. lenta strain C592 was sequenced and comparative genomics with the type strain (DSM 2243) revealed high conservation, but some notable differences. E. lenta strain C592 falls into group III, possessing 3α, 3β, 7α, and 12α-hydroxysteroid dehydrogenase (HSDH) activity, as determined by mass spectrometry of thin layer chromatography (TLC) separated metabolites of primary and secondary bile acids. Incubation of E. lenta oxo-bile acid and iso-bile acid metabolites with whole-cells of the high-activity bile acid 7α-dehydroxylating bacterium, Clostridium scindens VPI 12708, resulted in minimal conversion of oxo-derivatives to lithocholic acid (LCA). Further, Iso-chenodeoxycholic acid (iso-CDCA; 3β,7α-dihydroxy-5β-cholan-24-oic acid) was not metabolized by C. scindens. We then located a gene encoding a novel 12α-HSDH in E. lenta DSM 2243, also encoded by strain C592, and the recombinant purified enzyme was characterized and substrate-specificity determined. Genomic analysis revealed genes encoding an Rnf complex (rnfABCDEG), an energy conserving hydrogenase (echABCDEF) complex, as well as what appears to be a complete Wood-Ljungdahl pathway. Our prediction that by changing the gas atmosphere from nitrogen to hydrogen, bile acid oxidation would be inhibited, was confirmed. These results suggest that E. lenta is an important bile acid metabolizing gut microbe and that the gas atmosphere may be an important and overlooked regulator of bile acid metabolism in the gut.
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http://dx.doi.org/10.1080/19490976.2018.1458180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6287680PMC
November 2018

Defense against territorial intrusion is associated with DNA methylation changes in the honey bee brain.

BMC Genomics 2018 03 26;19(1):216. Epub 2018 Mar 26.

Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL, USA.

Background: Aggression is influenced by individual variation in temperament as well as behavioral plasticity in response to adversity. DNA methylation is stably maintained over time, but also reversible in response to specific environmental conditions, and may thus be a neuromolecular regulator of both of these processes. A previous study reported DNA methylation differences between aggressive Africanized and gentle European honey bees. We investigated whether threat-induced aggression altered DNA methylation profiles in the honey bee brain in response to a behavioral stimulus (aggression-provoking intruder bee or inert control). We sampled five minutes and two hours after stimulus exposure to examine the effect of time on epigenetic profiles of aggression.

Results: There were DNA methylation differences between aggressive and control bees for individual cytosine-guanine dinucleotides (CpGs) across the genome. Eighteen individual CpG sites showed significant difference between aggressive and control bees 120 min post stimulus. For clusters of CpGs, we report four genomic regions differentially methylated between aggressive and control bees at the 5-min time point, and 50 regions differentially methylated at the120-minute time point following intruder exposure. Differential methylation occurred at genes involved in neural plasticity, chromatin remodeling and hormone signaling. Additionally, there was a significant overlap of differential methylation with previously published epigenetic differences that distinguish aggressive Africanized and gentle European honey bees, suggesting an evolutionarily conserved use of brain DNA methylation in the regulation of aggression. Lastly, we identified individually statistically suggestive CpGs that as a group were significantly associated with differentially expressed genes underlying aggressive behavior and also co-localize with binding sites of transcription factors involved in neuroplasticity or neurodevelopment.

Conclusions: There were DNA methylation differences in the brain associated with response to an intruder. These differences increased in number a few hours after the initial exposure and overlap with previously reported aggression-associated genes and neurobiologically relevant transcription factor binding sites. Many DNA methylation differences that occurred in association with the expression of aggression in real time also exist between Africanized bees and European bees, suggesting an evolutionarily conserved role for epigenetic regulation in aggressive behavior.
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http://dx.doi.org/10.1186/s12864-018-4594-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5870497PMC
March 2018

A soft selective sweep during rapid evolution of gentle behaviour in an Africanized honeybee.

Nat Commun 2017 11 16;8(1):1550. Epub 2017 Nov 16.

State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, 650223, Kunming, China.

Highly aggressive Africanized honeybees (AHB) invaded Puerto Rico (PR) in 1994, displacing gentle European honeybees (EHB) in many locations. Gentle AHB (gAHB), unknown anywhere else in the world, subsequently evolved on the island within a few generations. Here we sequence whole genomes from gAHB and EHB populations, as well as a North American AHB population, a likely source of the founder AHB on PR. We show that gAHB retains high levels of genetic diversity after evolution of gentle behaviour, despite selection on standing variation. We observe multiple genomic loci with significant signatures of selection. Rapid evolution during colonization of novel habitats can generate major changes to characteristics such as morphological or colouration traits, usually controlled by one or more major genetic loci. Here we describe a soft selective sweep, acting at multiple loci across the genome, that occurred during, and may have mediated, the rapid evolution of a behavioural trait.
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http://dx.doi.org/10.1038/s41467-017-01800-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5688081PMC
November 2017

The 2017 Bioinformatics Open Source Conference (BOSC).

F1000Res 2017 19;6. Epub 2017 Oct 19.

Open Bioinformatics Foundation, Dublin, Ireland.

The Bioinformatics Open Source Conference (BOSC) is a meeting organized by the Open Bioinformatics Foundation (OBF), a non-profit group dedicated to promoting the practice and philosophy of Open Source software development and Open Science within the biological research community. The 18th annual BOSC ( http://www.open-bio.org/wiki/BOSC_2017) took place in Prague, Czech Republic in July 2017. The conference brought together nearly 250 bioinformatics researchers, developers and users of open source software to interact and share ideas about standards, bioinformatics software development, open and reproducible science, and this year's theme, open data. As in previous years, the conference was preceded by a two-day collaborative coding event open to the bioinformatics community, called the OBF Codefest.
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http://dx.doi.org/10.12688/f1000research.12929.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5649512PMC
October 2017

Adaptation to deep-sea chemosynthetic environments as revealed by mussel genomes.

Nat Ecol Evol 2017 Apr 3;1(5):121. Epub 2017 Apr 3.

Division of Life Science, Hong Kong University of Science and Technology, Hong Kong, China.

Hydrothermal vents and methane seeps are extreme deep-sea ecosystems that support dense populations of specialized macro-benthos such as mussels. But the lack of genome information hinders the understanding of the adaptation of these animals to such inhospitable environments. Here we report the genomes of a deep-sea vent/seep mussel (Bathymodiolus platifrons) and a shallow-water mussel (Modiolus philippinarum). Phylogenetic analysis shows that these mussel species diverged approximately 110.4 million years ago. Many gene families, especially those for stabilizing protein structures and removing toxic substances from cells, are highly expanded in B. platifrons, indicating adaptation to extreme environmental conditions. The innate immune system of B. platifrons is considerably more complex than that of other lophotrochozoan species, including M. philippinarum, with substantial expansion and high expression levels of gene families that are related to immune recognition, endocytosis and caspase-mediated apoptosis in the gill, revealing presumed genetic adaptation of the deep-sea mussel to the presence of its chemoautotrophic endosymbionts. A follow-up metaproteomic analysis of the gill of B. platifrons shows methanotrophy, assimilatory sulfate reduction and ammonia metabolic pathways in the symbionts, providing energy and nutrients, which allow the host to thrive. Our study of the genomic composition allowing symbiosis in extremophile molluscs gives wider insights into the mechanisms of symbiosis in other organisms such as deep-sea tubeworms and giant clams.
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http://dx.doi.org/10.1038/s41559-017-0121DOI Listing
April 2017

Assessing computational genomics skills: Our experience in the H3ABioNet African bioinformatics network.

PLoS Comput Biol 2017 Jun 1;13(6):e1005419. Epub 2017 Jun 1.

Sydney Brenner Institute for Molecular Bioscience, University of the Witwatersrand, Johannesburg, South Africa.

The H3ABioNet pan-African bioinformatics network, which is funded to support the Human Heredity and Health in Africa (H3Africa) program, has developed node-assessment exercises to gauge the ability of its participating research and service groups to analyze typical genome-wide datasets being generated by H3Africa research groups. We describe a framework for the assessment of computational genomics analysis skills, which includes standard operating procedures, training and test datasets, and a process for administering the exercise. We present the experiences of 3 research groups that have taken the exercise and the impact on their ability to manage complex projects. Finally, we discuss the reasons why many H3ABioNet nodes have declined so far to participate and potential strategies to encourage them to do so.
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http://dx.doi.org/10.1371/journal.pcbi.1005419DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5453403PMC
June 2017

Effect of pesticides on microbial communities in container aquatic habitats.

Sci Rep 2017 03 16;7:44565. Epub 2017 Mar 16.

Illinois Natural History Survey, University of Illinois at Urbana-Champaign, 1816 S. Oak St., Champaign IL 61820, USA.

Container aquatic habitats support a specialized community of macroinvertebrates (e.g. mosquitoes) that feed on microbial communities associated with decaying organic matter. These aquatic habitats are often embedded within and around agricultural lands and are frequently exposed to pesticides. We used a microcosm approach to examine the single and combined effects of two herbicides (atrazine, glyphosate), and three insecticides (malathion, carbaryl, permethrin) on microbial communities of container aquatic habitats. MiSeq sequencing of the V4 region of both bacterial and archaeal 16S rRNA gene was used to characterize the microbial communities of indoor microcosms that were either exposed to each pesticide alone, a mix of herbicides, a mix of insecticides, or a mix of all five insecticides. Individual insecticides but not herbicides reduced the microbial diversity and richness and two insecticides, carbaryl and permethrin, also altered the microbial community structure. A mixture of herbicides had no effect on microbial diversity or structure but a mixture of insecticides or all five pesticides reduced microbial diversity and altered the community structure. These findings suggest that exposure of aquatic ecosystems to individual pesticides or their mixtures can disrupt aquatic microbial communities and there is need to decipher how these changes affect resident macroinvertebrate communities.
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http://dx.doi.org/10.1038/srep44565DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5353589PMC
March 2017

Draft Assembly of Elite Inbred Line PH207 Provides Insights into Genomic and Transcriptome Diversity in Maize.

Plant Cell 2016 Nov 1;28(11):2700-2714. Epub 2016 Nov 1.

Roy J. Carver Biotechnology Center, University of Illinois, Urbana, Illinois 61801.

Intense artificial selection over the last 100 years has produced elite maize (Zea mays) inbred lines that combine to produce high-yielding hybrids. To further our understanding of how genome and transcriptome variation contribute to the production of high-yielding hybrids, we generated a draft genome assembly of the inbred line PH207 to complement and compare with the existing B73 reference sequence. B73 is a founder of the Stiff Stalk germplasm pool, while PH207 is a founder of Iodent germplasm, both of which have contributed substantially to the production of temperate commercial maize and are combined to make heterotic hybrids. Comparison of these two assemblies revealed over 2500 genes present in only one of the two genotypes and 136 gene families that have undergone extensive expansion or contraction. Transcriptome profiling revealed extensive expression variation, with as many as 10,564 differentially expressed transcripts and 7128 transcripts expressed in only one of the two genotypes in a single tissue. Genotype-specific genes were more likely to have tissue/condition-specific expression and lower transcript abundance. The availability of a high-quality genome assembly for the elite maize inbred PH207 expands our knowledge of the breadth of natural genome and transcriptome variation in elite maize inbred lines across heterotic pools.
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http://dx.doi.org/10.1105/tpc.16.00353DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5155341PMC
November 2016

The 2016 Bioinformatics Open Source Conference (BOSC).

F1000Res 2016 6;5. Epub 2016 Oct 6.

Plusvital, NovaUCD, Belfield, Dublin 4, Ireland.

The Bioinformatics Open Source Conference (BOSC) is a yearly meeting organized by the Open Bioinformatics Foundation (OBF), a non-profit group dedicated to promoting the practice and philosophy of Open Source software development and Open Science within the biological research community. BOSC has been run since 2000 as a two-day Special Interest Group (SIG) before the annual ISMB conference. The 17th annual BOSC ( http://www.open-bio.org/wiki/BOSC_2016) took place in Orlando, Florida in July 2016. As in previous years, the conference was preceded by a two-day collaborative coding event open to the bioinformatics community. The conference brought together nearly 100 bioinformatics researchers, developers and users of open source software to interact and share ideas about standards, bioinformatics software development, and open and reproducible science.
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http://dx.doi.org/10.12688/f1000research.9663.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5054807PMC
October 2016

Xylan degradation by the human gut Bacteroides xylanisolvens XB1A(T) involves two distinct gene clusters that are linked at the transcriptional level.

BMC Genomics 2016 05 4;17:326. Epub 2016 May 4.

Institut National de la recherche Agronomique (INRA), UR454 Microbiologie, Centre de Clermont-Ferrand-Theix, 63122, Saint-Genès-Champanelle, France.

Background: Plant cell wall (PCW) polysaccharides and especially xylans constitute an important part of human diet. Xylans are not degraded by human digestive enzymes in the upper digestive tract and therefore reach the colon where they are subjected to extensive degradation by some members of the symbiotic microbiota. Xylanolytic bacteria are the first degraders of these complex polysaccharides and they release breakdown products that can have beneficial effects on human health. In order to understand better how these bacteria metabolize xylans in the colon, this study was undertaken to investigate xylan breakdown by the prominent human gut symbiont Bacteroides xylanisolvens XB1A(T).

Results: Transcriptomic analyses of B. xylanisolvens XB1A(T) grown on insoluble oat-spelt xylan (OSX) at mid- and late-log phases highlighted genes in a polysaccharide utilization locus (PUL), hereafter called PUL 43, and genes in a fragmentary remnant of another PUL, hereafter referred to as rPUL 70, which were highly overexpressed on OSX relative to glucose. Proteomic analyses supported the up-regulation of several genes belonging to PUL 43 and showed the important over-production of a CBM4-containing GH10 endo-xylanase. We also show that PUL 43 is organized in two operons and that the knockout of the PUL 43 sensor/regulator HTCS gene blocked the growth of the mutant on insoluble OSX and soluble wheat arabinoxylan (WAX). The mutation not only repressed gene expression in the PUL 43 operons but also repressed gene expression in rPUL 70.

Conclusion: This study shows that xylan degradation by B. xylanisolvens XB1A(T) is orchestrated by one PUL and one PUL remnant that are linked at the transcriptional level. Coupled to studies on other xylanolytic Bacteroides species, our data emphasize the importance of one peculiar CBM4-containing GH10 endo-xylanase in xylan breakdown and that this modular enzyme may be used as a functional marker of xylan degradation in the human gut. Our results also suggest that B. xylanisolvens XB1A(T) has specialized in the degradation of xylans of low complexity. This functional feature may provide a niche to all xylanolytic bacteria harboring similar PULs. Further functional and ecological studies on fibrolytic Bacteroides species are needed to better understand their role in dietary fiber degradation and their impact on intestinal health.
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http://dx.doi.org/10.1186/s12864-016-2680-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4855328PMC
May 2016

Unraveling the pectinolytic function of Bacteroides xylanisolvens using a RNA-seq approach and mutagenesis.

BMC Genomics 2016 Feb 27;17:147. Epub 2016 Feb 27.

Institut National de la Recherche Agronomique (INRA), UR454 Microbiologie, Centre de Clermont-Ferrand/Theix, 63122, Saint-Genès Champanelle, France.

Background: Diet and particularly dietary fibres have an impact on the gut microbiome and play an important role in human health and disease. Pectin is a highly consumed dietary fibre found in fruits and vegetables and is also a widely used additive in the food industry. Yet there is no information on the effect of pectin on the human gut microbiome. Likewise, little is known on gut pectinolytic bacteria and their enzyme systems. This study was undertaken to investigate the mechanisms of pectin degradation by the prominent human gut symbiont Bacteroides xylanisolvens.

Results: Transcriptomic analyses of B. xylanisolvens XB1A grown on citrus and apple pectins at mid- and late-log phases highlighted six polysaccharide utilization loci (PUL) that were overexpressed on pectin relative to glucose. The PUL numbers used in this report are those given by Terrapon et al. (Bioinformatics 31(5):647-55, 2015) and found in the PUL database: http://www.cazy.org/PULDB/. Based on their CAZyme composition, we propose that PUL 49 and 50, the most overexpressed PULs on both pectins and at both growth phases, are involved in homogalacturonan (HG) and type I rhamnogalacturonan (RGI) degradation, respectively. PUL 13 and PUL 2 could be involved in the degradation of arabinose-containing side chains and of type II rhamnogalacturonan (RGII), respectively. Considering that HG is the most abundant moiety (>70%) within pectin, the importance of PUL 49 was further investigated by insertion mutagenesis into the susC-like gene. The insertion blocked transcription of the susC-like and the two downstream genes (susD-like/FnIII). The mutant showed strong growth reduction, thus confirming that PUL 49 plays a major role in pectin degradation.

Conclusion: This study shows the existence of six PULs devoted to pectin degradation by B. xylanisolvens, one of them being particularly important in this function. Hence, this species deploys a very complex enzymatic machinery that probably reflects the structural complexity of pectin. Our findings also highlight the metabolic plasticity of B. xylanisolvens towards dietary fibres that contributes to its competitive fitness within the human gut ecosystem. Wider functional and ecological studies are needed to understand how dietary fibers and especially plant cell wall polysaccharides drive the composition and metabolism of the fibrolytic and non-fibrolytic community within the gut microbial ecosystem.
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http://dx.doi.org/10.1186/s12864-016-2472-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4769552PMC
February 2016

Plasma Exosomal miRNAs in Persons with and without Alzheimer Disease: Altered Expression and Prospects for Biomarkers.

PLoS One 2015 1;10(10):e0139233. Epub 2015 Oct 1.

Department of Psychiatry and Psychiatric Institute, University of Illinois at Chicago, Chicago, Illinois, United States of America.

To assess the value of exosomal miRNAs as biomarkers for Alzheimer disease (AD), the expression of microRNAs was measured in a plasma fraction enriched in exosomes by differential centrifugation, using Illumina deep sequencing. Samples from 35 persons with a clinical diagnosis of AD dementia were compared to 35 age and sex matched controls. Although these samples contained less than 0.1 microgram of total RNA, deep sequencing gave reliable and informative results. Twenty miRNAs showed significant differences in the AD group in initial screening (miR-23b-3p, miR-24-3p, miR-29b-3p, miR-125b-5p, miR-138-5p, miR-139-5p, miR-141-3p, miR-150-5p, miR-152-3p, miR-185-5p, miR-338-3p, miR-342-3p, miR-342-5p, miR-548at-5p, miR-659-5p, miR-3065-5p, miR-3613-3p, miR-3916, miR-4772-3p, miR-5001-3p), many of which satisfied additional biological and statistical criteria, and among which a panel of seven miRNAs were highly informative in a machine learning model for predicting AD status of individual samples with 83-89% accuracy. This performance is not due to over-fitting, because a) we used separate samples for training and testing, and b) similar performance was achieved when tested on technical replicate data. Perhaps the most interesting single miRNA was miR-342-3p, which was a) expressed in the AD group at about 60% of control levels, b) highly correlated with several of the other miRNAs that were significantly down-regulated in AD, and c) was also reported to be down-regulated in AD in two previous studies. The findings warrant replication and follow-up with a larger cohort of patients and controls who have been carefully characterized in terms of cognitive and imaging data, other biomarkers (e.g., CSF amyloid and tau levels) and risk factors (e.g., apoE4 status), and who are sampled repeatedly over time. Integrating miRNA expression data with other data is likely to provide informative and robust biomarkers in Alzheimer disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0139233PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4591334PMC
May 2016