Publications by authors named "Christopher E Mason"

226 Publications

Sequencing of Circulating Microbial Cell-Free DNA Can Identify Pathogens in Periprosthetic Joint Infections.

J Bone Joint Surg Am 2021 Jul 22. Epub 2021 Jul 22.

Hospital for Special Surgery Research Institute, New York, NY.

Background: Over 1 million Americans undergo joint replacement each year, and approximately 1 in 75 will incur a periprosthetic joint infection. Effective treatment necessitates pathogen identification, yet standard-of-care cultures fail to detect organisms in 10% to 20% of cases and require invasive sampling. We hypothesized that cell-free DNA (cfDNA) fragments from microorganisms in a periprosthetic joint infection can be found in the bloodstream and utilized to accurately identify pathogens via next-generation sequencing.

Methods: In this prospective observational study performed at a musculoskeletal specialty hospital in the U.S., we enrolled 53 adults with validated hip or knee periprosthetic joint infections. Participants had peripheral blood drawn immediately prior to surgical treatment. Microbial cfDNA from plasma was sequenced and aligned to a genome database with >1,000 microbial species. Intraoperative tissue and synovial fluid cultures were performed per the standard of care. The primary outcome was accuracy in organism identification with use of blood cfDNA sequencing, as measured by agreement with tissue-culture results.

Results: Intraoperative and preoperative joint cultures identified an organism in 46 (87%) of 53 patients. Microbial cfDNA sequencing identified the joint pathogen in 35 cases, including 4 of 7 culture-negative cases (57%). Thus, as an adjunct to cultures, cfDNA sequencing increased pathogen detection from 87% to 94%. The median time to species identification for cases with genus-only culture results was 3 days less than standard-of-care methods. Circulating cfDNA sequencing in 14 cases detected additional microorganisms not grown in cultures. At postoperative encounters, cfDNA sequencing demonstrated no detection or reduced levels of the infectious pathogen.

Conclusions: Microbial cfDNA from pathogens causing local periprosthetic joint infections can be detected in peripheral blood. These circulating biomarkers can be sequenced from noninvasive venipuncture, providing a novel source for joint pathogen identification. Further development as an adjunct to tissue cultures holds promise to increase the number of cases with accurate pathogen identification and improve time-to-speciation. This test may also offer a novel method to monitor infection clearance during the treatment period.

Level Of Evidence: Diagnostic Level II. See Instructions for Authors for a complete description of levels of evidence.
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http://dx.doi.org/10.2106/JBJS.20.02229DOI Listing
July 2021

Betacoronavirus-specific alternate splicing.

bioRxiv 2021 Jul 2. Epub 2021 Jul 2.

Viruses can subvert a number of cellular processes in order to block innate antiviral responses, and many viruses interact with cellular splicing machinery. SARS-CoV-2 infection was shown to suppress global mRNA splicing, and at least 10 SARS-CoV-2 proteins bind specifically to one or more human RNAs. Here, we investigate 17 published experimental and clinical datasets related to SARS-CoV-2 infection as well as datasets from the betacoronaviruses SARS-CoV and MERS as well as Streptococcus pneumonia, HCV, Zika virus, Dengue virus, influenza H3N2, and RSV. We show that genes showing differential alternative splicing in SARS-CoV-2 have a similar functional profile to those of SARS-CoV and MERS and affect a diverse set of genes and biological functions, including many closely related to virus biology. Additionally, the differentially spliced transcripts of cells infected by coronaviruses were more likely to undergo intron-retention, contain a pseudouridine modification and a smaller number of exons than differentially spliced transcripts in the control groups. Viral load in clinical COVID-19 samples was correlated with isoform distribution of differentially spliced genes. A significantly higher number of ribosomal genes are affected by DAS and DGE in betacoronavirus samples, and the betacoronavirus differentially spliced genes are depleted for binding sites of RNA-binding proteins. Our results demonstrate characteristic patterns of differential splicing in cells infected by SARS-CoV-2, SARS-CoV, and MERS, potentially modifying a broad range of cellular functions and affecting a diverse set of genes and biological functions.
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http://dx.doi.org/10.1101/2021.07.02.450920DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8259905PMC
July 2021

Haplotype diversity and sequence heterogeneity of human telomeres.

Genome Res 2021 Jun 23. Epub 2021 Jun 23.

Department of Physiology and Biophysics, Weill Cornell Medicine, New York, New York 10065, USA.

Telomeres are regions of repetitive nucleotide sequences capping the ends of eukaryotic chromosomes that protect against deterioration, and whose lengths can be correlated with age and adverse health risk factors. Yet, given their length and repetitive nature, telomeric regions are not easily reconstructed from short-read sequencing, thus making telomere sequencing, mapping, and variant resolution challenging problems. Recently, long-read sequencing, with read lengths measuring in hundreds of kilobase pairs, has made it possible to routinely read into telomeric regions and inspect their sequence structure. Here, we describe a framework for extracting telomeric reads from whole-genome single-molecule sequencing experiments, including de novo identification of telomere repeat motifs and repeat types, and also describe their sequence variation. We find that long, complex telomeric stretches and repeats can be accurately captured with long-read sequencing, observe extensive sequence heterogeneity of human telomeres, discover and localize noncanonical telomere sequence motifs (both previously reported, as well as novel), and validate them in short-read sequence data. These data reveal extensive intra- and inter-population diversity of repeats in telomeric haplotypes, reveal higher paternal inheritance of telomeric variants, and represent the first motif composition maps of multi-kilobase-pair human telomeric haplotypes across three distinct ancestries (Ashkenazi, Chinese, and Utah), which can aid in future studies of genetic variation, aging, and genome biology.
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http://dx.doi.org/10.1101/gr.274639.120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8256856PMC
June 2021

Diurnal variation in the human skin microbiome affects accuracy of forensic microbiome matching.

Microbiome 2021 06 5;9(1):129. Epub 2021 Jun 5.

School of Energy and Environment, City University of Hong Kong, Hong Kong SAR, China.

Background: The human skin microbiome has been recently investigated as a potential forensic tool, as people leave traces of their potentially unique microbiomes on objects and surfaces with which they interact. In this metagenomic study of four people in Hong Kong, their homes, and public surfaces in their neighbourhoods, we investigated the stability and identifiability of these microbiota traces on a timescale of hours to days.

Results: Using a Canberra distance-based method of comparing skin and surface microbiomes, we found that a person could be accurately matched to their household in 84% of tests and to their neighbourhood in 50% of tests, and that matching accuracy did not decay for household surfaces over the 10-day study period, although it did for public surfaces. The time of day at which a skin or surface sample was taken affected matching accuracy, and 160 species across all sites were found to have a significant variation in abundance between morning and evening samples. We hypothesised that daily routines drive a rhythm of daytime dispersal from the pooled public surface microbiome followed by normalisation of a person's microbiome by contact with their household microbial reservoir, and Dynamic Bayesian Networks (DBNs) supported dispersal from public surfaces to skin as the major dispersal route among all sites studied.

Conclusions: These results suggest that in addition to considering the decay of microbiota traces with time, diurnal patterns in microbiome exposure that contribute to the human skin microbiome assemblage must also be considered in developing this as a potential forensic method. Video Abstract.
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http://dx.doi.org/10.1186/s40168-021-01082-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8180031PMC
June 2021

A global metagenomic map of urban microbiomes and antimicrobial resistance.

Cell 2021 Jun 26;184(13):3376-3393.e17. Epub 2021 May 26.

Weill Cornell Medicine, New York, NY, USA; The Bin Talal Bin Abdulaziz Alsaud Institute for Computational Biomedicine, New York, NY, USA.

We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms. Profiles of AMR genes varied widely in type and density across cities. Cities showed distinct microbial taxonomic signatures that were driven by climate and geographic differences. These results constitute a high-resolution global metagenomic atlas that enables discovery of organisms and genes, highlights potential public health and forensic applications, and provides a culture-independent view of AMR burden in cities.
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http://dx.doi.org/10.1016/j.cell.2021.05.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8238498PMC
June 2021

The impact of cross-kingdom molecular forensics on genetic privacy.

Microbiome 2021 05 20;9(1):114. Epub 2021 May 20.

Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY, 10021, USA.

Recent advances in metagenomic technology and computational prediction may inadvertently weaken an individual's reasonable expectation of privacy. Through cross-kingdom genetic and metagenomic forensics, we can already predict at least a dozen human phenotypes with varying degrees of accuracy. There is also growing potential to detect a "molecular echo" of an individual's microbiome from cells deposited on public surfaces. At present, host genetic data from somatic or germ cells provide more reliable information than microbiome samples. However, the emerging ability to infer personal details from different microscopic biological materials left behind on surfaces requires in-depth ethical and legal scrutiny. There is potential to identify and track individuals, along with new, surreptitious means of genetic discrimination. This commentary underscores the need to update legal and policy frameworks for genetic privacy with additional considerations for the information that could be acquired from microbiome-derived data. The article also aims to stimulate ubiquitous discourse to ensure the protection of genetic rights and liberties in the post-genomic era. Video abstract.
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http://dx.doi.org/10.1186/s40168-021-01076-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8138925PMC
May 2021

Investigation of product derived lymphoma following infusion of piggyBac modified CD19 chimeric antigen receptor T-cells.

Blood 2021 May 11. Epub 2021 May 11.

Westmead Hospital, Sydney, Australia.

We performed a Phase I clinical trial of donor derived CD19-specific chimeric antigen receptor T-cells (CAR T-cells) for B-cell malignancy that relapsed or persisted after matched related allogeneic hemopoietic stem cell transplant. To overcome the cost and transgene capacity limitations of traditional viral vectors, CAR T-cells were produced using the piggyBac transposon system of genetic modification. Following CAR T-cell infusion, one patient developed a gradually enlarging retroperitoneal tumor due to a CAR expressing CD4+ T-cell lymphoma. Screening of other patients led to the detection of a second CAR T-cell tumor in thoracic para-aortic lymph nodes in an asymptomatic patient. Analysis of the first lymphoma showed a high transgene copy number, but no insertion into typical oncogenes. There were also structural changes such as altered genomic copy number and point mutations unrelated to the insertion sites. Transcriptome analysis showed transgene promoter driven upregulation of transcription of surrounding regions despite insulator sequences surrounding the transgene. However, marked global changes in transcription predominantly correlated with gene copy number rather than insertion sites. In both patients, the CAR T-cell derived lymphoma progressed and one patient died. We describe the first two cases of malignant lymphoma derived from CAR gene modified T-cells. Although CAR T-cells have an enviable record of safety to date, our results emphasize the need for caution and regular follow up of CAR T recipients, especially when novel methods of gene transfer are used to create genetically modified immune therapies. The trial was registered at www.anzctr.org.au as ACTRN12617001579381.
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http://dx.doi.org/10.1182/blood.2021010858DOI Listing
May 2021

Multiplex qPCR discriminates variants of concern to enhance global surveillance of SARS-CoV-2.

PLoS Biol 2021 05 7;19(5):e3001236. Epub 2021 May 7.

Murphy Medical Associates, Greenwich, Connecticut, United States of America.

With the emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants that may increase transmissibility and/or cause escape from immune responses, there is an urgent need for the targeted surveillance of circulating lineages. It was found that the B.1.1.7 (also 501Y.V1) variant, first detected in the United Kingdom, could be serendipitously detected by the Thermo Fisher TaqPath COVID-19 PCR assay because a key deletion in these viruses, spike Δ69-70, would cause a "spike gene target failure" (SGTF) result. However, a SGTF result is not definitive for B.1.1.7, and this assay cannot detect other variants of concern (VOC) that lack spike Δ69-70, such as B.1.351 (also 501Y.V2), detected in South Africa, and P.1 (also 501Y.V3), recently detected in Brazil. We identified a deletion in the ORF1a gene (ORF1a Δ3675-3677) in all 3 variants, which has not yet been widely detected in other SARS-CoV-2 lineages. Using ORF1a Δ3675-3677 as the primary target and spike Δ69-70 to differentiate, we designed and validated an open-source PCR assay to detect SARS-CoV-2 VOC. Our assay can be rapidly deployed in laboratories around the world to enhance surveillance for the local emergence and spread of B.1.1.7, B.1.351, and P.1.
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http://dx.doi.org/10.1371/journal.pbio.3001236DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8133773PMC
May 2021

The Great Deceiver: miR-2392's Hidden Role in Driving SARS-CoV-2 Infection.

bioRxiv 2021 Apr 27. Epub 2021 Apr 27.

MicroRNAs (miRNAs) are small non-coding RNAs involved in post-transcriptional gene regulation that have a major impact on many diseases and provides an exciting avenue towards antiviral therapeutics. From patient transcriptomic data, we have discovered a circulating miRNA, miR-2392, that is directly involved with SARS-CoV-2 machinery during host infection. Specifically, we found that miR-2392 was key in driving downstream suppression of mitochondrial gene expression, increasing inflammation, glycolysis, and hypoxia as well as promoting many symptoms associated with COVID-19 infection. We demonstrate miR-2392 is present in the blood and urine of COVID-19 patients tested, but not detected in COVID-19 negative patients. These findings indicate the potential for developing a novel, minimally invasive, COVID-19 detection method. Lastly, using both human and hamster models, we have developed a novel miRNA-based antiviral therapeutic targeting miR-2392 that significantly reduces SARS-CoV-2 viability and may potentially inhibit a COVID-19 disease state in the host.
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http://dx.doi.org/10.1101/2021.04.23.441024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8095194PMC
April 2021

Pleckstrin-2 is essential for erythropoiesis in β-thalassemic mice, reducing apoptosis and enhancing enucleation.

Commun Biol 2021 05 3;4(1):517. Epub 2021 May 3.

Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA.

Erythropoiesis involves complex interrelated molecular signals influencing cell survival, differentiation, and enucleation. Diseases associated with ineffective erythropoiesis, such as β-thalassemias, exhibit erythroid expansion and defective enucleation. Clear mechanistic determinants of what make erythropoiesis effective are lacking. We previously demonstrated that exogenous transferrin ameliorates ineffective erythropoiesis in β-thalassemic mice. In the current work, we utilize transferrin treatment to elucidate a molecular signature of ineffective erythropoiesis in β-thalassemia. We hypothesize that compensatory mechanisms are required in β-thalassemic erythropoiesis to prevent apoptosis and enhance enucleation. We identify pleckstrin-2-a STAT5-dependent lipid binding protein downstream of erythropoietin-as an important regulatory node. We demonstrate that partial loss of pleckstrin-2 leads to worsening ineffective erythropoiesis and pleckstrin-2 knockout leads to embryonic lethality in β-thalassemic mice. In addition, the membrane-associated active form of pleckstrin-2 occurs at an earlier stage during β-thalassemic erythropoiesis. Furthermore, membrane-associated activated pleckstrin-2 decreases cofilin mitochondrial localization in β-thalassemic erythroblasts and pleckstrin-2 knockdown in vitro induces cofilin-mediated apoptosis in β-thalassemic erythroblasts. Lastly, pleckstrin-2 enhances enucleation by interacting with and activating RacGTPases in β-thalassemic erythroblasts. This data elucidates the important compensatory role of pleckstrin-2 in β-thalassemia and provides support for the development of targeted therapeutics in diseases of ineffective erythropoiesis.
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http://dx.doi.org/10.1038/s42003-021-02046-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8093212PMC
May 2021

Draft Genome Sequences of Various Bacterial Phyla Isolated from the International Space Station.

Microbiol Resour Announc 2021 Apr 29;10(17). Epub 2021 Apr 29.

Jet Propulsion Laboratory, California Institute of Technology, Pasadena, California, USA

Whole-genome sequences were generated from 96 bacterial strains of 14 species that were isolated from International Space Station surfaces during the Microbial Tracking 2 study. Continued characterization of this closed habitat's microbiome enables tracking of the spread and evolution of secondary pathogens, which is vital for astronaut health.
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http://dx.doi.org/10.1128/MRA.00214-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8086211PMC
April 2021

RP58 Represses Transcriptional Programs Linked to Nonneuronal Cell Identity and Glioblastoma Subtypes in Developing Neurons.

Mol Cell Biol 2021 06 23;41(7):e0052620. Epub 2021 Jun 23.

Weill Cornell Medical College, Department of Neurological Surgery, New York, New York, USA.

How mammalian neuronal identity is progressively acquired and reinforced during development is not understood. We have previously shown that loss of RP58 (ZNF238 or ZBTB18), a BTB/POZ-zinc finger-containing transcription factor, in the mouse brain leads to microcephaly, corpus callosum agenesis, and cerebellum hypoplasia and that it is required for normal neuronal differentiation. The transcriptional programs regulated by RP58 during this process are not known. Here, we report for the first time that in embryonic mouse neocortical neurons a complex set of genes normally expressed in other cell types, such as those from mesoderm derivatives, must be actively repressed and that RP58 is a critical regulator of these repressed transcriptional programs. Importantly, gene set enrichment analysis (GSEA) analyses of these transcriptional programs indicate that repressed genes include distinct sets of genes significantly associated with glioma progression and/or pluripotency. We also demonstrate that reintroducing RP58 in glioma stem cells leads not only to aspects of neuronal differentiation but also to loss of stem cell characteristics, including loss of stem cell markers and decrease in stem cell self-renewal capacities. Thus, RP58 acts as an master guardian of the neuronal identity transcriptome, and its function may be required to prevent brain disease development, including glioma progression.
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http://dx.doi.org/10.1128/MCB.00526-20DOI Listing
June 2021

Early introductions and transmission of SARS-CoV-2 variant B.1.1.7 in the United States.

Cell 2021 05 3;184(10):2595-2604.e13. Epub 2021 Apr 3.

Connecticut State Department of Public Health, Rocky Hill, CT 06067, USA; Centers for Disease Control and Prevention, Atlanta, GA 30329, USA.

The emergence and spread of SARS-CoV-2 lineage B.1.1.7, first detected in the United Kingdom, has become a global public health concern because of its increased transmissibility. Over 2,500 COVID-19 cases associated with this variant have been detected in the United States (US) since December 2020, but the extent of establishment is relatively unknown. Using travel, genomic, and diagnostic data, we highlight that the primary ports of entry for B.1.1.7 in the US were in New York, California, and Florida. Furthermore, we found evidence for many independent B.1.1.7 establishments starting in early December 2020, followed by interstate spread by the end of the month. Finally, we project that B.1.1.7 will be the dominant lineage in many states by mid- to late March. Thus, genomic surveillance for B.1.1.7 and other variants urgently needs to be enhanced to better inform the public health response.
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http://dx.doi.org/10.1016/j.cell.2021.03.061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8018830PMC
May 2021

A verified genomic reference sample for assessing performance of cancer panels detecting small variants of low allele frequency.

Genome Biol 2021 Apr 16;22(1):111. Epub 2021 Apr 16.

Marketing, Integrated DNA Technologies, Inc., 1710 Commercial Park, Coralville, IA, 52241, USA.

Background: Oncopanel genomic testing, which identifies important somatic variants, is increasingly common in medical practice and especially in clinical trials. Currently, there is a paucity of reliable genomic reference samples having a suitably large number of pre-identified variants for properly assessing oncopanel assay analytical quality and performance. The FDA-led Sequencing and Quality Control Phase 2 (SEQC2) consortium analyze ten diverse cancer cell lines individually and their pool, termed Sample A, to develop a reference sample with suitably large numbers of coding positions with known (variant) positives and negatives for properly evaluating oncopanel analytical performance.

Results: In reference Sample A, we identify more than 40,000 variants down to 1% allele frequency with more than 25,000 variants having less than 20% allele frequency with 1653 variants in COSMIC-related genes. This is 5-100× more than existing commercially available samples. We also identify an unprecedented number of negative positions in coding regions, allowing statistical rigor in assessing limit-of-detection, sensitivity, and precision. Over 300 loci are randomly selected and independently verified via droplet digital PCR with 100% concordance. Agilent normal reference Sample B can be admixed with Sample A to create new samples with a similar number of known variants at much lower allele frequency than what exists in Sample A natively, including known variants having allele frequency of 0.02%, a range suitable for assessing liquid biopsy panels.

Conclusion: These new reference samples and their admixtures provide superior capability for performing oncopanel quality control, analytical accuracy, and validation for small to large oncopanels and liquid biopsy assays.
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http://dx.doi.org/10.1186/s13059-021-02316-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8051128PMC
April 2021

Cross-oncopanel study reveals high sensitivity and accuracy with overall analytical performance depending on genomic regions.

Genome Biol 2021 Apr 16;22(1):109. Epub 2021 Apr 16.

Greenwood Genetic Center, 106 Gregor Mendel Circle, Greenwood, SC, 29646, USA.

Background: Targeted sequencing using oncopanels requires comprehensive assessments of accuracy and detection sensitivity to ensure analytical validity. By employing reference materials characterized by the U.S. Food and Drug Administration-led SEquence Quality Control project phase2 (SEQC2) effort, we perform a cross-platform multi-lab evaluation of eight Pan-Cancer panels to assess best practices for oncopanel sequencing.

Results: All panels demonstrate high sensitivity across targeted high-confidence coding regions and variant types for the variants previously verified to have variant allele frequency (VAF) in the 5-20% range. Sensitivity is reduced by utilizing VAF thresholds due to inherent variability in VAF measurements. Enforcing a VAF threshold for reporting has a positive impact on reducing false positive calls. Importantly, the false positive rate is found to be significantly higher outside the high-confidence coding regions, resulting in lower reproducibility. Thus, region restriction and VAF thresholds lead to low relative technical variability in estimating promising biomarkers and tumor mutational burden.

Conclusion: This comprehensive study provides actionable guidelines for oncopanel sequencing and clear evidence that supports a simplified approach to assess the analytical performance of oncopanels. It will facilitate the rapid implementation, validation, and quality control of oncopanels in clinical use.
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http://dx.doi.org/10.1186/s13059-021-02315-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8051090PMC
April 2021

Evaluating the analytical validity of circulating tumor DNA sequencing assays for precision oncology.

Nat Biotechnol 2021 Apr 12. Epub 2021 Apr 12.

R&D Genomics MPS, Institute for Clinical and Experimental Pathology ARUP Laboratories, Salt Lake City, UT, USA.

Circulating tumor DNA (ctDNA) sequencing is being rapidly adopted in precision oncology, but the accuracy, sensitivity and reproducibility of ctDNA assays is poorly understood. Here we report the findings of a multi-site, cross-platform evaluation of the analytical performance of five industry-leading ctDNA assays. We evaluated each stage of the ctDNA sequencing workflow with simulations, synthetic DNA spike-in experiments and proficiency testing on standardized, cell-line-derived reference samples. Above 0.5% variant allele frequency, ctDNA mutations were detected with high sensitivity, precision and reproducibility by all five assays, whereas, below this limit, detection became unreliable and varied widely between assays, especially when input material was limited. Missed mutations (false negatives) were more common than erroneous candidates (false positives), indicating that the reliable sampling of rare ctDNA fragments is the key challenge for ctDNA assays. This comprehensive evaluation of the analytical performance of ctDNA assays serves to inform best practice guidelines and provides a resource for precision oncology.
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http://dx.doi.org/10.1038/s41587-021-00857-zDOI Listing
April 2021

Draft Genome Sequences of and Species Isolated from the International Space Station and Crew Resupply Vehicle Capsule.

Microbiol Resour Announc 2021 Apr 1;10(13). Epub 2021 Apr 1.

Biotechnology and Planetary Protection Group, Jet Propulsion Laboratory, California Institute of Technology, Pasadena, California, USA

The draft whole-genome sequences (WGS) of 30 fungal strains isolated from the International Space Station and belonging to the and genera were assembled. The WGS will allow for detailed genomic characterization to determine the possible applications and importance for space and biotechnological industries.
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http://dx.doi.org/10.1128/MRA.01398-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8104057PMC
April 2021

A comprehensive metagenomics framework to characterize organisms relevant for planetary protection.

Microbiome 2021 04 1;9(1):82. Epub 2021 Apr 1.

Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY, 10065, USA.

Background: Clean rooms of the Space Assembly Facility (SAF) at the Jet Propulsion Laboratory (JPL) at NASA are the final step of spacecraft cleaning and assembly before launching into space. Clean rooms have stringent methods of air-filtration and cleaning to minimize microbial contamination for exoplanetary research and minimize the risk of human pathogens, but they are not sterile. Clean rooms make a selective environment for microorganisms that tolerate such cleaning methods. Previous studies have attempted to characterize the microbial cargo through sequencing and culture-dependent protocols. However, there is not a standardized metagenomic workflow nor analysis pipeline for spaceflight hardware cleanroom samples to identify microbial contamination. Additionally, current identification methods fail to characterize and profile the risk of low-abundance microorganisms.

Results: A comprehensive metagenomic framework to characterize microorganisms relevant for planetary protection in multiple cleanroom classifications (from ISO-5 to ISO-8.5) and sample types (surface, filters, and debris collected via vacuum devices) was developed. Fifty-one metagenomic samples from SAF clean rooms were sequenced and analyzed to identify microbes that could potentially survive spaceflight based on their microbial features and whether the microbes expressed any metabolic activity or growth. Additionally, an auxiliary testing was performed to determine the repeatability of our techniques and validate our analyses. We find evidence that JPL clean rooms carry microbes with attributes that may be problematic in space missions for their documented ability to withstand extreme conditions, such as psychrophilia and ability to form biofilms, spore-forming capacity, radiation resistance, and desiccation resistance. Samples from ISO-5 standard had lower microbial diversity than those conforming to ISO-6 or higher filters but still carried a measurable microbial load.

Conclusions: Although the extensive cleaning processes limit the number of microbes capable of withstanding clean room condition, it is important to quantify thresholds and detect organisms that can inform ongoing Planetary Protection goals, provide a biological baseline for assembly facilities, and guide future mission planning. Video Abstract.
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http://dx.doi.org/10.1186/s40168-021-01020-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8016160PMC
April 2021

sp. nov., Isolated From the International Space Station.

Front Microbiol 2021 15;12:639396. Epub 2021 Mar 15.

Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA, United States.

Four strains belonging to the family of were isolated from different locations on the International Space Station (ISS) across two consecutive flights. Of these, three were identified as Gram-negative, rod-shaped, catalase-positive, oxidase-positive, motile bacteria, designated as IF7SW-B2, IIF1SW-B5, and IIF4SW-B5, whereas the fourth was identified as . The sequence similarity of these three ISS strains, designated as IF7SW-B2, IIF1SW-B5, and IIF4SW-B5, was <99.4% for 16S rRNA genes and <97.3% for gene, with the closest being SE2.11. Furthermore, the multi-locus sequence analysis placed these three ISS strains in the same clade of The average nucleotide identity (ANI) values of these three ISS strains were <93% and digital DNA-DNA hybridization (dDDH) values were <46.4% with any described species. Based on the ANI and dDDH analyses, these three ISS strains were considered as novel species belonging to the genus The three ISS strains showed 100% ANI similarity and dDDH values with each other, indicating that these three ISS strains, isolated during various flights and from different locations, belong to the same species. These three ISS strains were found to grow optimally at temperatures from 25 to 30°C, pH 6.0 to 8.0, and NaCl 0 to 1%. Phenotypically, these three ISS strains resemble and since they assimilate similar sugars as sole carbon substrate when compared to other species. Fatty acid analysis showed that the major fatty acid produced by the ISS strains are C -ω7 and C -ω6. The predominant quinone was ubiquinone 10, and the major polar lipids were diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, and an unidentified lipid. Therefore, based on genomic, phylogenetic, biochemical, and fatty acid analyses, strains IF7SW-B2, IIF1SW-B5, and IIF4SW-B5, are assigned to a novel species within the genus , and the name sp. nov. is proposed. The type strain is IF7SW-B2 (NRRL B-65601 and LMG 32165).
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http://dx.doi.org/10.3389/fmicb.2021.639396DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8005752PMC
March 2021

The spatial landscape of lung pathology during COVID-19 progression.

Nature 2021 05 29;593(7860):564-569. Epub 2021 Mar 29.

Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medicine, New York, NY, USA.

Recent studies have provided insights into the pathology of and immune response to COVID-19. However, a thorough investigation of the interplay between infected cells and the immune system at sites of infection has been lacking. Here we use high-parameter imaging mass cytometry that targets the expression of 36 proteins to investigate the cellular composition and spatial architecture of acute lung injury in humans (including injuries derived from SARS-CoV-2 infection) at single-cell resolution. These spatially resolved single-cell data unravel the disordered structure of the infected and injured lung, alongside the distribution of extensive immune infiltration. Neutrophil and macrophage infiltration are hallmarks of bacterial pneumonia and COVID-19, respectively. We provide evidence that SARS-CoV-2 infects predominantly alveolar epithelial cells and induces a localized hyperinflammatory cell state that is associated with lung damage. We leverage the temporal range of fatal outcomes of COVID-19 in relation to the onset of symptoms, which reveals increased macrophage extravasation and increased numbers of mesenchymal cells and fibroblasts concomitant with increased proximity between these cell types as the disease progresses-possibly as a result of attempts to repair the damaged lung tissue. Our data enable us to develop a biologically interpretable landscape of lung pathology from a structural, immunological and clinical standpoint. We use this landscape to characterize the pathophysiology of the human lung from its macroscopic presentation to the single-cell level, which provides an important basis for understanding COVID-19 and lung pathology in general.
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http://dx.doi.org/10.1038/s41586-021-03475-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204801PMC
May 2021

PCR assay to enhance global surveillance for SARS-CoV-2 variants of concern.

medRxiv 2021 Mar 12. Epub 2021 Mar 12.

Connecticut State Department of Public Health, Rocky Hill, CT 06067, USA.

With the emergence of SARS-CoV-2 variants that may increase transmissibility and/or cause escape from immune responses , there is an urgent need for the targeted surveillance of circulating lineages. It was found that the B.1.1.7 (also 501Y.V1) variant first detected in the UK could be serendipitously detected by the ThermoFisher TaqPath COVID-19 PCR assay because a key deletion in these viruses, spike Δ69-70, would cause a "spike gene target failure" (SGTF) result. However, a SGTF result is not definitive for B.1.1.7, and this assay cannot detect other variants of concern that lack spike Δ69-70, such as B.1.351 (also 501Y.V2) detected in South Africa and P.1 (also 501Y.V3) recently detected in Brazil . We identified a deletion in the ORF1a gene (ORF1a Δ3675-3677) in all three variants, which has not yet been widely detected in other SARS-CoV-2 lineages. Using ORF1a Δ3675-3677 as the primary target and spike Δ69-70 to differentiate, we designed and validated an open source PCR assay to detect SARS-CoV-2 variants of concern . Our assay can be rapidly deployed in laboratories around the world to enhance surveillance for the local emergence spread of B.1.1.7, B.1.351, and P.1.
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http://dx.doi.org/10.1101/2021.01.28.21250486DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987060PMC
March 2021

Systemic Tissue and Cellular Disruption from SARS-CoV-2 Infection revealed in COVID-19 Autopsies and Spatial Omics Tissue Maps.

bioRxiv 2021 Mar 9. Epub 2021 Mar 9.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virus has infected over 115 million people and caused over 2.5 million deaths worldwide. Yet, the molecular mechanisms underlying the clinical manifestations of COVID-19, as well as what distinguishes them from common seasonal influenza virus and other lung injury states such as Acute Respiratory Distress Syndrome (ARDS), remains poorly understood. To address these challenges, we combined transcriptional profiling of 646 clinical nasopharyngeal swabs and 39 patient autopsy tissues, matched with spatial protein and expression profiling (GeoMx) across 357 tissue sections. These results define both body-wide and tissue-specific (heart, liver, lung, kidney, and lymph nodes) damage wrought by the SARS-CoV-2 infection, evident as a function of varying viral load (high vs. low) during the course of infection and specific, transcriptional dysregulation in splicing isoforms, T cell receptor expression, and cellular expression states. In particular, cardiac and lung tissues revealed the largest degree of splicing isoform switching and cell expression state loss. Overall, these findings reveal a systemic disruption of cellular and transcriptional pathways from COVID-19 across all tissues, which can inform subsequent studies to combat the mortality of COVID-19, as well to better understand the molecular dynamics of lethal SARS-CoV-2 infection and other viruses.
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http://dx.doi.org/10.1101/2021.03.08.434433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987017PMC
March 2021

Shotgun transcriptome, spatial omics, and isothermal profiling of SARS-CoV-2 infection reveals unique host responses, viral diversification, and drug interactions.

Nat Commun 2021 03 12;12(1):1660. Epub 2021 Mar 12.

DNAnexus, Inc., Mountain View, CA, USA.

In less than nine months, the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) killed over a million people, including >25,000 in New York City (NYC) alone. The COVID-19 pandemic caused by SARS-CoV-2 highlights clinical needs to detect infection, track strain evolution, and identify biomarkers of disease course. To address these challenges, we designed a fast (30-minute) colorimetric test (LAMP) for SARS-CoV-2 infection from naso/oropharyngeal swabs and a large-scale shotgun metatranscriptomics platform (total-RNA-seq) for host, viral, and microbial profiling. We applied these methods to clinical specimens gathered from 669 patients in New York City during the first two months of the outbreak, yielding a broad molecular portrait of the emerging COVID-19 disease. We find significant enrichment of a NYC-distinctive clade of the virus (20C), as well as host responses in interferon, ACE, hematological, and olfaction pathways. In addition, we use 50,821 patient records to find that renin-angiotensin-aldosterone system inhibitors have a protective effect for severe COVID-19 outcomes, unlike similar drugs. Finally, spatial transcriptomic data from COVID-19 patient autopsy tissues reveal distinct ACE2 expression loci, with macrophage and neutrophil infiltration in the lungs. These findings can inform public health and may help develop and drive SARS-CoV-2 diagnostic, prevention, and treatment strategies.
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http://dx.doi.org/10.1038/s41467-021-21361-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7954844PMC
March 2021

Post-transcriptional regulation of antiviral gene expression by N6-methyladenosine.

Cell Rep 2021 Mar;34(9):108798

Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA; Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA. Electronic address:

Type I interferons (IFNs) induce hundreds of IFN-stimulated genes (ISGs) in response to viral infection. Induction of these ISGs must be regulated for an efficient and controlled antiviral response, but post-transcriptional controls of these genes have not been well defined. Here, we identify a role for the RNA base modification N6-methyladenosine (mA) in the regulation of ISGs. Using ribosome profiling and quantitative mass spectrometry, coupled with mA-immunoprecipitation and sequencing, we identify a subset of ISGs, including IFITM1, whose translation is enhanced by mA and the mA methyltransferase proteins METTL3 and METTL14. We further determine that the mA reader YTHDF1 increases the expression of IFITM1 in an mA-binding-dependent manner. Importantly, we find that the mA methyltransferase complex promotes the antiviral activity of type I IFN. Thus, these studies identify mA as having a role in post-transcriptional control of ISG translation during the type I IFN response for antiviral restriction.
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http://dx.doi.org/10.1016/j.celrep.2021.108798DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7981787PMC
March 2021

Telomere length dynamics in response to DNA damage in malaria parasites.

iScience 2021 Feb 20;24(2):102082. Epub 2021 Jan 20.

Department of Microbiology and Immunology, Weill Cornell Medical College, New York, NY, USA.

Malaria remains a major cause of morbidity and mortality in the developing world. Recent work has implicated chromosome end stability and the repair of DNA breaks through telomere healing as potent drivers of variant antigen diversification, thus associating basic mechanisms for maintaining genome integrity with aspects of host-parasite interactions. Here we applied long-read sequencing technology to precisely examine the dynamics of telomere addition and chromosome end stabilization in response to double-strand breaks within subtelomeric regions. We observed that the process of telomere healing induces the initial synthesis of telomere repeats well in excess of the minimal number required for end stability. However, once stabilized, these newly created telomeres appear to function normally, eventually returning to a length nearing that of intact chromosome ends. These results parallel recent observations in humans, suggesting an evolutionarily conserved mechanism for chromosome end repair.
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http://dx.doi.org/10.1016/j.isci.2021.102082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7887396PMC
February 2021

SARS-CoV-2 genomic diversity and the implications for qRT-PCR diagnostics and transmission.

Genome Res 2021 04 18;31(4):635-644. Epub 2021 Feb 18.

Department of Computer Science, Rice University, Houston, Texas 77005, USA.

The COVID-19 pandemic has sparked an urgent need to uncover the underlying biology of this devastating disease. Though RNA viruses mutate more rapidly than DNA viruses, there are a relatively small number of single nucleotide polymorphisms (SNPs) that differentiate the main SARS-CoV-2 lineages that have spread throughout the world. In this study, we investigated 129 RNA-seq data sets and 6928 consensus genomes to contrast the intra-host and inter-host diversity of SARS-CoV-2. Our analyses yielded three major observations. First, the mutational profile of SARS-CoV-2 highlights intra-host single nucleotide variant (iSNV) and SNP similarity, albeit with differences in C > U changes. Second, iSNV and SNP patterns in SARS-CoV-2 are more similar to MERS-CoV than SARS-CoV-1. Third, a significant fraction of insertions and deletions contribute to the genetic diversity of SARS-CoV-2. Altogether, our findings provide insight into SARS-CoV-2 genomic diversity, inform the design of detection tests, and highlight the potential of iSNVs for tracking the transmission of SARS-CoV-2.
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http://dx.doi.org/10.1101/gr.268961.120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8015855PMC
April 2021

Characterization of complete lncRNAs transcriptome reveals the functional and clinical impact of lncRNAs in multiple myeloma.

Leukemia 2021 05 17;35(5):1438-1450. Epub 2021 Feb 17.

Área de Oncología, Centro de Investigación Médica Aplicada (CIMA), Universidad de Navarra, IDISNA, Pamplona, Spain.

Multiple myeloma (MM) is an incurable disease, whose clinical heterogeneity makes its management challenging, highlighting the need for biological features to guide improved therapies. Deregulation of specific long non-coding RNAs (lncRNAs) has been shown in MM, nevertheless, the complete lncRNA transcriptome has not yet been elucidated. In this work, we identified 40,511 novel lncRNAs in MM samples. lncRNAs accounted for 82% of the MM transcriptome and were more heterogeneously expressed than coding genes. A total of 10,351 overexpressed and 9,535 downregulated lncRNAs were identified in MM patients when compared with normal bone-marrow plasma cells. Transcriptional dynamics study of lncRNAs in the context of normal B-cell maturation revealed 989 lncRNAs with exclusive expression in MM, among which 89 showed de novo epigenomic activation. Knockdown studies on one of these lncRNAs, SMILO (specific myeloma intergenic long non-coding RNA), resulted in reduced proliferation and induction of apoptosis of MM cells, and activation of the interferon pathway. We also showed that the expression of lncRNAs, together with clinical and genetic risk alterations, stratified MM patients into several progression-free survival and overall survival groups. In summary, our global analysis of the lncRNAs transcriptome reveals the presence of specific lncRNAs associated with the biological and clinical behavior of the disease.
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http://dx.doi.org/10.1038/s41375-021-01147-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8102198PMC
May 2021

Early introductions and community transmission of SARS-CoV-2 variant B.1.1.7 in the United States.

medRxiv 2021 Feb 12. Epub 2021 Feb 12.

Connecticut State Department of Public Health, Rocky Hill, CT 06067, USA.

The emergence and spread of SARS-CoV-2 lineage B.1.1.7, first detected in the United Kingdom, has become a national public health concern in the United States because of its increased transmissibility. Over 500 COVID-19 cases associated with this variant have been detected since December 2020, but its local establishment and pathways of spread are relatively unknown. Using travel, genomic, and diagnostic testing data, we highlight the primary ports of entry for B.1.1.7 in the US and locations of possible underreporting of B.1.1.7 cases. New York, which receives the most international travel from the UK, is likely one of the key hubs for introductions and domestic spread. Finally, we provide evidence for increased community transmission in several states. Thus, genomic surveillance for B.1.1.7 and other variants urgently needs to be enhanced to better inform the public health response.
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http://dx.doi.org/10.1101/2021.02.10.21251540DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7885932PMC
February 2021
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