scholarly journals Discriminating between JCPyV and BKPyV in Urinary Virome Data Sets

Viruses ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 1041
Author(s):  
Rita Mormando ◽  
Alan J. Wolfe ◽  
Catherine Putonti
Keyword(s):  
Jc Virus ◽  
Sequence Similarity ◽  
Bk Virus ◽  
Data Sets ◽  
Metagenomic Sequencing ◽  
Significant Sequence Similarity ◽  
Sequence Identity ◽  
Shotgun Metagenomic Sequencing ◽  
Urinary Microbiome ◽  
Six Genes

Polyomaviruses are abundant in the human body. The polyomaviruses JC virus (JCPyV) and BK virus (BKPyV) are common viruses in the human urinary tract. Prior studies have estimated that JCPyV infects between 20 and 80% of adults and that BKPyV infects between 65 and 90% of individuals by age 10. However, these two viruses encode for the same six genes and share 75% nucleotide sequence identity across their genomes. While prior urinary virome studies have repeatedly reported the presence of JCPyV, we were interested in seeing how JCPyV prevalence compares to BKPyV. We retrieved all publicly available shotgun metagenomic sequencing reads from urinary microbiome and virome studies (n = 165). While one third of the data sets produced hits to JCPyV, upon further investigation were we able to determine that the majority of these were in fact BKPyV. This distinction was made by specifically mining for JCPyV and BKPyV and considering uniform coverage across the genome. This approach provides confidence in taxon calls, even between closely related viruses with significant sequence similarity.

scholarly journals Methods for Automatic Reference Trees and Multilevel Phylogenetic Placement

2018 ◽  
Author(s):  
Lucas Czech ◽  
Alexandros Stamatakis
Keyword(s):  
Large Scale ◽  
Sequence Data ◽  
Sequence Similarity ◽  
Computational Effort ◽  
Supplementary Information ◽  
Data Sets ◽  
Metagenomic Sequencing ◽  
Sequencing Studies ◽  
Manual Selection ◽  
Supplementary Material

AbstractMotivationIn most metagenomic sequencing studies, the initial analysis step consists in assessing the evolutionary provenance of the sequences. Phylogenetic (or Evolutionary) Placement methods can be employed to determine the evolutionary position of sequences with respect to a given reference phylogeny. These placement methods do however face certain limitations: The manual selection of reference sequences is labor-intensive; the computational effort to infer reference phylogenies is substantially larger than for methods that rely on sequence similarity; the number of taxa in the reference phylogeny should be small enough to allow for visually inspecting the results.ResultsWe present algorithms to overcome the above limitations. First, we introduce a method to automatically construct representative sequences from databases to infer reference phylogenies. Second, we present an approach for conducting large-scale phylogenetic placements on nested phylogenies. Third, we describe a preprocessing pipeline that allows for handling huge sequence data sets. Our experiments on empirical data show that our methods substantially accelerate the workflow and yield highly accurate placement results.ImplementationFreely available under GPLv3 at http://github.com/lczech/[email protected] InformationSupplementary data are available at Bioinformatics online.

scholarly journals T-cell responses to peptide fragments of the BK virus T antigen: implications for cross-reactivity of immune response to JC virus

2006 ◽  
Vol 87 (10) ◽  
pp. 2951-2960 ◽  
Author(s):  
Jongming Li ◽  
Jos Melenhorst ◽  
Nancy Hensel ◽  
Katyoun Rezvani ◽  
Giuseppe Sconocchia ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Jc Virus ◽  
Sequence Similarity ◽  
Simian Virus 40 ◽  
T Cell Responses ◽  
T Antigen ◽  
Bk Virus ◽  
Cell Responses ◽  
Cross Immunity

Infection with BK virus (BKV) induces both humoral and cellular immunity, but the viral antigens of T-antigen (T-ag) stimulating T-cell responses are largely unknown. To identify BKV-specific T cells in healthy individuals, peripheral blood lymphocytes were cultured with autologous dendritic cells (DCs) loaded with BKV lysate and T cells were screened for intracellular gamma interferon production after stimulation with an overlapping 15mer peptide library of the BKV T-ag. Among many immunogenic peptides identified, four T-ag peptides were identified as candidate major histocompatibility complex class I and II T-cell epitopes, restricted to human leukocyte antigen (HLA)-B*0702, -B*08, -DRB1*0301 and -DRB1*0901. Further, a candidate 9mer peptide, LPLMRKAYL, was confirmed to be restricted to HLA-B*0702 and -B*08. Because the polyomaviruses BKV, JC virus (JCV) and Simian virus 40 (SV40) share extensive sequence similarity in the immunogenic proteins T-ag and VP1, it was hypothesized that, in humans, these proteins contain conserved cytotoxic T-lymphocyte (CTL) target epitopes. Four HLA-restricted conserved epitopes of BKV, JCV and SV40 were identified: HLA-B*07, -B*08 and -DRB1*0901 for T-ag and -A*0201 for VP1. T cells cultured in vitro that were specific for one viral antigen recognized other conserved epitopes. CTLs generated from BKV T-ag and VP1 peptide were cytotoxic to DC targets pulsed with either BKV or JCV. Therefore, infection by one of the two viruses (BKV and JCV) could establish cross-immunity against the other. Although cross-cytotoxicity experiments were not performed with SV40, cross-recognition data from conserved antigen epitopes of polyomaviruses suggest strongly that cross-immunity might also exist among the three viruses.

scholarly journals Metagenomic Insights into The Diversity and Functions of Microbial Assemblages in Tasik Kenyir Ecosystem

2017 ◽  
Author(s):  
Mohd Ezhar Mohd Noor ◽  
Sharifah Noor Emilia Syed Jamil Fadaak ◽  
Mohd Noor Mat Isa ◽  
Mohd Faizal Abu Bakar ◽  
Muhd Danish-Daniel Abdullah
Keyword(s):  
Carbon Fixation ◽  
Dna Isolation ◽  
Metagenomic Data ◽  
Data Sets ◽  
Metagenomic Sequencing ◽  
Microbial Species ◽  
Disturbed Area ◽  
Shotgun Metagenomic Sequencing ◽  
Pristine Area ◽  
Disturbed Areas

AbstractTropical freshwater lake such as Tasik Kenyir are underrepresented among the growing number of environmental metagenomic data sets. In Tasik Kenyir, water from two different sites, pristine and disturbed areas were sampled. After the filtration process, genomic DNA from both sites were extracted using Meta-G-nome DNA isolation kit and shotgun metagenomic sequencing was carried out on Illumina HiSeq2500 Desktop Sequencer (Illumina, Inc.). Raw data were then trimmed and assembled using Metagenomic Assembler program, MetaVelvet. Data analysis was carried out using software Blast2GO (BioBam Bioinformatic S.L). The total number of sequence reads was 189,158 from TKS1.5m (disturbed area) and 246,577 from TKS2.5m (pristine area).The results indicate that sequence reads of microbial species were presence at disturbed area near the aquaculture zone was lower than the sequence reads of microbial species were presence at pristine area. When compared to archaea, both samples were dominated by bacteria (more than 90%) suggesting that bacteria are absolutely dominant in the prokaryotic communities in the freshwater samples. The lake appears to contain a mixture of autotrophs and heterotrophs capable of performing main biogeochemical cycles like nitrogen fixation byKlebsiellasp for TKS1.5m andPontibactersp. for TKS2.5m. and carbon fixation by heterotrophicAlcaligenessp. andShewanella decolorationiin TKS1.5m, and byPantoeasp. in TKS2.5m. Present study will advance our understanding of the importance of freshwater microbial communities for ecosystem and human health.

scholarly journals Independent pseudogenizations and losses of sox15 during amniote diversification following asymmetric ohnolog evolution

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yusaku Ogita ◽  
Kei Tamura ◽  
Shuuji Mawaribuchi ◽  
Nobuhiko Takamatsu ◽  
Michihiko Ito
Keyword(s):  
Sequence Similarity ◽  
The Other ◽  
Vertebrate Evolution ◽  
Skeletal Muscle Regeneration ◽  
Cns Development ◽  
Placental Development ◽  
Relaxed Selection ◽  
Significant Sequence Similarity ◽  
Anuran Amphibians ◽  
Divergent Gene

Abstract Background Four ohnologous genes (sox1, sox2, sox3, and sox15) were generated by two rounds of whole-genome duplication in a vertebrate ancestor. In eutherian mammals, Sox1, Sox2, and Sox3 participate in central nervous system (CNS) development. Sox15 has a function in skeletal muscle regeneration and has little functional overlap with the other three ohnologs. In contrast, the frog Xenopus laevis and zebrafish orthologs of sox15 as well as sox1-3 function in CNS development. We previously reported that Sox15 is involved in mouse placental development as neofunctionalization, but is pseudogenized in the marsupial opossum. These findings suggest that sox15 might have evolved with divergent gene fates during vertebrate evolution. However, knowledge concerning sox15 in other vertebrate lineages than therian mammals, anuran amphibians, and teleost fish is scarce. Our purpose in this study was to clarify the fate and molecular evolution of sox15 during vertebrate evolution. Results We searched for sox15 orthologs in all vertebrate classes from agnathans to mammals by significant sequence similarity and synteny analyses using vertebrate genome databases. Interestingly, sox15 was independently pseudogenized at least twice during diversification of the marsupial mammals. Moreover, we observed independent gene loss of sox15 at least twice during reptile evolution in squamates and crocodile-bird diversification. Codon-based phylogenetic tree and selective analyses revealed an increased dN/dS ratio for sox15 compared to the other three ohnologs during jawed vertebrate evolution. Conclusions The findings revealed an asymmetric evolution of sox15 among the four ohnologs during vertebrate evolution, which was supported by the increased dN/dS values in cartilaginous fishes, anuran amphibians, and amniotes. The increased dN/dS value of sox15 may have been caused mainly by relaxed selection. Notably, independent pseudogenizations and losses of sox15 were observed during marsupial and reptile evolution, respectively. Both might have been caused by strong relaxed selection. The drastic gene fates of sox15, including neofunctionalization and pseudogenizations/losses during amniote diversification, might be caused by a release from evolutionary constraints.

scholarly journals Culture-Independent Genotyping, Virulence and Antimicrobial Resistance Gene Identification of Staphylococcus aureus from Orthopaedic Implant-Associated Infections

2021 ◽  
Vol 9 (4) ◽  
pp. 707
Author(s):  
J. Christopher Noone ◽  
Fabienne Antunes Ferreira ◽  
Hege Vangstein Aamot
Keyword(s):  
Staphylococcus Aureus ◽  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Virulence Gene ◽  
Orthopaedic Implant ◽  
Metagenomic Sequencing ◽  
Gene Detection ◽  
Shotgun Metagenomic Sequencing ◽  
Antimicrobial Resistance Genes ◽  
Culture Independent

Our culture-independent nanopore shotgun metagenomic sequencing protocol on biopsies has the potential for same-day diagnostics of orthopaedic implant-associated infections (OIAI). As OIAI are frequently caused by Staphylococcus aureus, we included S. aureus genotyping and virulence gene detection to exploit the protocol to its fullest. The aim was to evaluate S. aureus genotyping, virulence and antimicrobial resistance genes detection using the shotgun metagenomic sequencing protocol. This proof of concept study included six patients with S. aureus-associated OIAI at Akershus University Hospital, Norway. Five tissue biopsies from each patient were divided in two: (1) conventional microbiological diagnostics and genotyping, and whole genome sequencing (WGS) of S. aureus isolates; (2) shotgun metagenomic sequencing of DNA from the biopsies. Consensus sequences were analysed using spaTyper, MLST, VirulenceFinder, and ResFinder from the Center for Genomic Epidemiology (CGE). MLST was also compared using krocus. All spa-types, one CGE and four krocus MLST results matched Sanger sequencing results. Virulence gene detection matched between WGS and shotgun metagenomic sequencing. ResFinder results corresponded to resistance phenotype. S. aureus spa-typing, and identification of virulence and antimicrobial resistance genes are possible using our shotgun metagenomics protocol. MLST requires further optimization. The protocol has potential application to other species and infection types.

scholarly journals Specific Detection of Human BK Polyomavirus in Urine Samples of Immunocompromised Patients

2003 ◽  
Vol 10 (1) ◽  
pp. 66-69 ◽  
Author(s):  
Carol J. Holman ◽  
Jo-Anne H. van Burik ◽  
Steven H. Hinrichs ◽  
Henry H. Balfour
Keyword(s):  
Jc Virus ◽  
Medical Center ◽  
Bk Virus ◽  
Urine Samples ◽  
Proteinase K ◽  
Restriction Enzyme Digestion ◽  
Immunocompromised Patients ◽  
Pcr Assay ◽  
Dna Precipitation ◽  
Serial Dilutions

ABSTRACT A semiquantitative PCR assay for the detection of BK virus in urine was developed using primers for BK virus that specifically amplified BK but not JC virus. DNA was extracted from urine through treatment with proteinase K followed by DNA precipitation with sodium acetate. Semiquantitation was achieved by amplifying serial dilutions (1:1, 1:10, 1:100, and 1:1,000) of the urine specimens. Each assay included both positive (stock BK virus and previously positive patient urine) and negative (no template) controls. A urine sample was interpreted as positive if any of the serial dilutions showed amplification of the DNA fragment of the expected size. For some patient-derived samples, amplification of the expected-size fragment was achieved with a dilute template whereas no amplification was achieved with a concentrated template. This was attributed to interfering substances in the urine. PCR results were compared with urine cytology and shown to be more sensitive. Validation studies were performed at the University of Nebraska Medical Center, utilizing a separate qualitative PCR assay that detects both BK and JC virus and distinguishes between them by restriction enzyme digestion patterns. Of 46 urine samples analyzed using both methods, 22 were positive by both assays, 18 were negative by both assays, 5 were positive only by the Nebraska method, and 1 was positive only by our method. In comparison with the Nebraska PCR, our PCR assay had a sensitivity of 81% and specificity of 95%. For twenty-one (43%) of 49 immunocompromised patients, tests were postive when specimens were submitted because of clinical suspicion of BK virus infection.

An intergenic G-rich region in Leishmania tarentolae kinetoplast maxicircle DNA is a pan-edited cryptogene encoding ribosomal protein S12

1992 ◽  
Vol 12 (1) ◽  
pp. 56-67
Author(s):  
D A Maslov ◽  
N R Sturm ◽  
B M Niner ◽  
E S Gruszynski ◽  
M Peris ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Sequence Similarity ◽  
Rich Region ◽  
Leishmania Tarentolae ◽  
Significant Sequence Similarity ◽  
The Family ◽  
Intergenic Regions ◽  
Ribosomal Protein S12

Six short G-rich intergenic regions in the maxicircle of Leishmania tarentolae are conserved in location and polarity in two other kinetoplastid species. We show here that G-rich region 6 (G6) represents a pan-edited cryptogene which contains at least two domains edited independently in a 3'-to-5' manner connected by short unedited regions. In the completely edited RNA, 117 uridines are added at 49 sites and 32 uridines are deleted at 13 sites, creating a translated 85-amino-acid polypeptide. Similar polypeptides are probably encoded by pan-edited G6 transcripts in two other species. The G6 polypeptide has significant sequence similarity to the family of S12 ribosomal proteins. A minicircle-encoded gRNA overlaps 12 editing sites in G6 mRNA, and chimeric gRNA/mRNA molecules were shown to exist, in agreement with the transesterification model for editing.

scholarly journals Evaluation of the CosmosID Bioinformatics Platform for Prosthetic Joint-Associated Sonicate Fluid Shotgun Metagenomic Data Analysis

2018 ◽  
Vol 57 (2) ◽  
Author(s):  
Qun Yan ◽  
Yu Mi Wi ◽  
Matthew J. Thoendel ◽  
Yash S. Raval ◽  
Kerryl E. Greenwood-Quaintance ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Metagenomic Data ◽  
Metagenomic Sequencing ◽  
Antibacterial Resistance ◽  
Sequencing Data ◽  
Bacterial Detection ◽  
Shotgun Metagenomic Sequencing ◽  
Prosthetic Joint ◽  
Validation Set ◽  
Fluid Culture

ABSTRACT We previously demonstrated that shotgun metagenomic sequencing can detect bacteria in sonicate fluid, providing a diagnosis of prosthetic joint infection (PJI). A limitation of the approach that we used is that data analysis was time-consuming and specialized bioinformatics expertise was required, both of which are barriers to routine clinical use. Fortunately, automated commercial analytic platforms that can interpret shotgun metagenomic data are emerging. In this study, we evaluated the CosmosID bioinformatics platform using shotgun metagenomic sequencing data derived from 408 sonicate fluid samples from our prior study with the goal of evaluating the platform vis-à-vis bacterial detection and antibiotic resistance gene detection for predicting staphylococcal antibacterial susceptibility. Samples were divided into a derivation set and a validation set, each consisting of 204 samples; results from the derivation set were used to establish cutoffs, which were then tested in the validation set for identifying pathogens and predicting staphylococcal antibacterial resistance. Metagenomic analysis detected bacteria in 94.8% (109/115) of sonicate fluid culture-positive PJIs and 37.8% (37/98) of sonicate fluid culture-negative PJIs. Metagenomic analysis showed sensitivities ranging from 65.7 to 85.0% for predicting staphylococcal antibacterial resistance. In conclusion, the CosmosID platform has the potential to provide fast, reliable bacterial detection and identification from metagenomic shotgun sequencing data derived from sonicate fluid for the diagnosis of PJI. Strategies for metagenomic detection of antibiotic resistance genes for predicting staphylococcal antibacterial resistance need further development.

scholarly journals Legionella pneumophila CRISPR-Cas suggests recurrent encounters with one or more phages in the family Microviridae .

2021 ◽  
Author(s):  
Shayna R. Deecker ◽  
Malene L. Urbanus ◽  
Beth Nicholson ◽  
Alexander W. Ensminger
Keyword(s):  
Legionella Pneumophila ◽  
Sequence Similarity ◽  
Mobile Element ◽  
Current Data ◽  
Significant Sequence Similarity ◽  
Remediation Strategies ◽  
The Family ◽  
Legionnaires Disease ◽  
Outstanding Question

Legionella pneumophila is a ubiquitous freshwater pathogen and the causative agent of Legionnaires’ disease. L. pneumophila growth within protists provides a refuge from desiccation, disinfection, and other remediation strategies. One outstanding question has been whether this protection extends to phages. L. pneumophila isolates are remarkably devoid of prophages and to date no Legionella phages have been identified. Nevertheless, many L. pneumophila isolates maintain active CRISPR-Cas defenses. So far, the only known target of these systems is an episomal element that we previously named Legionella Mobile Element-1 (LME-1). The continued expansion of publicly available genomic data promises to further our understanding of the role of these systems. We now describe over 150 CRISPR-Cas systems across 600 isolates to establish the clearest picture yet of L. pneumophila ’s adaptive defenses. By searching for targets of 1,500 unique CRISPR-Cas spacers, LME-1 remains the only identified CRISPR-Cas targeted integrative element. We identified 3 additional LME-1 variants - all targeted by previously and newly identified CRISPR-Cas spacers - but no other similar elements. Notably, we also identified several spacers with significant sequence similarity to microviruses, specifically those within the subfamily Gokushovirinae . These spacers are found across several different CRISPR-Cas arrays isolated from geographically diverse isolates, indicating recurrent encounters with these phages. Our analysis of the extended Legionella CRISPR-Cas spacer catalog leads to two main conclusions: current data argue against CRISPR-Cas targeted integrative elements beyond LME-1, and the heretofore unknown L. pneumophila phages are most likely lytic gokushoviruses. IMPORTANCE Legionnaires’ disease is an often-fatal pneumonia caused by Legionella pneumophila , which normally grows inside amoebae and other freshwater protists. L. pneumophila trades diminished access to nutrients for the protection and isolation provided by the host. One outstanding question is whether L. pneumophila is susceptible to phages, given the protection provided by its intracellular lifestyle. In this work, we use Legionella CRISPR spacer sequences as a record of phage infection to predict that the “missing” L. pneumophila phages belong to the microvirus subfamily Gokushovirinae . Gokushoviruses are known to infect another intracellular pathogen, Chlamydia . How do gokushoviruses access L. pneumophila (and Chlamydia ) inside their “cozy niches”? Does exposure to phages happen during a transient extracellular period (during cell-to-cell spread) or is it indicative of a more complicated environmental lifestyle? One thing is clear, 100 years after their discovery, phages continue to hold important secrets about the bacteria upon which they prey.

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