scholarly journals Predicting Intra- and Intertypic Recombination in Enterovirus 71

2018 ◽  
Author(s):  
Andrew Woodman ◽  
Kuo-Ming Lee ◽  
Richard Janissen ◽  
Yu-Nong Gong ◽  
Nynke Dekker ◽  
...  
Keyword(s):  
Acute Flaccid Paralysis ◽  
Recombination Frequency ◽  
Enterovirus 71 ◽  
Template Switching ◽  
Neurological Damage ◽  
A Cell ◽  
Strain Type ◽  
Intertypic Recombinant ◽  
Recombination Junction ◽  
Recombination Assay

AbstractEnteroviruses are well known for their ability to cause neurological damage and paralysis. The model enterovirus is poliovirus (PV), the causative agent of poliomyelitis, a condition characterized by acute flaccid paralysis. A related virus, enterovirus 71 (EV-A71), causes similar clinical outcomes in recurrent outbreaks throughout Asia. Retrospective phylogenetic analysis has shown that recombination between circulating strains of EV-A71 produces the outbreak-associated strains which exhibit increased virulence and/or transmissibility. While studies on the mechanism(s) of recombination in PV are ongoing in several laboratories, little is known about factors that influence recombination in EV-A71. We have developed a cell-based assay to study recombination of EV-A71 based upon previously reported assays for poliovirus recombination. Our results show that: (1) EV-A71 strain-type and RNA sequence diversity impacts recombination frequency in a predictable manner that mimics the observations found in nature; (2) recombination is primarily a replicative process mediated by the RNA-dependent RNA polymerase (RdRp); (3) a mutation shown to reduce recombination in PV (L420A) similarly reduces EV-A71 recombination suggesting conservation in mechanism(s); and (4) sequencing of intertypic recombinant genomes indicates that template-switching is by a mechanism that requires some sequence homology at the recombination junction and that the triggers for template-switching may be sequence independent. The development of this recombination assay will permit further investigation on the interplay between replication, recombination and disease.

scholarly journals Predicting Intraserotypic Recombination in Enterovirus 71

2018 ◽  
Vol 93 (4) ◽  
Author(s):  
Andrew Woodman ◽  
Kuo-Ming Lee ◽  
Richard Janissen ◽  
Yu-Nong Gong ◽  
Nynke H. Dekker ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Iii ◽  
Enterovirus 71 ◽  
Template Switching ◽  
Rna Dependent Rna Polymerase ◽  
A Cell ◽  
Strain Type ◽  
Species Groups ◽  
Recombination Junction ◽  
Recombination Assay

ABSTRACTEnteroviruses are well known for their ability to cause neurological damage and paralysis. The model enterovirus is poliovirus (PV), the causative agent of poliomyelitis, a condition characterized by acute flaccid paralysis. A related virus, enterovirus 71 (EV-A71), causes similar clinical outcomes in recurrent outbreaks throughout Asia. Retrospective phylogenetic analysis has shown that recombination between circulating strains of EV-A71 produces the outbreak-associated strains which exhibit increased virulence and/or transmissibility. While studies on the mechanism(s) of recombination in PV are ongoing in several laboratories, little is known about factors that influence recombination in EV-A71. We have developed a cell-based assay to study recombination of EV-A71 based upon previously reported assays for poliovirus recombination. Our results show that (i) EV-A71 strain type and RNA sequence diversity impacts recombination frequency in a predictable manner that mimics the observations found in nature; (ii) recombination is primarily a replicative process mediated by the RNA-dependent RNA polymerase; (iii) a mutation shown to reduce recombination in PV (L420A) similarly reduces EV-A71 recombination, suggesting conservation in mechanism(s); and (iv) sequencing of intraserotypic recombinant genomes indicates that template switching occurs by a mechanism that may require some sequence homology at the recombination junction and that the triggers for template switching may be sequence independent. The development of this recombination assay will permit further investigation on the interplay between replication, recombination and disease.IMPORTANCERecombination is a mechanism that contributes to genetic diversity. We describe the first assay to study EV-A71 recombination. Results from this assay mimic what is observed in nature and can be used by others to predict future recombination events within the enterovirus species A group. In addition, our results highlight the central role played by the viral RNA-dependent RNA polymerase (RdRp) in the recombination process. Further, our results show that changes to a conserved residue in the RdRp from different species groups have a similar impact on viable recombinant virus yields, which is indicative of conservation in mechanism.

First detection of enterovirus 71 from an acute flaccid paralysis case with residual paralysis in Iran

2008 ◽  
Vol 42 (4) ◽  
pp. 409-411 ◽  
Author(s):  
Shohreh Shahmahmoodi ◽  
Zahra Mehrabi ◽  
Mohammad Reza Eshraghian ◽  
Talat Mokhtari Azad ◽  
Hamideh Tabatabaie ◽  
...  
Keyword(s):  
Acute Flaccid Paralysis ◽  
Enterovirus 71 ◽  
Flaccid Paralysis ◽  
Residual Paralysis ◽  
Acute Flaccid Paralysis Case

scholarly journals Identification of a DNA Segment Exhibiting Rearrangement Modifying Effects upon Transgenic δ-deleting Elements

1997 ◽  
Vol 186 (1) ◽  
pp. 91-100 ◽  
Author(s):  
Karen M. Janowski ◽  
Stephanie Ledbetter ◽  
Matthew S. Mayo ◽  
Richard D. Hockett
Keyword(s):  
T Cells ◽  
T Cell ◽  
Recombination Frequency ◽  
Cell Lineage ◽  
Cell Receptor ◽  
Cell Type ◽  
Reporter Construct ◽  
In Vitro Recombination ◽  
Recombination Assay

Control of the rearrangement and expression of the T cell receptor α and δ chains is critical for determining T cell type. The process of δ deletion is a candidate mechanism for maintaining separation of the α and δ loci. Mice harboring a transgenic reporter δ deletion construct show α/β T cell lineage–specific use of the transgenic elements. A 48-basepair segment of DNA, termed HPS1A, when deleted from this reporter construct, loses tight lineage-specific rearrangement control of transgenic elements, with abundant rearrangements of transgenic δ-deleting elements now in γ/δ T cells. Furthermore, HPS1A augments recombination frequency of extrachromosomal substrates in an in vitro recombination assay. DNA binding proteins recognizing HPS1A have been identified and are restricted to early B and T cells, during the time of active rearrangement of endogenous TCR and immunoglobulin loci. These data are consistent with δ deletion playing an important role in maintaining separate TCR α and δ loci.

scholarly journals Direct Detection and Identification of Enteroviruses from Faeces of Healthy Nigerian Children Using a Cell-Culture Independent RT-Seminested PCR Assay

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Temitope Oluwasegun Cephas Faleye ◽  
Moses Olubusuyi Adewumi ◽  
Bamidele Atinuke Coker ◽  
Felix Yasha Nudamajo ◽  
Johnson Adekunle Adeniji
Keyword(s):  
Cell Culture ◽  
Sequence Data ◽  
Direct Detection ◽  
Clinical Specimen ◽  
Enterovirus 71 ◽  
Healthy Children ◽  
Molecular Sequence Data ◽  
Band Size ◽  
Culture Independent ◽  
A Cell

Recently, a cell-culture independent protocol for detection of enteroviruses from clinical specimen was recommended by the WHO for surveillance alongside the previously established protocols. Here, we investigated whether this new protocol will show the same enterovirus diversity landscape as the established cell-culture dependent protocols. Faecal samples were collected from sixty apparently healthy children in Ibadan, Nigeria. Samples were resuspended in phosphate buffered saline, RNA was extracted, and the VP1 gene was amplified using WHO recommended RT-snPCR protocol. Amplicons were sequenced and sequences subjected to phylogenetic analysis. Fifteen (25%) of the 60 samples yielded the expected band size. Of the 15 amplicons sequenced, 12 were exploitable. The remaining 3 had electropherograms with multiple peaks and were unexploitable. Eleven of the 12 exploitable sequences were identified as Coxsackievirus A1 (CVA1), CVA3, CVA4, CVA8, CVA20, echovirus 32 (E32), enterovirus 71 (EV71), EVB80, and EVC99. Subsequently, the last exploitable sequence was identified as enterobacteriophage baseplate gene by nucleotide BLAST. The results of this study document the first description of molecular sequence data on CVA1, CVA8, and E32 strains present in Nigeria. The result further showed that species A enteroviruses were more commonly detected in the region when cell-culture bias is bypassed.

Involvement of Coprinus endonuclease in preparing substrate for in vitro recombination

Genome ◽  
1990 ◽  
Vol 33 (1) ◽  
pp. 101-108 ◽  
Author(s):  
G. P. Montgomery ◽  
B. C. Lu
Keyword(s):  
Plasmid Dna ◽  
Genetic Recombination ◽  
Recombination Frequency ◽  
Fold Increase ◽  
Electrophoretic Analysis ◽  
Site Specific ◽  
Circular Plasmid ◽  
In Vitro Recombination ◽  
Recombination Assay

A functional recombination assay involving the tetracycline mutant plasmids, pUW1 and pUW4, was used to assess (i) the nature of the DNA substrates needed and (ii) the involvement of Coprinus endonuclease in preparing substrate, for the RecA-directed recombination process. A gapped circular plasmid and a linear or a nicked circular plasmid are efficient substrate combinations in this system to achieve a 160-fold increase in the in vitro recombination frequency over the control levels. The Coprinus endonuclease obtained from early meiotic prophase can produce such substrates. The recombination frequency obtained with the combination of gapped pUW1 plasmids initially relaxed by the Coprinus endonuclease and linear pUW4 plasmids produced by the site-specific BamHI digest is 10-fold lower than that obtained when both substrates are digested by BamHI. The results suggest that the Coprinus endonuclease creates random nicks on plasmid DNA. Glyoxal gel electrophoretic analysis was used to confirm this random nicking activity of Coprinus endonuclease.Key words: Coprinus, genetic recombination, endonuclease, recA.

scholarly journals Recent advances of enterovirus 71 $$3{\rm C}^{{\rm pro}}$$ targeting Inhibitors

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Rominah Onintsoa Diarimalala ◽  
Meichun Hu ◽  
Yanhong Wei ◽  
Kanghong Hu
Keyword(s):  
Public Health ◽  
Life Cycle ◽  
Acute Flaccid Paralysis ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Health Issues ◽  
Mild Disease ◽  
Further Development ◽  
Brain Stem Encephalitis ◽  
Children Under 5

Abstract With CA16, enterovirus-71 is the causative agent of hand foot and mouth disease (HFMD) which occurs mostly in children under 5 years-old and responsible of several outbreaks since a decade. Most of the time, HFMD is a mild disease but can progress to severe complications such as meningitis, brain stem encephalitis, acute flaccid paralysis (AFP) and even death; EV71 has been identified in all severe cases. Therefore, it is actually one of the most public health issues that threatens children’s life. $$3{\rm C}^{{\rm pro}}$$ 3 C pro is a protease which plays important functions in EV71 infection. To date, a lot of $$3{\rm C}^{{\rm pro}}$$ 3 C pro inhibitors have been tested but none of them has been approved yet. Therefore, a drug screening is still an utmost importance in order to treat and/or prevent EV71 infections. This work highlights the EV71 life cycle, $$3{\rm C}^{{\rm pro}}$$ 3 C pro functions and $$3{\rm C}^{{\rm pro}}$$ 3 C pro inhibitors recently screened. It permits to well understand all mechanisms about $$3{\rm C}^{{\rm pro}}$$ 3 C pro and consequently allow further development of drugs targeting $$3{\rm C}^{{\rm pro}}$$ 3 C pro . Thus, this review is helpful for screening of more new $$3{\rm C}^{{\rm pro}}$$ 3 C pro inhibitors or for designing analogues of well known $$3{\rm C}^{{\rm pro}}$$ 3 C pro inhibitors in order to improve its antiviral activity.

scholarly journals The AU-Rich RNA Recombination Hot Spot Sequence of Brome Mosaic Virus Is Functional in Tombusviruses: Implications for the Mechanism of RNA Recombination

2004 ◽  
Vol 78 (5) ◽  
pp. 2288-2300 ◽  
Author(s):  
Natalia Shapka ◽  
Peter D. Nagy
Keyword(s):  
Mosaic Virus ◽  
Hot Spots ◽  
Hot Spot ◽  
Brome Mosaic Virus ◽  
Template Switching ◽  
Rna Recombination ◽  
Replicase Proteins ◽  
Recombination Hot Spots ◽  
Recombination Junction ◽  
Selection Of

ABSTRACT RNA recombination can be facilitated by recombination signals present in viral RNAs. Among such signals are short sequences with high AU contents that constitute recombination hot spots in Brome mosaic virus (BMV) and retroviruses. In this paper, we demonstrate that a defective interfering (DI) RNA, a model template associated with Tomato bushy stunt virus (TBSV), a tombusvirus, undergoes frequent recombination in plants and protoplast cells when it carries the AU-rich hot spot sequence from BMV. Similar to the situation with BMV, most of the recombination junction sites in the DI RNA recombinants were found within the AU-rich region. However, unlike BMV or retroviruses, where recombination usually occurred with precision between duplicated AU-rich sequences, the majority of TBSV DI RNA recombinants were imprecise. In addition, only one copy of the AU-rich sequence was essential to promote recombination in the DI RNA. The selection of junction sites was also influenced by a putative cis-acting element present in the DI RNA. We found that this RNA sequence bound to the TBSV replicase proteins more efficiently than did control nonviral sequences, suggesting that it might be involved in replicase “landing” during the template switching events. In summary, evidence is presented that a tombusvirus can use the recombination signal of BMV. This supports the idea that common AU-rich recombination signals might promote interviral recombination between unrelated viruses.

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