19 research outputs found
The human sodium iodide symporter as a reporter gene for studying Middle East respiratory syndrome coronavirus pathogenesis
Single photon emission computed tomography (SPECT) is frequently used in oncology and cardiology to evaluate disease progression and/or treatment efficacy. Such technology allows for real-time evaluation of disease progression and when applied to studying infectious diseases may provide insight into pathogenesis. Insertion of a SPECT-compatible reporter gene into a virus may provide insight into mechanisms of pathogenesis and viral tropism. The human sodium iodide symporter (hNIS), a SPECT and positron emission tomography reporter gene, was inserted into Middle East respiratory syndrome coronavirus (MERS-CoV), a recently emerged virus that can cause severe respiratory disease and death in afflicted humans to obtain a quantifiable and sensitive marker for viral replication to further MERS-CoV animal model development. The recombinant virus was evaluated for fitness, stability, and reporter gene functionality. The recombinant and parental viruses demonstrated equal fitness in terms of peak titer and replication kinetics, were stable for up to six in vitro passages, and were functional. Further in vivo evaluation indicated variable stability, but resolution limits hampered in vivo functional evaluation. These data support the further development of hNIS for monitoring infection in animal models of viral disease.IMPORTANCE Advanced medical imaging such as single photon emission computed tomography with computed tomography (SPECT/CT) enhances fields such as oncology and cardiology. Application of SPECT/CT, magnetic resonance imaging, and positron emission tomography to infectious disease may enhance pathogenesis studies and provide alternate biomarkers of disease progression. The experiments described in this article focus on insertion of a SPECT/CT-compatible reporter gene into MERS-CoV to demonstrate that a functional SPECT/CT reporter gene can be inserted into a virus
Filovirus RefSeq Entries: Evaluation and Selection of Filovirus Type Variants, Type Sequences, and Names
Sequence determination of complete or coding-complete genomes of viruses is becoming common practice for supporting the work of epidemiologists, ecologists, virologists, and taxonomists. Sequencing duration and costs are rapidly decreasing, sequencing hardware is under modification for use by non-experts, and software is constantly being improved to simplify sequence data management and analysis. Thus, analysis of virus disease outbreaks on the molecular level is now feasible, including characterization of the evolution of individual virus populations in single patients over time. The increasing accumulation of sequencing data creates a management problem for the curators of commonly used sequence databases and an entry retrieval problem for end users. Therefore, utilizing the data to their fullest potential will require setting nomenclature and annotation standards for virus isolates and associated genomic sequences. The National Center for Biotechnology Informationâs (NCBIâs) RefSeq is a non-redundant, curated database for reference (or type) nucleotide sequence records that supplies source data to numerous other databases. Building on recently proposed templates for filovirus variant naming [ ()////-], we report consensus decisions from a majority of past and currently active filovirus experts on the eight filovirus type variants and isolates to be represented in RefSeq, their final designations, and their associated sequences
Virus nomenclature below the species level : a standardized nomenclature for filovirus strains and variants rescued from cDNA
Specific alterations (mutations, deletions,
insertions) of virus genomes are crucial for the functional
characterization of their regulatory elements and their expression products, as well as a prerequisite for the creation
of attenuated viruses that could serve as vaccine
candidates. Virus genome tailoring can be performed either
by using traditionally cloned genomes as starting materials,
followed by site-directed mutagenesis, or by de novo synthesis
of modified virus genomes or parts thereof. A systematic
nomenclature for such recombinant viruses is
necessary to set them apart from wild-type and laboratoryadapted
viruses, and to improve communication and collaborations
among researchers who may want to use
recombinant viruses or create novel viruses based on them.
A large group of filovirus experts has recently proposed
nomenclatures for natural and laboratory animal-adapted
filoviruses that aim to simplify the retrieval of sequence
data from electronic databases. Here, this work is extended
to include nomenclature for filoviruses obtained in the
laboratory via reverse genetics systems. The previously
developed template for natural filovirus genetic variant
naming,\virus name[(\strain[/)\isolation host-suffix[/
\country of sampling[/\year of sampling[/\genetic
variant designation[-\isolate designation[, is retained, but we propose to adapt the type of information added to each
field for cDNA clone-derived filoviruses. For instance, the
full-length designation of an Ebola virus Kikwit variant
rescued from a plasmid developed at the US Centers for
Disease Control and Prevention could be akin to ââEbola
virus H.sapiens-rec/COD/1995/Kikwit-abc1ââ (with the
suffix âârecââ identifying the recombinant nature of the virus
and ââabc1ââ being a placeholder for any meaningful isolate
designator). Such a full-length designation should be used
in databases and the methods section of publications.
Shortened designations (such as ââEBOV H.sap/COD/95/
Kik-abc1ââ) and abbreviations (such as ââEBOV/Kik-abc1ââ)
could be used in the remainder of the text, depending on
how critical it is to convey information contained in the
full-length name. ââEBOVââ would suffice if only one
EBOV strain/variant/isolate is addressed.http://link.springer.com/journal/705hb201
Virus nomenclature below the species level : a standardized nomenclature for laboratory animal-adapted strains and variants of viruses assigned to the family Filoviridae
The International Committee on Taxonomy of Viruses (ICTV) organizes the classification of
viruses into taxa, but is not responsible for the nomenclature for taxa members. International
experts groups, such as the ICTV Study Groups, recommend the classification and naming of
viruses and their strains, variants, and isolates. The ICTV Filoviridae Study Group has recently
introduced an updated classification and nomenclature for filoviruses. Subsequently, and
together with numerous other filovirus experts, a consistent nomenclature for their natural
genetic variants and isolates was developed that aims at simplifying the retrieval of sequence
data from electronic databases. This is a first important step toward a viral genome annotation
standard as sought by the US National Center for Biotechnology Information (NCBI). Here, this
work is extended to include filoviruses obtained in the laboratory by artificial selection through
passage in laboratory hosts. The previously developed template for natural filovirus genetic
variant naming ( //<year of
sampling>/-) is retained, but it is proposed to
adapt the type of information added to each field for laboratory animal-adapted variants. For
instance, the full-length designation of an Ebola virus Mayinga variant adapted at the State
Research Center for Virology and Biotechnology âVectorâ to cause disease in guinea pigs after
seven passages would be akin to âEbola virus VECTOR/C.porcellus-lab/COD/1976/Mayinga-
GPA-P7â. As was proposed for the names of natural filovirus variants, we suggest using the fulllength
designation in databases, as well as in the method section of publications. Shortened
designations (such as âEBOV VECTOR/C.por/COD/76/May-GPA-P7â) and abbreviations (such
as âEBOV/May-GPA-P7â) could be used in the remainder of the text depending on how critical it is to convey information contained in the full-length name. âEBOVâ would suffice if only one
EBOV strain/variant/isolate is addressed.This work was funded in part by the Joint Science and Technology Office for Chem Bio Defense (proposal #TMTI0048_09_RD_T to SB).http://www.springerlink.com/content/0304-8608/hb2013ab201
Figure 1
<p>Body weight and temperature in Syrian hamsters inoculated with MERS-CoV isolate HCoV-EMC/2012. </p
A fixed moderate-dose combination of tiletamine+zolazepam outperforms midazolam in induction of short-term immobilization of ball pythons (Python regius).
Laboratory animals are commonly anesthetized to prevent pain and distress and to provide safe handling. Anesthesia procedures are well-developed for common laboratory mammals, but not as well established in reptiles. We assessed the performance of intramuscularly injected tiletamine (dissociative anesthetic) and zolazepam (benzodiazepine sedative) in fixed combination (2 mg/kg and 3 mg/kg) in comparison to 2 mg/kg of midazolam (benzodiazepine sedative) in ball pythons (Python regius). We measured heart and respiratory rates and quantified induction parameters (i.e., time to loss of righting reflex, time to loss of withdrawal reflex) and recovery parameters (i.e., time to regain righting reflex, withdrawal reflex, normal behavior). Mild decreases in heart and respiratory rates (median decrease of <10 beats per minute and <5 breaths per minute) were observed for most time points among all three anesthetic dose groups. No statistically significant difference between the median time to loss of righting reflex was observed among animals of any group (p = 0.783). However, the withdrawal reflex was lost in all snakes receiving 3mg/kg of tiletamine+zolazepam but not in all animals of the other two groups (p = 0.0004). In addition, the time for animals to regain the righting reflex and resume normal behavior was longer in the drug combination dose groups compared to the midazolam group (p = 0.0055). Our results indicate that midazolam is an adequate sedative for ball pythons but does not suffice to achieve reliable immobilization or anesthesia, whereas tiletamine+zolazepam achieves short-term anesthesia in a dose-dependent manner