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A novel ViewRNA in situ hybridization method for the detection of the dynamic distribution of Classical Swine Fever Virus RNA in PK15 cells

Overview of attention for article published in Virology Journal, April 2017
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  • Above-average Attention Score compared to outputs of the same age (53rd percentile)
  • Above-average Attention Score compared to outputs of the same age and source (58th percentile)

Mentioned by

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4 tweeters

Citations

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2 Dimensions

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14 Mendeley
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Title
A novel ViewRNA in situ hybridization method for the detection of the dynamic distribution of Classical Swine Fever Virus RNA in PK15 cells
Published in
Virology Journal, April 2017
DOI 10.1186/s12985-017-0734-4
Pubmed ID
Authors

Qianyi Zhang, Lu Xu, Yujie Zhang, Tuanjie Wang, Xingqi Zou, Yuanyuan Zhu, Yan Zhao, Cui Li, Kai Chen, Yongfang Sun, Junxiang Sun, Qizu Zhao, Qin Wang

Abstract

Classical swine fever (CSF) is a highly contagious fatal infectious disease caused by classical swine fever virus (CSFV). A better understanding of CSFV replication is important for the study of pathogenic mechanism of CSF. With the development of novel RNA in situ Hybridization method, quantitatively localization and visualization of the virus RNA molecular in cultured cell or tissue section becomes very important tool to address these pivotal pathogenic questions. In this study, we established ViewRNA ISH method to reveal the dynamic distribution of CSFV RNA in PK15 cells. We designed several specific probes of CSFV RNA and reference gene β-actin for host PK15 cells to monitor the relative location of CSFV RNA and house-keeping gene in the infected cells. After determining the titer of reference strain CSFV (HeBHH1/95) with the 50% tissue culture infective dose (TCID50), we optimized the protease K concentration and formalin fixation time to analyze the hybridization efficiency, fluorescence intensity and repeatability. In order to measure the sensitivity of this assay, we compared it with the fluorescent antibody test (FAT) and immunohistochemical(IHC) method. Specificity of the ViewRNA ISH was tested by detecting several sub genotypes of CSFV (sub genotype 1.1, 2.1, 2.2 and 2.3) which are present in China and other normal pig infectious virus (bovine viral diarrhea virus (BVDV), porcine parvovirus (PPV), porcine pseudorabies virus (PRV) and porcine circovirusII(PCV-2). The lowest detection threshold of the ViewRNA ISH method was 10(-8), while the sensitivity of FAT and IHC were 10(-5) and 10(-4), respectively. The ViewRNA ISH was specific for CSFV RNA including 1.1, 2.1, 2.2 and 2.3 subtypes, meanwhile, there was no cross-reaction with negative control and other viruses including BVDV, PPV, PRV and PCV-2. Our results showed that after infection at 0.5 hpi (hours post inoculation, hpi), the CSFV RNA can be detected in nucleus and cytoplasm; during 3-9 hpi, RNA was mainly distributed in nucleus and reached a maximum at 12hpi, then RNA copy number was gradually increased around the cell nucleus during 24-48 hpi and reached the peak at 72hpi. To our knowledge, this is the first to reveal the dynamic distribution of medium virulence CSFV RNA in PK15 cells using the ViewRNA ISH method. The sensitivity of the ViewRNA ISH was three to four orders of magnitude higher than that of FAT and IHC methods. The specificity experiment showed that the ViewRNA ISH was highly specific for CSFV and no cross-reaction occurred to negative control and other pig infectious virus. This assay is more suitable for studying the CSFV RNA life cycle in cell nucleus. The results proved that CSFV RNA enters into PK15 cells earlier than 0.5hpi, relative to the eclipse period of cytoplasm is 6-9 hpi and CSFV RNA has ever existed in nucleus.

Twitter Demographics

The data shown below were collected from the profiles of 4 tweeters who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 14 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Unknown 14 100%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 3 21%
Student > Doctoral Student 3 21%
Professor > Associate Professor 2 14%
Researcher 2 14%
Student > Master 1 7%
Other 1 7%
Unknown 2 14%
Readers by discipline Count As %
Veterinary Science and Veterinary Medicine 6 43%
Agricultural and Biological Sciences 2 14%
Neuroscience 1 7%
Medicine and Dentistry 1 7%
Unknown 4 29%

Attention Score in Context

This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 01 May 2017.
All research outputs
#6,372,648
of 11,379,716 outputs
Outputs from Virology Journal
#929
of 1,959 outputs
Outputs of similar age
#119,591
of 264,772 outputs
Outputs of similar age from Virology Journal
#13
of 36 outputs
Altmetric has tracked 11,379,716 research outputs across all sources so far. This one is in the 43rd percentile – i.e., 43% of other outputs scored the same or lower than it.
So far Altmetric has tracked 1,959 research outputs from this source. They receive a mean Attention Score of 3.5. This one is in the 48th percentile – i.e., 48% of its peers scored the same or lower than it.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 264,772 tracked outputs that were published within six weeks on either side of this one in any source. This one has gotten more attention than average, scoring higher than 53% of its contemporaries.
We're also able to compare this research output to 36 others from the same source and published within six weeks on either side of this one. This one has gotten more attention than average, scoring higher than 58% of its contemporaries.