Generation of duck Tembusu virus using a simple reverse genetic system in duck embryo fibroblast cells

Outbreaks of duck Tembusu virus (DTMUV) have caused substantial economic losses in the major duck-producing regions of China since 2010. To improve our understanding of the host cellular responses to virus infection and the pathogenesis of DTMUV infection, we applied isobaric tags for relative and absolute quantification (iTRAQ) labeling coupled with multidimensional liquid chromatography-tandem mass spectrometry to detect the protein changes in duck embryo fibroblast cells (DEFs) infected and mock-infected with DTMUV. In total, 434 cellular proteins were differentially expressed, among which 116, 76, and 339 proteins were differentially expressed in the DTMUV-infected DEFs at 12, 24, and 42 hours postinfection, respectively. The Gene Ontology analysis indicated that the biological processes of the differentially expressed proteins were primarily related to cellular processes, metabolic processes, biological regulation, response to stimulus, and cellular organismal processes and that the molecular functions in which the differentially expressed proteins were mainly involved were binding and catalytic activity. Some selected proteins that were found to be differentially expressed in DTMUV-infected DEFs were further confirmed by real-time PCR. The results of this study provide valuable insight into DTMUV-host interactions. This could lead to a better understanding of DTMUV infection mechanisms.

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Development of a Reverse Genetic System to Generate Recombinant Chimeric Tacaribe Virus that Expresses Junín Virus Glycoproteins

Pathogens ◽

10.3390/pathogens9110948 ◽

2020 ◽

Vol 9 (11) ◽

pp. 948

Author(s):

Sabrina Foscaldi ◽

María Eugenia Loureiro ◽

Claudia Sepúlveda ◽

Carlos Palacios ◽

María Belén Forlenza ◽

...

Keyword(s):

Hemorrhagic Fever ◽

Genetic System ◽

Reverse Genetic System ◽

Dependent Manner ◽

Reverse Genetic ◽

Junin Virus ◽

Tacaribe Virus ◽

Growth Properties ◽

Intracellular Expression ◽

Junín Virus

Mammarenaviruses are enveloped and segmented negative-stranded RNA viruses that comprise several pathogenic members associated with severe human hemorrhagic fevers. Tacaribe virus (TCRV) is the prototype for the New World group of mammarenaviruses and is not only naturally attenuated but also phylogenetically and antigenically related to all South American pathogenic mammarenaviruses, particularly the Junín virus (JUNV), which is the etiological agent of Argentinian hemorrhagic fever (AHF). Moreover, since TCRV protects guinea pigs and non-human primates from lethal challenges with pathogenic strains of JUNV, it has already been considered as a potential live-attenuated virus vaccine candidate against AHF. Here, we report the development of a reverse genetic system that relies on T7 polymerase-driven intracellular expression of the complementary copy (antigenome) of both viral S and L RNA segments. Using this approach, we successfully recovered recombinant TCRV (rTCRV) that displayed growth properties resembling those of authentic TCRV. We also generated a chimeric recombinant TCRV expressing the JUNV glycoproteins, which propagated similarly to wild-type rTCRV. Moreover, a controlled modification within the S RNA 5′ non-coding terminal sequence diminished rTCRV propagation in a cell-type dependent manner, giving rise to new perspectives where the incorporation of additional attenuation markers could contribute to develop safe rTCRV-based vaccines against pathogenic mammarenaviruses.

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Establishment of an efficient reverse genetic system of Mumps virus S79 from cloned DNA

World Journal of Pediatrics ◽

10.1007/s12519-019-00286-8 ◽

2019 ◽

Vol 15 (5) ◽

pp. 499-505 ◽

Cited By ~ 1

Author(s):

Duo Zhou ◽

Meng-Ying Zhu ◽

Yi-Long Wang ◽

Xiao-Qiang Hao ◽

Dong-Ming Zhou ◽

...

Keyword(s):

Genetic System ◽

Mumps Virus ◽

Reverse Genetic System ◽

Reverse Genetic

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Characterization and reverse genetic establishment of cattle derived Akabane virus in China

BMC Veterinary Research ◽

10.1186/s12917-021-03054-x ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Dongjie Chen ◽

Di Wang ◽

Fang Wei ◽

Yufang Kong ◽

Junhua Deng ◽

...

Keyword(s):

Genetic System ◽

Reverse Genetic System ◽

Akabane Virus ◽

Reverse Genetic ◽

Wild Type ◽

T7 Promoter ◽

Cytopathic Effects ◽

The World ◽

L Segment ◽

Genome Manipulation

Abstract Background Akabane virus (AKAV) is an important insect-borne virus which is widely distributed throughout the world except the Europe and is considered as a great threat to herbivore health. Results An AKAV strain defined as TJ2016 was firstly isolated from the bovine sera in China in 2016. Sequence analysis of the S and M segments suggested that the isolated AKAV strain was closely related to the AKAV strains JaGAr39 and JaLAB39, which belonged to AKAV genogroup II. To further study the pathogenic mechanism of AKAV, the full-length cDNA clone of TJ2016 S, M, and L segment was constructed separately into the TVT7R plasmid at the downsteam of T7 promoter and named as TVT7R-S, TVT7R-M, and TVT7R-L, respectively. The above three plasmids were further transfected into the BSR-T7/5 cells simultaneously with a ratio of 1:1:1 to produce the rescued virus AKAV. Compared with the parental wild type AKAV (wtAKAV), the rescued virus (rAKAV) was proved to be with similar cytopathic effects (CPE), plaque sizes and growth kinetics in BHK-21 cells. Conclusion We successfully isolated a AKAV strain TJ2016 from the sera of cattle and established a reverse genetic platform for AKAV genome manipulation. The established reverse genetic system is also a powerful tool for further research on AKAV pathogenesis and even vaccine studies.

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