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RNA 1 and RNA 2 Genomic Segments of Chronic Bee Paralysis Virus Are Infectious and Induce Chronic Bee Paralysis Disease.

Youssef I, Schurr F, Goulet A, Cougoule N, Ribière-Chabert M, Darbon H, Thiéry R, Dubois E - J Immunol Res (2015)

Bottom Line: Taken together, these results show that CBPV RNA 1 and CBPV RNA 2 segments can induce virus replication and produce CBPV virus particles.Therefore, the three minor RNAs described in early studies are not essential for virus replication.These data are crucial for the development of a reverse genetic system for CBPV.

View Article: PubMed Central - PubMed

Affiliation: Anses Sophia Antipolis, Unit of Bee Pathology, 105 route des Chappes, CS 20111, 06902 Sophia Antipolis, France ; Architecture et Fonction des Macromolécules Biologiques (AFMB), Aix Marseille University, CNRS, UMR 7257, Case 932, Campus de Luminy, 163 avenue de Luminy, 13288 Marseille Cedex 09, France.

ABSTRACT
Chronic bee paralysis virus (CBPV) causes an infectious and contagious disease of adult honeybees. Its segmented genome is composed of two major positive single-stranded RNAs, RNA 1 (3,674 nt) and RNA 2 (2,305 nt). Three minor RNAs (about 1,000 nt each) have been described earlier but they were not detected by sequencing of CBPV genome. In this study, the results of in vivo inoculation of the two purified CBPV major RNAs are presented and demonstrate that RNA 1 and RNA 2 are infectious. Honeybees inoculated with 10(9) RNA copies per bee developed paralysis symptoms within 6 days after inoculation. The number of CBPV RNA copies increased significantly throughout the infection. Moreover, the negative strand of CBPV RNA was detected by RT-PCR, and CBPV particles were visualized by electronic microscopy in inoculated honeybees. Taken together, these results show that CBPV RNA 1 and CBPV RNA 2 segments can induce virus replication and produce CBPV virus particles. Therefore, the three minor RNAs described in early studies are not essential for virus replication. These data are crucial for the development of a reverse genetic system for CBPV.

No MeSH data available.


Related in: MedlinePlus

DNA standard curve of CBPV RNA 2 RT-qPCR obtained with a 10-fold serial dilution (108 to 102 DNA copies per reaction) of 2,710 kb plasmid including the coding sequence of the predicted structural protein pSP on RNA 2-ORF3. Four independent runs were performed and allowed to obtain the linear regression analysis of the Ct measured for each amplification (y-axis) versus log10 of DNA concentration of each dilution (x-axis). The equation of the linear regression and the correlation coefficient (R2) are indicated.
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fig2: DNA standard curve of CBPV RNA 2 RT-qPCR obtained with a 10-fold serial dilution (108 to 102 DNA copies per reaction) of 2,710 kb plasmid including the coding sequence of the predicted structural protein pSP on RNA 2-ORF3. Four independent runs were performed and allowed to obtain the linear regression analysis of the Ct measured for each amplification (y-axis) versus log10 of DNA concentration of each dilution (x-axis). The equation of the linear regression and the correlation coefficient (R2) are indicated.

Mentions: In order to evaluate the CBPV RNA 2 RT-qPCR, four independent runs were performed using a 10-fold serial dilution of a plasmid DNA control as standard. The standard curve (Figure 2) showed a linear correlation between Ct and log10 DNA concentration of each run (R2 = 0.998). The slope of the DNA standard curve was −3.27 and the average efficiency of RNA 2 RT-qPCR was 102%. The limit of quantification was 100 DNA copies per reaction.


RNA 1 and RNA 2 Genomic Segments of Chronic Bee Paralysis Virus Are Infectious and Induce Chronic Bee Paralysis Disease.

Youssef I, Schurr F, Goulet A, Cougoule N, Ribière-Chabert M, Darbon H, Thiéry R, Dubois E - J Immunol Res (2015)

DNA standard curve of CBPV RNA 2 RT-qPCR obtained with a 10-fold serial dilution (108 to 102 DNA copies per reaction) of 2,710 kb plasmid including the coding sequence of the predicted structural protein pSP on RNA 2-ORF3. Four independent runs were performed and allowed to obtain the linear regression analysis of the Ct measured for each amplification (y-axis) versus log10 of DNA concentration of each dilution (x-axis). The equation of the linear regression and the correlation coefficient (R2) are indicated.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4637131&req=5

fig2: DNA standard curve of CBPV RNA 2 RT-qPCR obtained with a 10-fold serial dilution (108 to 102 DNA copies per reaction) of 2,710 kb plasmid including the coding sequence of the predicted structural protein pSP on RNA 2-ORF3. Four independent runs were performed and allowed to obtain the linear regression analysis of the Ct measured for each amplification (y-axis) versus log10 of DNA concentration of each dilution (x-axis). The equation of the linear regression and the correlation coefficient (R2) are indicated.
Mentions: In order to evaluate the CBPV RNA 2 RT-qPCR, four independent runs were performed using a 10-fold serial dilution of a plasmid DNA control as standard. The standard curve (Figure 2) showed a linear correlation between Ct and log10 DNA concentration of each run (R2 = 0.998). The slope of the DNA standard curve was −3.27 and the average efficiency of RNA 2 RT-qPCR was 102%. The limit of quantification was 100 DNA copies per reaction.

Bottom Line: Taken together, these results show that CBPV RNA 1 and CBPV RNA 2 segments can induce virus replication and produce CBPV virus particles.Therefore, the three minor RNAs described in early studies are not essential for virus replication.These data are crucial for the development of a reverse genetic system for CBPV.

View Article: PubMed Central - PubMed

Affiliation: Anses Sophia Antipolis, Unit of Bee Pathology, 105 route des Chappes, CS 20111, 06902 Sophia Antipolis, France ; Architecture et Fonction des Macromolécules Biologiques (AFMB), Aix Marseille University, CNRS, UMR 7257, Case 932, Campus de Luminy, 163 avenue de Luminy, 13288 Marseille Cedex 09, France.

ABSTRACT
Chronic bee paralysis virus (CBPV) causes an infectious and contagious disease of adult honeybees. Its segmented genome is composed of two major positive single-stranded RNAs, RNA 1 (3,674 nt) and RNA 2 (2,305 nt). Three minor RNAs (about 1,000 nt each) have been described earlier but they were not detected by sequencing of CBPV genome. In this study, the results of in vivo inoculation of the two purified CBPV major RNAs are presented and demonstrate that RNA 1 and RNA 2 are infectious. Honeybees inoculated with 10(9) RNA copies per bee developed paralysis symptoms within 6 days after inoculation. The number of CBPV RNA copies increased significantly throughout the infection. Moreover, the negative strand of CBPV RNA was detected by RT-PCR, and CBPV particles were visualized by electronic microscopy in inoculated honeybees. Taken together, these results show that CBPV RNA 1 and CBPV RNA 2 segments can induce virus replication and produce CBPV virus particles. Therefore, the three minor RNAs described in early studies are not essential for virus replication. These data are crucial for the development of a reverse genetic system for CBPV.

No MeSH data available.


Related in: MedlinePlus