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  • PeerJ  (3)
  • Li, Ming  (3)
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  • 1
    In: PeerJ, PeerJ, Vol. 8 ( 2020-03-11), p. e8750-
    Abstract: Deformed wing virus (DWV) is a serious threat to honey bees ( Apis mellifera ) and is considered a major cause of elevated losses of honey bee colonies. However, lack of information on the immunogenicity of DWV structural proteins has hindered the development of effective biocontrol drugs. Methods We optimized the VP1 , VP2 and VP3 codons of DWV surface capsid protein genes on the basis of an Escherichia coli codon bias, and the optimized genes of roVP1 , roVP2 and roVP3 were separately expressed in E. coli and purified. Next, the three recombinant proteins of roVP1 , roVP2 and roVP3 were intramuscularly injected into BALB/c and the immunogenicity was evaluated by the levels of specific IgG and cytokines. Furthermore, anti- roVP -antisera ( roVP1 or roVP2 or roVP3 ) from the immunized mice was incubated with DWV for injecting healthy white-eyed pupae for the viral challenge test, respectively. Results The optimized genes roVP1 , roVP2 and roVP3 achieved the expression in E. coli using SDS-PAGE and Western blotting. Post-immunization, roVP2 and roVP3 exhibited higher immunogenicity than roVP1 and stimulated a stronger humoral immune response in the mice, which showed that the recombinant proteins of roVP3 and roVP2 induced a specific immune response in the mice. In the challenge test, data regarding quantitative real-time RT-PCR (qRT-PCR) from challenged pupae showed that the level of virus copies in the recombinant protein groups was significantly lower than that of the virus-only group at 96 h post-inoculation ( P 〈 0.05). Among them, the degree of neutralization using antibodies raised to the recombinant proteins are between approximately 2-fold and 4-fold and the virus copies of the roVP3 group are the lowest in the three recombinant protein groups, which indicated that specific antibodies against recombinant proteins roVP1 , roVP2 and roVP3 of DWV could neutralize DWV to reduce the virus titer in the pupae. Collectively, these results demonstrated that the surface capsid protein of DWV acted as candidates for the development of therapeutic antibodies against the virus.
    Type of Medium: Online Resource
    ISSN: 2167-8359
    Language: English
    Publisher: PeerJ
    Publication Date: 2020
    detail.hit.zdb_id: 2703241-3
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  • 2
    Online Resource
    Online Resource
    PeerJ ; 2019
    In:  PeerJ Vol. 7 ( 2019-11-14), p. e8003-
    In: PeerJ, PeerJ, Vol. 7 ( 2019-11-14), p. e8003-
    Abstract: Sacbrood virus (SBV) is one of the most pathogenic honeybee viruses that exhibits host specificity and regional variations. The SBV strains that infect the Chinese honeybee Apis cerana are called Chinese SBVs (CSBVs). Methods In this study, a CSBV strain named AmCSBV-SDLY-2016 (GenBank accession No. MG733283 ) infecting A. mellifera was identified by electron microscopy, its protein composition was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and agar gel immunodiffusion assay, and its nucleotide sequence was identified using a series of reverse-transcription polymerase chain reaction fragments of AmCSBV-SDLY-2016 generated using SBV/CSBV-specific primers. To investigate phylogenetic relationships of the CSBV isolates, a phylogenetic tree of the complete open reading frames (ORF) of the CSBV sequences was constructed using MEGA 6.0; then, the similarity and recombination events among the isolated CSBV strains were analyzed using SimPlot and RDP4 software, respectively. Results Sequencing results revealed the complete 8,794-nucleotide long complete genomic RNA of the strain, with a single large ORF (189–8,717) encoding 2,843 amino acids. Comparison of the deduced amino acid sequence with the SBV/CSBV reference sequences deposited in the GenBank database identified helicase, protease, and RNA-dependent RNA polymerase domains; the structural genes were located at the 5′ end, whereas the non-structural genes were found at the 3′ end. Multiple sequence alignment showed that AmCSBV-SDLY-2016 had a 17-amino acid (aa) and a single aa deletion at positions 711–729 and 2,128, respectively, as compared with CSBV-GD-2002, and a 16-aa deletion (positions 711–713 and 715–728) as compared with AmSBV-UK-2000. However, AmCSBV-SDLY-2016 was similar to the CSBV-JLCBS-2014 strain, which infects A. cerana . AmCSBV-SDLY-2016 ORF shared 92.4–97.1% identity with the genomes of other CSBV strains (94.5–97.7% identity for deduced amino acids). AmCSBV-SDLY-2016 was least similar (89.5–90.4% identity) to other SBVs but showed maximum similarity with the previously reported CSBV-FZ-2014 strain. The phylogenetic tree constructed from AmCSBV-SDLY-2016 and 43 previously reported SBV/CSBV sequences indicated that SBV/CSBV strains clustered according to the host species and country of origin; AmCSBV-SDLY-2016 clustered with other previously reported Chinese and Asian strains (AC genotype SBV, as these strains originated from A. cerana ) but was separate from the SBV genomes originating from Europe (AM genotype SBV, originating from A. mellifera ). A SimPlot graph of SBV genomes confirmed the high variability, especially between the AC genotype SBV and AM genotype SBV. This genomic diversity may reflect the adaptation of SBV to specific hosts, ability of CSBV to cross the species barrier, and the spatial distances that separate CSBVs from other SBVs.
    Type of Medium: Online Resource
    ISSN: 2167-8359
    Language: English
    Publisher: PeerJ
    Publication Date: 2019
    detail.hit.zdb_id: 2703241-3
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  • 3
    In: PeerJ, PeerJ, Vol. 7 ( 2019-06-28), p. e7214-
    Abstract: Deformed wing virus (DWV) is one of many viruses that infect honeybees and has been extensively studied because of its close association with honeybee colony collapse that is induced by Varroa destructor . However, virus genotypes, sequence characteristics, and genetic variations of DWV remain unknown in China. Methods Two DWV strains were isolated from Jinzhou and Qinhuangdao cities in China, and were named China1-2017 (accession number: MF770715 ) and China2-2018 (accession number: MH165180 ), respectively, and their complete genome sequences were analyzed. To investigate the phylogenetic relationships of the DWV isolates, a phylogenetic tree of the complete open reading frame (ORF), structural protein VP1, and non-structural protein 3C+RdRp of the DWV sequences was constructed using the MEGA 5.0 software program. Then, the similarity and recombinant events of the DWV isolated strains were analyzed using recombination detection program (RDP4) software and genetic algorithm for recombination detection (GARD). Results The complete genomic analysis showed that the genomes of the China1-2017 and China2-2018 DWV strains consisted of 10,141 base pairs (bp) and 10,105 bp, respectively, and contained a single, large ORF (China1-2017: 1,146–9,827 bp; China2-2018: 1,351–9,816 bp) that encoded 2,894 amino acids. The sequences were compared with 20 previously reported DWV sequences from different countries and with sequences of two closely related viruses, Kakugo virus (KV) and V. destructor virus-1. Multiple sequence comparisons revealed a nucleotide identity of 84.3–96.7%, and identity of 94.7–98.6% in amino acids between the two isolate strains and 20 reference strains. The two novel isolates showed 96.7% nucleotide identity and 98.1% amino acid identity. The phylogenetic analyses showed that the two isolates belonged to DWV Type A and were closely related to the KV-2001 strain from Japan. Based on the RDP4 and GARD analyses, the recombination of the China2-2018 strain was located at the 4,266–7,507 nt region, with Korea I-2012 as an infer unknown parent and China-2017 as a minor parent, which spanned the entire helicase ORF. To the best of our knowledge, this is the first study to the complete sequence of DWV isolated from Apis cerana and the possible DWV recombination events in China. Our findings are important for further research of the phylogenetic relationship of DWVs in China with DWV strains from other countries and also contribute to the understanding of virological properties of these complex DWV recombinants.
    Type of Medium: Online Resource
    ISSN: 2167-8359
    Language: English
    Publisher: PeerJ
    Publication Date: 2019
    detail.hit.zdb_id: 2703241-3
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