Genetic and phylogenetic analysis of Chinese sacbrood virus isolates fromApis mellifera

Abstract

BackgroundSacbrood 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).MethodsIn this study, a CSBV strain named AmCSBV-SDLY-2016 (GenBank accession No. ) 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.ResultsSequencing 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.

Genomic Sequencing and Comparison of Sacbrood Viruses from Apis cerana and Apis mellifera in Taiwan

“…The detection of AcSBV prevalence in A. mellifera populations from the sampling sites where A. cerana and A. mellifera were crossbreeding confirmed that AcSBV prevalence rates gradually developed a similar trend in the A. cerana and A. mellifera crossbreeding apiaries, and the existence of AcSBV cross-infection between A. cerana and A. mellifera was also confirmed by phylogenetic analysis based on partial VP1 sequences [33]. Similar to our case, some SBV strains from A. mellifera included in this study were found to be distinct from other AmSBV strains in terms of genomic features and were clustered with AcSBVs based on whole genome comparisons and phylogenetic analysis [24,30,36,40]. Strains from the same or closer geography distance showed higher similarity, and the phylogenetic analysis also indicated the same result [24,30,36,40,44].…”

“…Similar to our case, some SBV strains from A. mellifera included in this study were found to be distinct from other AmSBV strains in terms of genomic features and were clustered with AcSBVs based on whole genome comparisons and phylogenetic analysis [24,30,36,40]. Strains from the same or closer geography distance showed higher similarity, and the phylogenetic analysis also indicated the same result [24,30,36,40,44]. Indeed, it was shown that the cross-infection of SBV strains occurs between two honeybee species in other countries, including China, Vietnam, and Korea, leading to the high genetic divergence among SBV strains [24,30,36,40].…”

“…Strains from the same or closer geography distance showed higher similarity, and the phylogenetic analysis also indicated the same result [ 24 , 30 , 36 , 40 , 44 ]. Indeed, it was shown that the cross-infection of SBV strains occurs between two honeybee species in other countries, including China, Vietnam, and Korea, leading to the high genetic divergence among SBV strains [ 24 , 30 , 36 , 40 ].…”

“…It should be noted that deletions in the 712–730 amino acid (VP1) region of the ORF were found in most AcSBVs [ 24 , 30 , 40 ]. Since it has been mentioned that VP1 has the highest sequence variation among SBVs [ 24 , 30 , 40 , 48 ].…”

“…It should be noted that deletions in the 712–730 amino acid (VP1) region of the ORF were found in most AcSBVs [ 24 , 30 , 40 ]. Since it has been mentioned that VP1 has the highest sequence variation among SBVs [ 24 , 30 , 40 , 48 ]. Based on our comparisons, there are three types of deletion patterns: 17-amino-acid deletions, 10–13-amino-acid deletions, and deletions of less than 10 amino acids ( Figure 2 ).…”