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

Sacbrood virus (SBV) was the first identified bee virus and shown to cause serious epizootic infections in the population of Apis cerana in Taiwan in 2015. Herein, the whole genome sequences of SBVs in A. cerana and A. mellifera were decoded and designated AcSBV-TW and AmSBV-TW, respectively. The whole genomes of AcSBV-TW and AmSBV-TW were 8776 and 8885 bp, respectively, and shared 90% identity. Each viral genome encoded a polyprotein, which consisted of 2841 aa in AcSBV-TW and 2859 aa in AmSBV-TW, and these sequences shared 95% identity. Compared to 54 other SBVs, the structural protein and protease regions showed high variation, while the helicase was the most highly conserved region among SBVs. Moreover, a 17-amino-acid deletion was found in viral protein 1 (VP1) region of AcSBV-TW compared to AmSBV-TW. The phylogenetic analysis based on the polyprotein sequences and partial VP1 region indicated that AcSBV-TW was grouped into the SBV clade with the AC-genotype (17-aa deletion) and was closely related to AmSBV-SDLY and CSBV-FZ, while AmSBV-TW was grouped into the AM-genotype clade but branched independently from other AmSBVs, indicating that the divergent genomic characteristics of AmSBV-TW might be a consequence of geographic distance driving evolution, and AcSBV-TW was closely related to CSBV-FZ, which originated from China. This 17-amino-acid deletion could be found in either AcSBV or AmSBV in Taiwan, indicating cross-infection between the two viruses. Our data revealed geographic and host specificities between SBVs. The amino acid difference in the VP1 region might serve as a molecular marker for describing SBV cross-infection.

Complete Genome Sequence of a Novel Putative RNA Virus, RiPV-2, from the Bean Bug Riptortus pedestris.

A novel putative single-stranded RNA virus was discovered from the transcriptome of a bean bug, Riptortus pedestris, infected with the entomopathogenic fungus Beaveria bassiana JEF-007. The complete genome sequence was 9,915 nucleotides long and encoded a 2,916-amino-acid polyprotein. This virus belonged to Iflaviridae based on phylogenetic analysis and was named RiPV-2.

Genomic sequencing of Troides aeacus nucleopolyhedrovirus (TraeNPV) from golden birdwing larvae (Troides aeacus formosanus) to reveal defective Autographa californica NPV genomic features.

BACKGROUND: The golden birdwing butterfly (Troides aeacus formosanus) is a rarely observed species in Taiwan. Recently, a typical symptom of nuclear polyhedrosis was found in reared T. aeacus larvae. From the previous Kimura-2 parameter (K-2-P) analysis based on the nucleotide sequence of three genes in this isolate, polh, lef-8 and lef-9, the underlying virus did not belong to any known nucleopolyhedrovirus (NPV) species. Therefore, this NPV was provisionally named "TraeNPV". To understand this NPV, the nucleotide sequence of the whole TraeNPV genome was determined using next-generation sequencing (NGS) technology. RESULTS: The genome of TraeNPV is 125,477 bp in length with 144 putative open reading frames (ORFs) and its GC content is 40.45%. A phylogenetic analysis based on the 37 baculoviral core genes suggested that TraeNPV is a Group I NPV that is closely related to Autographa californica nucleopolyhedrovirus (AcMNPV). A genome-wide analysis showed that TraeNPV has some different features in its genome compared with other NPVs. Two novel ORFs (Ta75 and Ta139), three truncated ORFs (pcna, he65 and bro) and one duplicated ORF (38.7 K) were found in the TraeNPV genome; moreover, there are fewer homologous regions (hrs) than there are in AcMNPV, which shares eight hrs within the TraeNPV genome. TraeNPV shares similar genomic features with AcMNPV, including the gene content, gene arrangement and gene/genome identity, but TraeNPV lacks 15 homologous ORFs from AcMNPV in its genome, such as ctx, host cell-specific factor 1 (hcf-1), PNK/PNL, vp15, and apsup, which are involved in the auxiliary functions of alphabaculoviruses. CONCLUSIONS: Based on these data, TraeNPV would be clarified as a new NPV species with defective AcMNPV genomic features. The precise relationship between TraeNPV and other closely related NPV species were further investigated. This report could provide comprehensive information on TraeNPV for evolutionary insights into butterfly-infected NPV.

Mechanistic insight into the attenuation of gouty inflammation by Taiwanese green propolis via inhibition of the NLRP3 inflammasome.

Dysregulation of NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome is involved in many chronic inflammatory diseases, including gouty arthritis. Activation of the NLRP3 inflammasome requires priming and activation signals: the priming signal controls the expression of NLRP3 and interleukin (IL)-1ß precursor (proIL-1ß), while the activation signal leads to the assembly of the NLRP3 inflammasome and to caspase-1 activation. Here, we reported the effects of the alcoholic extract of Taiwanese green propolis (TGP) on the NLRP3 inflammasome in vitro and in vivo. TGP inhibited proIL-1ß expression by reducing nuclear factor kappa B activation and reactive oxygen species (ROS) production in lipopolysaccharide-activated macrophages. Additionally, TGP also suppressed the activation signal by reducing mitochondrial damage, ROS production, lysosomal rupture, c-Jun N-terminal kinases 1/2 phosphorylation and apoptosis-associated speck-like protein oligomerization. Furthermore, we found that TGP inhibited the NLRP3 inflammasome partially via autophagy induction. In the in vivo mouse model of uric acid crystal-induced peritonitis, TGP attenuated the peritoneal recruitment of neutrophils, and the levels of IL-1ß, active caspase-1, IL-6 and monocyte chemoattractant protein-1 in lavage fluids. As a proof of principle, in this study, we purified a known compound, propolin G, from TGP and identified this compound as a potential inhibitor of the NLRP3 inflammasome. Our results indicated that TGP might be useful for ameliorating gouty inflammation via inhibition of the NLRP3 inflammasome.

A novel picorna-like virus, Riptortus pedestris virus-1 (RiPV-1), found in the bean bug, R. pedestris, after fungal infection.

A viral genome was assembled de novo from next-generation sequencing (NGS) data from bean bugs, Riptortus pedestris, infected with an entomopathogenic fungus, Beauveria bassiana (Bb), and was further confirmed via the RACE method. This is a novel insect positive-sense single-stranded RNA virus, which we named Riptortus pedestris virus-1 (RiPV-1) (GenBank accession no. KU958718). The genome of RiPV-1 consists of 10,554 nucleotides (nt), excluding the poly(A) tail, which contains a single large open reading frame (ORF) of 10,371 nt encoding a polyprotein (3456 aa) and flanked by 71 and 112 nt at the 5' and 3' untranslated regions (UTR), respectively. RiPV-1 genome organization from the 5' end contains a consensus organization of picorna-like RNA virus helicase, cysteine protease, and RNA-dependent RNA polymerase (RdRp), in addition to two putative structural proteins located at the 3' region and a poly(A) tail at the 3' end. The viral particles were approximately 30nm in diameter with some dispersal distinctive surface projections. Based on the phylogenetic analysis of the RdRp sequences, RiPV-1 was clustered in the unassigned insect RNA viruses with two other viruses, APV and KFV. These three viruses were suggested to constitute a new group of insect RNA viruses. RiPV-1 could be found in all stages of lab-reared bean bugs and was detected abundantly in the thorax, abdomen, midgut and fat body, but not in the reproductive organs and muscle. Interestingly, RiPV-1 replication was increased dramatically in bean bugs 2-6days after fungal infection. In conclusion, a novel insect RNA virus was found by NGS data assembly. This virus can provide further insight into the interaction between virus, fungus and the host.

A new nucleopolyhedrovirus strain (LdMNPV-like virus) with a defective fp25 gene from Lymantria xylina (Lepidoptera: Lymantriidae) in Taiwan.

A new multiple nucleopolyhedrovirus strain was isolated from casuarina moth, Lymantria xylina Swinhoe, (Lepidoptera: Lymantriidae) in Taiwan. This Lymantria-derived virus can be propagated in IPLB-LD-652Y and NTU-LY cell lines and showed a few polyhedra (occlusion bodies) CPE in the infected cells. The restriction fragment length polymorphism (RFLP) profiles of whole genome indicated that this virus is distinct from LyxyMNPV and the virus genome size was approximately 139 kbps, which was smaller than that of LyxyMNPV. The molecular phylogenetic analyses of three important genes (polyhedrin, lef-8 and lef-9) were performed. Polyhedrin, LEF-8 and LEF-9 putative amino acid analyses of this virus revealed that this virus belongs to Group II NPV and closely related to LdMNPV than to LyxyMNPV. The phylogenetic distance analysis was further clarified the relationship to LdMNPV and this virus provisionally named LdMNPV-like virus. A significant deletion of a 44bp sequence found in LdMNPV-like virus was noted in the fp25k sequences of LdMNPV and LyxyMNPV and may play an important role in the few polyhedra CPE. In ultrastructural observations, the nuclei of the infected LD host cells contained large occlusion bodies (OBs), and few OBs, which presented as one or two OBs in a nucleus that was otherwise filled with free nuclocapsids and virions. We concluded that this LdMNPV-like virus is a new LdMNPV strain from L. xylina.