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.

A new microsporidium, Triwangia caridinae gen. nov., sp. nov. parasitizing fresh water shrimp, Caridina formosae (Decapoda: Atyidae) in Taiwan.

A new microsporidium was isolated from the endemic, Taiwanese shrimp, Caridina formosae (Decapoda, Atyidae) from northern Taiwan. A conspicuous symptom of infection was presence of opaque white xenomas located in the proximity of the alimentary tract, the surface of the hepatopancreas, and the gills. A fully developed xenoma consisted of a hard, thick capsule filled with sporophorous vesicles containing multiple spores. Microsporidia developed synchronously within the same sporophorous vesicle, although the stage of parasite development differed among the vesicles. Fresh spores were pyriform, mononucleated and measured 6.53 × 4.38 µm. The polar filament was anisofilar with 9-11 coils. Phylogenetic analysis based on the small subunit ribosomal DNA sequence showed that the isolate is most similar to the fish microsporidian clade containing the genera Kabatana, Microgemma, Potaspora, Spraguea, and Teramicra. The highest sequence identity, 80%, was with Spraguea spp. Based on pathogenesis, life cycle and phylogenetic analysis, we erect a new genus and species, Triwangia caridinae for the novel microsporidium.

Genomic sequencing and analyses of Lymantria xylina multiple nucleopolyhedrovirus.

BACKGROUND: Outbreaks of the casuarina moth, Lymantria xylina Swinehoe (Lepidoptera: Lymantriidae), which is a very important forest pest in Taiwan, have occurred every five to 10 years. This moth has expanded its range of host plants to include more than 65 species of broadleaf trees. LyxyMNPV (L. xylina multiple nucleopolyhedrovirus) is highly virulent to the casuarina moth and has been investigated as a possible biopesticide for controlling this moth. LdMNPV-like virus has also been isolated from Lymantria xylina larvae but LyxyMNPV was more virulent than LdMNPV-like virus both in NTU-LY and IPLB-LD-652Y cell lines. To better understand LyxyMNPV, the nucleotide sequence of the LyxyMNPV DNA genome was determined and analysed. RESULTS: The genome of LyxyMNPV consists of 156,344 bases, has a G+C content of 53.4% and contains 157 putative open reading frames (ORFs). The gene content and gene order of LyxyMNPV were similar to those of LdMNPV, with 151 ORFs identified as homologous to those reported in the LdMNPV genome. Two genes (Lyxy49 and Lyxy123) were homologous to other baculoviruses, and four unique LyxyMNPV ORFs (Lyxy11, Lyxy19, Lyxy130 and Lyxy131) were identified in the LyxyMNPV genome, including a gag-like gene that was not reported in baculoviruses. LdMNPV contains 23 ORFs that are absent in LyxyMNPV. Readily identifiable homologues of the gene host range factor-1 (hrf-1), which appears to be involved in the susceptibility of L. dispar to NPV infection, were not present in LyxyMNPV. Additionally, two putative odv-e27 homologues were identified in LyxyMNPV. The LyxyMNPV genome encoded 14 bro genes compared with 16 in LdMNPV, which occupied more than 8% of the LyxyMNPV genome. Thirteen homologous regions (hrs) were identified containing 48 repeated sequences composed of 30-bp imperfect palindromes. However, they differed in the relative positions, number of repeats and orientation in the genome compared to LdMNPV. CONCLUSION: The gene parity plot analysis, percent identity of the gene homologues and a phylogenetic analysis suggested that LyxyMNPV is a Group II NPV that is most closely related to LdMNPV but with a highly distinct genomic organisation.

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.

Collagen IX is required for the integrity of collagen II fibrils and the regulation of vascular plexus formation in zebrafish caudal fins.

Capillary plexuses form during both vasculogenesis and angiogenesis and are remodeled into mature vessel types and patterns which are delicately orchestrated with the sizes and shapes of other tissues and organs. We isolated a zebrafish mutation named prp (for persistent plexus) that causes persistent formation of vascular plexuses in the caudal fins and consequent mispatterning of bony fin rays and the fin shape. Detailed analyses revealed that the prp mutation causes a significant reduction in the size and dramatic structural defects in collagen II-rich extracellular matrices called actinotrichia of both embryonic finfolds and adult fins. prp was mapped to chromosome 19 and found to encode the zebrafish collagen9alpha1 (col9alpha1) gene which is abundantly expressed in developing finfolds. A point mutation resulting in a leucine-to-histidine change was detected in the thrombospondin domain of the col9alpha1 gene in prp. Morpholino-mediated knockdown of col9alpha1 phenocopied the prp small-finfold phenotype in wild-type embryos, and an injection of plasmids containing the col9alpha1 cDNA into prp embryos locally restored the finfold size. Furthermore, we found that osteoblasts in prp mutants were mispatterned apparently following the abnormal vascular plexus pattern, demonstrating that blood vessels play an important role in the patterning of bony rays in zebrafish caudal fins.