Foot-and-mouth disease (FMD) is a highly contagious disease of livestock that causes severe economic loss in susceptible cloven-hoofed animals. Although the traditional inactivated vaccine has been proved effective, it may lead to a new outbreak of FMD because of either incomplete inactivation of FMDV or the escape of live virus from vaccine production workshop. Thus, it is urgent to develop a novel FMDV vaccine that is safer, more effective and more economical than traditional vaccines. A recombinant silkworm baculovirus Bm-P12A3C which contained the intact P1-2A and 3C protease coding regions of FMDV Asia 1/HNK/CHA/05 was developed. Indirect immunofluorescence test and sandwich-ELISA were used to verify that Bm-P12A3C could express the target cassette. Expression products from silkworm were diluted to 30 folds and used as antigen to immunize cattle. Specific antibody was induced in all vaccinated animals. After challenge with virulent homologous virus, four of the five animals were completely protected, and clinical symptoms were alleviated and delayed in the remaining one. Furthermore, a PD₅₀ (50% bovine protective dose) test was performed to assess the bovine potency of the subunit vaccine. The result showed the subunit vaccine could achieve 6.34 PD₅₀ per dose. The results suggest that this strategy might be used to develop the new subunit FMDV vaccine.

Background: The major royal jelly proteins/yellow (MRJP/YELLOW) family possesses several physiological and chemical functions in the development of Apis mellifera and Drosophila melanogaster. Each protein of the family has a conserved domain named MRJP. However, there is no report of MRJP/YELLOW family proteins in the Lepidoptera. Results: Using the YELLOW protein sequence in Drosophila melanogaster to BLAST silkworm EST database, we found a gene family composed of seven members with a conserved MRJP domain each and named it YELLOW protein family of Bombyx mori. We completed the cDNA sequences with RACE method. The protein of each member possesses a MRJP domain and a putative cleavable signal peptide consisting of a hydrophobic sequence. In view of genetic evolution, the whole Bm YELLOW protein family composes a monophyletic group, which is distinctly separate from Drosophila melanogaster and Apis mellifera. We then showed the tissue expression profiles of Bm YELLOW protein family genes by RT-PCR. Conclusion: A Bombyx mori YELLOW protein family is found to be composed of at least seven members. The low homogeneity and unique pattern of gene expression by each member among the family ensure us to prophesy that the members of Bm YELLOW protein family would play some important physiological functions in silkworm development.

The domestic silkworm, Bombyx mori, is a model insect with important economic value for silk production that also acts as a bioreactor for biomaterial production. The functional complexity of the silkworm transcriptome has not yet been fully elucidated, although genomic sequencing and other tools have been widely used in its study. We explored the transcriptome of silkworm at different developmental stages using high-throughput paired-end RNA sequencing. A total of about 3.3 gigabases (Gb) of sequence was obtained, representing about a 7-fold coverage of the B. mori genome. From the reads that were mapped to the genome sequence; 23,461 transcripts were obtained, 5,428 of them were novel. Of the 14,623 predicted protein-coding genes in the silkworm genome database, 11,884 of them were found to be expressed in the silkworm transcriptome, giving a coverage of 81.3%. A total of 13,195 new exons were detected, of which, 5,911 were found in the annotated genes in the Silkworm Genome Database (SilkDB). An analysis of alternative splicing in the transcriptome revealed that 3,247 genes had undergone alternative splicing. To help with the data analysis, a transcriptome database that integrates our transcriptome data with the silkworm genome data was constructed and is publicly available at http://124.17.27.136/gbrowse2/. To our knowledge, this is the first study to elucidate the silkworm transcriptome using high-throughput RNA sequencing technology. Our data indicate that the transcriptome of silkworm is much more complex than previously anticipated. This work provides tools and resources for the identification of new functional elements and paves the way for future functional genomics studies.

Newcastle disease virus (NDV) causes severe economic losses through severe morbidity and mortality and poses a significant threat to the global poultry industry. Significant efforts have been made to develop novel vaccines and therapeutics; however, the interaction of NDV with the host is not yet fully understood. Interferons (IFNs), an integral component of innate immune signaling, act as the first line of defense against invading viruses. Compared with the mammalian repertoire of IFNs, limited information is available on the antiviral potential of IFNs in chickens. Here, we expressed chicken IFN-γ (chIFN-γ) using a baculovirus expression vector system, characterized its antiviral potential against NDV, and determined its antiviral potential. Priming of chicken embryo fibroblasts with chIFN-γ elicited an antiviral environment in primary cells, which was mainly due to interferon-stimulated genes (ISGs). A genome-wide transcriptomics approach was used to elucidate the possible signaling pathways associated with IFN-γ-induced immune responses. RNA-sequencing (RNA-seq) data revealed significant induction of ISG-associated pathways, activated temporal expression of ISGs, antiviral mediators, and transcriptional regulators in a cascade of antiviral responses. Collectively, we found that IFN-γ significantly elicited an antiviral response against NDV infection. These data provide a foundation for chIFN-γ-mediated antiviral responses and underpin functional annotation of these important chIFN-γ-induced antiviral influencers.

Background: HearMNPV, a nucleopolyhedrovirus (NPV), which infects the cotton bollworm, Helicoverpa armigera, comprises multiple rod-shaped nucleocapsids in virion(as detected by electron microscopy). HearMNPV shows a different host range compared with H. armigera single-nucleocapsid NPV (HearSNPV). To better understand HearMNPV, the HearMNPV genome was sequenced and analyzed. Methods: The morphology of HearMNPV was observed by electron microscope. The qPCR was used to determine the replication kinetics of HearMNPV infectious for H. armigera in vivo. A random genomic library of HearMNPV was constructed according to the “partial filling-in” method, the sequence and organization of the HearMNPV genome was analyzed and compared with sequence data from other baculoviruses. Results: Real time qPCR showed that HearMNPV DNA replication included a decreasing phase, latent phase, exponential phase, and a stationary phase during infection of H. armigera. The HearMNPV genome consists of 154,196 base pairs, with a G + C content of 40.07%. 162 putative ORFs were detected in the HearMNPV genome, which represented 90.16% of the genome. The remaining 9.84% constitute four homologous regions and other non-coding regions. The gene content and gene arrangement in HearMNPV were most similar to those of Mamestra configurata NPV-B (MacoNPV-B), but was different to HearSNPV. Comparison of the genome of HearMNPV and MacoNPV-B suggested that HearMNPV has a deletion of a 5.4-kb fragment containing five ORFs. In addition, HearMNPV orf66, bro genes, and hrs are different to the corresponding parts of the MacoNPV-B genome. Conclusions: HearMNPV can replicate in vivo in H. armigera and in vitro, and is a new NPV isolate distinguished from HearSNPV. HearMNPV is most closely related to MacoNPV-B, but has a distinct genomic structure, content, and organization.

The 30 K proteins, the major group of hemolymph proteins in the silkworm, Bombyx mori (Lepidoptera: Bombycidae), are structurally related with molecular masses of ∼30 kDa and are involved in various physiological processes, e.g., energy storage, embryonic development, and immune responses. For this report, known 30 K protein gene sequences were used as Blastn queries against sequences in the B. mori transcriptome (SilkTransDB). Twenty-nine cDNAs ( Bm30K-1–29) were retrieved, including four being previously unidentified in the Lipoprotein_11 family. The genomic structures of the 29 genes were analyzed and they were mapped to their corresponding chromosomes. Furthermore, phylogenetic analysis revealed that the 29 genes encode three types of 30 K proteins. The members increased in each type is mainly a result of gene duplication with the appearance of each type preceding the differentiation of each species included in the tree. Real-Time Quantitative Polymerase Chain Reaction (Q-PCR) confirmed that the genes could be expressed, and that the three types have different temporal expression patterns. Proteins from the hemolymph was separated by SDS-PAGE, and those with molecular mass of ∼30 kDa were isolated and identified by mass spectrometry sequencing in combination with searches of various databases containing B. mori 30K protein sequences. Of the 34 proteins identified, 13 are members of the 30 K protein family, with one that had not been found in the SilkTransDB, although it had been found in the B. mori genome. Taken together, our results indicate that the 30 K protein family contains many members with various functions. Other methods will be required to find more members of the family.