BackgroundThe continuing spread of the newly emerged H7N9 virus among poultry in China, as well as the possibility of human-to-human transmission, has attracted numerous efforts to develop an effective vaccine against H7N9. The use of nanoparticles in vaccinology is inspired by the fact that most pathogens have a dimension within the nano-size range and therefore can be processed efficiently by the immune system, which leads to a potent immune response. Herein, we report a facile approach to increase antigen size to achieve not only fast but also effective responses against the recombinant HA/H7N9 protein via a simple conjugation of the protein onto the surface of nanodiamond particles.ResultsIn this study, trimeric Haemagglutinin (H7) that is transiently expressed in N. benthamiana was purified using affinity chromatography, and its trimeric state was revealed successfully by the cross-linking reaction. The trimeric H7 solution was subsequently mixed with a nanodiamond suspension in different ratios. The successful conjugation of the trimeric H7 onto the surface of nanodiamond particles was demonstrated by the changes in size and Zeta-potential of the particles before and after protein coating, Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and Western-blot analysis. Next, biofunction of the protein-nanodiamond conjugates was screened using a haemagglutination assay. A mixture containing 5 µg of trimeric H7 and 60 µg of nanodiamond corresponds to a ratio of 1:12 (w/w) of agglutinated chicken red blood cells at HA titer of 1024, which is 512-fold higher than the HA titer of free trimeric H7. After the 2nd and 3rd immunization in mice, ELISA and Western blot analyses demonstrated that the physical mixture of trimeric H7 protein and nanodiamond (1:12, w/w) elicited statistically significant stronger H7-specific-IgG response demonstrated by higher amounts of H7N9-specific IgG (over 15.4-fold with P < 0.05 after the second immunization).ConclusionsThese results indicated a potential effect inherent to nanodiamond towards modulating immune systems, which should be further evaluated and broadly applied in nanovaccine development.

Ascorbic acid (AsA) is an important antioxidant in plants and regulates various physiological processes. In this study, we show that exogenous treatments with the oxidized form of AsA, that is, dehydroascorbate (DHA), activates induced systemic resistance in rice against the root‐knot nematode Meloidogyne graminicola , and investigate the molecular and biochemical mechanisms underlying this phenotype. Detailed transcriptome analysis on roots of rice plants showed an early and robust transcriptional response on foliar DHA treatment, with induction of several genes related to plant stress responses, immunity, antioxidant activity, and secondary metabolism already at 1 day after treatment. Quantitative and qualitative evaluation of H 2 O 2 levels confirmed the appearance of a reactive oxygen species (ROS) burst on DHA treatment, both at the site of treatment and systemically. Experiments using chemical ROS inhibitors or scavengers confirmed that H 2 O 2 accumulation contributes to DHA‐based induced resistance. Furthermore, hormone measurements in DHA‐treated plants showed a significant systemic accumulation of the defence hormone salicylic acid (SA). The role of the SA pathway in DHA‐based induced resistance was confirmed by nematode infection experiments using an SA‐signalling deficient WRKY45 ‐RNAi line and reverse transcription‐quantitative PCR on SA marker genes. Our results collectively reveal that DHA activates induced systemic resistance in rice against the root‐knot nematode M. graminicola , mediated through the production of ROS and activation of the SA pathway.

Porcine epidemic diarrhea virus (PEDV) is a causative agent of a highly infectious disease with a high mortality rate, especially in newborn piglets in Asian countries resulting in serious economic loss. The development of a rapid, safe, effective and cost-efficient vaccine is crucial to protect pigs against PEDV infection. The COE antigen is regarded to be a major target for subunit vaccine development against PEDV infection. The naturally assembled COE protein forms a homotrimeric structure. In the present study, we successfully produced a trimeric COE protein as a native structure by fusion with the C-terminal isoleucine zipper trimerization (GCN4pII) motif in Nicotiana benthamiana, with a high expression level shown via semi-quantified Western blots. Trimeric COE protein was purified via immobilized metal affinity chromatography (IMAC), and its trimeric structure was successfully demonstrated by a cross-linking reaction, and a native PAGE gel. A crude extract containing the COE trimer was used for evaluating immunogenicity in mice. After 1 and 2 booster immunizations, the crude extract containing trimeric COE elicited elevated PEDV-specific humoral responses, as demonstrated by ELISA and Western blot analyses. Notably, a virus-neutralizing antibody assay indicated that the neutralization activities of sera of mice vaccinated with the crude extract containing COE-GCN4pII were similar to those of mice vaccinated with a commercial vaccine. These results suggest that crude extract containing trimeric COE is a promising plant-based subunit vaccine candidate for PEDV prevention.

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Pinus merkusii is an important species in Vietnam with many economic and biological contributions. The information on diversity within and between populations of a species is necessary for plantation programs, breeding and conservation strategies. Genetic diversity of three Vietnamese populations (NA, QB and QN) was analyzed using the random amplified polymorphic DNA (RAPD) markers. Nine RAPD primers produced 82 markers, 77 of which were polymorphic with 93.9% of polymorphism. The results showed higher genetic variation within populations (72%) than between populations (28%) and low Nei's genetic differentiation index among populations (0.1867). The populations also clustered based on PCoA analysis where cluster I included NA and QB populations and Cluster II, the QN population. These results suggest that P. merkusii populations in Vietnam is necessary to develop the genetic resources.