Assessment of the immune interference effects of multivalent vaccine for influenza epidemic strain in 2022-2023 and evaluation of its efficacy.

The various strains of influenza virus cause respiratory symptoms in humans every year and annual vaccinations are recommended. Due to its RNA-type genes and segmented state, it belongs to a virus that mutates frequently with antigenic drift and shift, giving rise to various strains. Each year, the World Health Organization identifies the epidemic strains and operates a global surveillance system to suggest the viral composition for the influenza vaccine. Influenza viruses, which have multiple viral strains, are produced in the format of multivalent vaccine. However, the multivalent vaccine has a possibility of causing immune interference by introducing multiple strain-specific antigens in a single injection. Therefore, evaluating immune interference phenomena is essential when assessing multivalent vaccines. In this study, the protective ability and immunogenicity of multivalent and monovalent vaccines were evaluated in mice to assess immune interference in the multivalent vaccine. Monovalent and multivalent vaccines were manufactured using the latest strain of the 2022-2023 seasonal influenza virus selected by the World Health Organization. The protective abilities of both types of vaccines were tested through hemagglutination inhibition test. The immunogenicity of multivalent and monovalent vaccines were tested through enzyme-linked immunosorbent assay to measure the cellular and humoral immunity expression rates. As a result of the protective ability and immunogenicity test, higher level of virus neutralizing ability and greater amount of antibodies in both IgG1 and IgG2 were confirmed in the multivalent vaccine. No immune interference was found to affect the protective capacity and immune responses of the multivalent vaccines.

A comprehensive analysis integrating phenotypic assessment uncovering thornless cultivar lineages in Aralia elata.

Aralia elata is an Araliaceae woody plant species found in Northeastern Asia. To understand how genetic pools are distributed for A.elata clones, we were to analyze the population structure of A.elata cultivars and identify how these are correlated with thorn-related phenotype which determines the utility of A.elata. We found that the de novo assembled genome of 'Yeongchun' shared major genomic compartments with the public A.elata genome assembled from the wild-type from China. To identify the population structure of the 32 Korean and Japanese cultivars, we identified 44 SSR markers and revealed three main sub-clusters using ΔK analysis with one isolated cultivar. Machine-learning based clustering with thorn-related phenotype correlated moderately with population structure based on SSR analysis suggested multi-layered genetic regulation of thorn-related phenotypes. Thus, we revealed genetic lineage of A.elata and uncovered isolated cultivar which can provide new genetic material for further breeding.

Genetic characterization and pathogenicity in a mouse model of newly isolated bat-originated mammalian orthoreovirus in South Korea.

Mammalian orthoreoviruses (MRVs) infect a wide range of hosts, including humans, livestock, and wildlife. In the present study, we isolated a novel Mammalian orthoreovirus from the intestine of a microbat (Myotis aurascens) and investigated its biological and pathological characteristics. Phylogenetic analysis indicated that the new isolate was serotype 2, sharing the segments with those from different hosts. Our results showed that it can infect a wide range of cell lines from different mammalian species, including human, swine, and non-human primate cell lines. Additionally, media containing trypsin, yeast extract, and tryptose phosphate broth promoted virus propagation in primate cell lines and most human cell lines, but not in A549 and porcine cell lines. Mice infected with this strain via the intranasal route, but not via the oral route, exhibited weight loss and respiratory distress. The virus is distributed in a broad range of organs and causes lung damage. In vitro and in vivo experiments also suggested that the new virus could be a neurotropic infectious strain that can infect a neuroblastoma cell line and replicate in the brains of infected mice. Additionally, it caused a delayed immune response, as indicated by the high expression levels of cytokines and chemokines only at 14 days post-infection (dpi). These data provide an important understanding of the genetics and pathogenicity of mammalian orthoreoviruses in bats at risk of spillover infections.IMPORTANCEMammalian orthoreoviruses (MRVs) have a broad range of hosts and can cause serious respiratory and gastroenteritis diseases in humans and livestock. Some strains infect the central nervous system, causing severe encephalitis. In this study, we identified BatMRV2/SNU1/Korea/2021, a reassortment of MRV serotype 2, isolated from bats with broad tissue tropism, including the neurological system. In addition, it has been shown to cause respiratory syndrome in mouse models. The given data will provide more evidence of the risk of mammalian orthoreovirus transmission from wildlife to various animal species and the sources of spillover infections.

Self-Assembled Nanostructures Presenting Repetitive Arrays of Subunit Antigens for Enhanced Immune Response.

Infectious diseases pose persistent threats to public health, demanding advanced vaccine technologies. Nanomaterial-based delivery systems offer promising solutions to enhance immunogenicity while minimizing reactogenicity. We introduce a self-assembled vaccine (SAV) platform employing antigen-polymer conjugates designed to facilitate robust immune responses. The SAVs exhibit efficient cellular uptake by dendritic cells (DCs) and macrophages, which are crucial players in the innate immune system. The high-density antigen presentation of this SAV platform enhances the affinity for DCs through multivalent recognition, significantly augmenting humoral immunity. SAV induced high levels of immunoglobulin G (IgG), IgG1, and IgG2a, suggesting that mature DCs efficiently induced B cell activation through multivalent antigen recognition. Universality was confirmed by applying it to respiratory viruses, showcasing its potential as a versatile vaccine platform. Furthermore, we have also demonstrated strong protection against influenza A virus infection with SAV containing hemagglutinin, which is used in influenza A virus subunit vaccines. The efficacy and adaptability of this nanostructured vaccine present potential utility in combating infectious diseases.

A COVID-19 Vaccine for Dogs Prevents Reverse Zoonosis.

COVID-19 is caused by severe acute respiratory syndrome virus type 2 (SARS-CoV-2), which can infect both humans and animals. SARS-CoV-2 originated from bats and can affect various species capable of crossing the species barrier due to active mutation. Although reports on reverse zoonosis (human-to-animal transmission) of SARS-CoV-2 remain limited, reverse zoonosis has been reported in many species such as cats, tigers, minks, etc. Therefore, transmission to more animals cannot be ruled out. Moreover, the wide distribution of SARS-CoV-2 in the human population could result in an increased risk of reverse zoonosis. To counteract reverse zoonosis, we developed the first COVID-19 subunit vaccines for dogs, which are representative companion animals, and the vaccine includes the SARS-CoV-2 recombinant protein of whole S1 protein and the receptor-binding domain (RBD). A subunit vaccine is a vaccine developed by purifying only the protein region that induces an immune response instead of the whole pathogen. This type of vaccine is safer than the whole virus vaccine because there is no risk of infection and proliferation through back-mutation of the virus. Vaccines were administered to beagles twice at an interval of 3 weeks subcutaneously and antibody formation rates were assessed in serum. We identified a titer, comparable to that of vaccinated people, shown to be sufficient to protect against SARS-CoV-2. Therefore, the vaccination of companion animals, such as dogs, may prevent reverse zoonosis by protecting animals from SARS-CoV-2; thus, reverse zoonosis of COVID-19 is preventable.

Construction of a CRISPR/Cas9-Mediated Genome Editing System in Lentinula edodes.

CRISPR/Cas9 genome editing systems have been established in a broad range of eukaryotic species. Herein, we report the first method for genetic engineering in pyogo (shiitake) mushrooms (Lentinula edodes) using CRISPR/Cas9. For in vivo expression of guide RNAs (gRNAs) targeting the mating-type gene HD1 (LeA1), we identified an endogenous LeU6 promoter in the L. edodes genome. We constructed a plasmid containing the LeU6 and glyceraldehyde-3-phosphate dehydrogenase (LeGPD) promoters to express the Cas9 protein. Among the eight gRNAs we tested, three successfully disrupted the LeA1 locus. Although the CRISPR-Cas9-induced alleles did not affect mating with compatible monokaryotic strains, disruption of the transcription levels of the downstream genes of LeHD1 and LeHD2 was detected. Based on this result, we present the first report of a simple and powerful genetic manipulation tool using the CRISPR/Cas9 toolbox for the scientifically and industrially important edible mushroom, L. edodes.

Functional Analysis of a Novel ABL (Abnormal Browning Related to Light) Gene in Mycelial Brown Film Formation of Lentinula edodes.

Lentinula edodes is a globally important edible mushroom species that is appreciated for its medicinal properties as well as its nutritional value. During commercial cultivation, a mycelial brown film forms on the surface of the sawdust growth medium at the late vegetative stage. Mycelial film formation is a critical developmental process that contributes to the quantity and quality of the mushroom yield. However, little is known regarding the genetic underpinnings of brown film formation on the surface of mycelial tissue. A novel causal gene associated with the formation of the mycelial brown film, named ABL (Abnormal browning related to light), was identified in this study. The comparative genetic analysis by dihybrid crosses between normal and abnormal browning film cultivars demonstrated that a single dominant allele was responsible for the abnormal mycelium browning phenotype. Whole-genome sequencing analysis of hybrid isolates revealed five missense single-nucleotide polymorphisms (SNPs) in the ABL locus of individuals forming abnormal partial brown films. Additional whole-genome resequencing of a further 16 cultivars showed that three of the five missense SNPs were strongly associated with the abnormal browning phenotype. Overexpression of the dominant abl-D allele in a wild-type background conferred the abnormal mycelial browning phenotype upon transformants, with slender hyphae observed as a general defective mycelial growth phenotype. Our methodology will aid the future discovery of candidate genes associated with favorable traits in edible mushrooms. The discovery of a novel gene, ABL, associated with mycelial film formation will facilitate marker-associated breeding in L. edodes.

Analysis of Genetic Diversity and Population Structure of Wild Strains and Cultivars Using Genomic SSR Markers in Lentinula edodes.

In this study, the genetic diversity and the population structure of 77 wild strains and 23 cultivars of Lentinula edodes from Korea were analyzed using 20 genomic SSRs, and their genetic relationship was investigated. The tested strains of L. edodes were divided into three sub-groups consisting of only wild strains, mainly wild strains and several cultivars, and mainly cultivars and several wild strains by distance-based analysis. Using model-based analysis, L. edodes strains were divided into two subpopulations; the first one consisting of only wild strains and the second one with mainly cultivars and several wild strains. Moreover, AMOVA analysis revealed that the genetic variation in the cultivars was higher than that in the wild strains. The expected and observed heterozygosity and values indicating the polymorphic information content of L. edodes cultivars from Korea were also higher than that of the wild strains. Based on these results, we presume that the cultivars in Korea have developed by using numerous strains from other countries. In conclusion, the usage of wild strains for the development of new cultivars could improve the adaptability of L. edodes to biotic and abiotic stress.

Transcriptional network regulation of the brassinosteroid signaling pathway by the BES1-TPL-HDA19 co-repressor complex.

MAIN CONCLUSION: The brassinosteroid-related BES1 and BZR1 transcription factors dynamically modulate downstream gene networks via the TPL-HDA19 co-repressor complex in BR-signaling pathways in Arabidopsis thaliana. Brassinosteroids (BRs) are plant steroid hormones that are essential for diverse growth and developmental processes across the whole life cycle of plants. In Arabidopsis thaliana, the BR-related transcription factors BRI1-EMS-SUPPRESSOR 1 (BES1) and BRASSINAZOLE-RESISTANT 1 (BZR1) regulate a range of global gene expression in response to BR and several external signaling cues; however, the molecular mechanisms by which they mediate the reprogramming of downstream transcription remain unclear. We here report that formation of a protein complex between BES1 and BZR1 and Histone Deacetylase 19 (HDA19) via the conserved ERF-associated amphiphilic repression (EAR) motif proved essential for regulation of BR-signaling-related gene expression. Defects in BR-related functions of BES1 and BZR1 proteins containing a mutated EAR motif were completely rescued by artificial fusion with EAR-repression domain (SRDX), TOPLESS (TPL), or HDA19 proteins. RNA-sequencing analysis of Arabidopsis plants over-expressing bes1-DmEAR or bes1-DmEAR-HDA19 revealed an essential role for HDA19 activity in regulation of BES1/BZR1-mediated BR signaling. In addition to BR-related gene expression, the BES1-HDA19 transcription factor complex was important for abiotic stress-related drought stress tolerance and organ boundary formation. These results suggested that integrating activation of BR-signaling pathways with the formation of the protein complex containing BES1/BZR1 and TPL-HDA19 via the EAR motif was important in fine-tuning BR-related gene networks in plants.

Comparative transcriptome analysis identified candidate genes involved in mycelium browning in Lentinula edodes.

BACKGROUND: Lentinula edodes is one of the most popular edible mushroom species in the world and contains useful medicinal components, such as lentinan. The light-induced formation of brown film on the vegetative mycelial tissues of L. edodes is an important process for ensuring the quantity and quality of this edible mushroom. To understand the molecular mechanisms underlying this critical developmental process in L. edodes, we characterized the morphological phenotypic changes in a strain, Chamaram, associated with abnormal brown film formation and compared its genome-wide transcriptional features. RESULTS: In the present study, we performed genome-wide transcriptome analyses of different vegetative mycelium growth phenotypes, namely, early white, normal brown, and defective dark yellow partial brown films phenotypes which were exposed to different light conditions. The analysis revealed the identification of clusters of genes specific to the light-induced brown film phenotypes. These genes were significantly associated with light sensing via photoreceptors such as FMN- and FAD-bindings, signal transduction by kinases and GPCRs, melanogenesis via activation of tyrosinases, and cell wall degradation by glucanases, chitinases, and laccases, which suggests these processes are involved in the formation of mycelial browning in L. edodes. Interestingly, hydrophobin genes such as SC1 and SC3 exhibited divergent expression levels in the normal and abnormal brown mycelial films, indicating the ability of these genes to act in fruiting body initiation and formation of dikaryotic mycelia. Furthermore, we identified the up-regulation of glycoside hydrolase domain-containing genes in the normal brown film but not in the abnormal film phenotype, suggesting that cell wall degradation in the normal brown film phenotype is crucial in the developmental processes related to the initiation and formation of fruiting bodies. CONCLUSIONS: This study systematically analysed the expression patterns of light-induced browning-related genes in L. edodes. Our findings provide information for further investigations of browning formation mechanisms in L. edodes and a foundation for future L. edodes breeding.

The complete mitochondrial genome of the edible and phytopathogenic fungus Desarmillaria tabescens.

Desarmillaria tabescens is one of the most important edible, medicinal, and phytopathogenic basidiomycetes. The complete mitochondrial genome of this species was determined using next-generation sequencing technology. This mitogenome is a circular molecule of 93,439 bp with a GC content of 29.28% and contains 15 protein-coding, two rRNA (rnl and rns), and 24 tRNA genes. Phylogenetic analysis revealed that D. tabescens is genetically closest to Agrocybe aegerita. Desarmillaria tabescens mitogenome can contribute to our understanding of the phylogeny and evolution of this species.

Genome-wide transcriptomic analysis of BR-deficient Micro-Tom reveals correlations between drought stress tolerance and brassinosteroid signaling in tomato.

Brassinosteroids (BRs) are plant steroid hormones that play crucial roles in a range of growth and developmental processes. Although BR signal transduction and biosynthetic pathways have been well characterized in model plants, their biological roles in an important crop, tomato (Solanum lycopersicum), remain unknown. Here, cultivated tomato (WT) and a BR synthesis mutant, Micro-Tom (MT), were compared using physiological and transcriptomic approaches. The cultivated tomato showed higher tolerance to drought and osmotic stresses than the MT tomato. However, BR-defective phenotypes of MT, including plant growth and stomatal closure defects, were completely recovered by application of exogenous BR or complementation with a SlDWARF gene. Using genome-wide transcriptome analysis, 619 significantly differentially expressed genes (DEGs) were identified between WT and MT plants. Several DEGs were linked to known signaling networks, including those related to biotic/abiotic stress responses, lignification, cell wall development, and hormone responses. Consistent with the higher susceptibility of MT to drought stress, several gene sets involved in responses to drought and osmotic stress were differentially regulated between the WT and MT tomato plants. Our data suggest that BR signaling pathways are involved in mediating the response to abiotic stress via fine-tuning of abiotic stress-related gene networks in tomato plants.

Development and Molecular Characterization of Novel Polymorphic Genomic DNA SSR Markers in Lentinula edodes.

Sixteen genomic DNA simple sequence repeat (SSR) markers of Lentinula edodes were developed from 205 SSR motifs present in 46.1-Mb long L. edodes genome sequences. The number of alleles ranged from 3-14 and the major allele frequency was distributed from 0.17-0.96. The values of observed and expected heterozygosity ranged from 0.00-0.76 and 0.07-0.90, respectively. The polymorphic information content value ranged from 0.07-0.89. A dendrogram, based on 16 SSR markers clustered by the paired hierarchical clustering' method, showed that 33 shiitake cultivars could be divided into three major groups and successfully identified. These SSR markers will contribute to the efficient breeding of this species by providing diversity in shiitake varieties. Furthermore, the genomic information covered by the markers can provide a valuable resource for genetic linkage map construction, molecular mapping, and marker-assisted selection in the shiitake mushroom.

Development of 44 Novel Polymorphic SSR Markers for Determination of Shiitake Mushroom (Lentinula edodes) Cultivars.

The shiitake mushroom (Lentinulaedodes) is one of the most popular edible mushrooms in the world and has attracted attention for its value in medicinal and pharmacological uses. With recent advanced research and techniques, the agricultural cultivation of the shiitake mushroom has been greatly increased, especially in East Asia. Additionally, demand for the development of new cultivars with good agricultural traits has been greatly enhanced, but the development processes are complicated and more challenging than for other edible mushrooms. In this study, we developed 44 novel polymorphic simple sequence repeat (SSR) markers for the determination of shiitake mushroom cultivars based on a whole genome sequencing database of L. edodes. These markers were found to be polymorphic and reliable when screened in 23 shiitake mushroom cultivars. For the 44 SSR markers developed in this study, the major allele frequency ranged from 0.13 to 0.94; the number of genotypes and number of alleles were each 2-11; the observed and expected heterozygosity were 0.00-1.00 and 0.10-0.90, respectively; and the polymorphic information content value ranged from 0.10 to 0.89. These new markers can be used for molecular breeding, the determination of cultivars, and other applications.