Trimerized S expressed by modified vaccinia virus Ankara (MVA) confers superior protection against lethal SARS-CoV-2 challenge in mice.

The reoccurrence of successive waves of SARS-CoV-2 variants suggests the exploration of more vaccine alternatives is imperative. Modified vaccinia virus Ankara (MVA) is a virus vector exhibiting excellent safety as well as efficacy for vaccine development. Here, a series of recombinant MVAs (rMVAs) expressing monomerized or trimerized S proteins from different SARS-CoV-2 variants are engineered. Trimerized S expressed from rMVAs is found predominantly as trimers on the surface of infected cells. Remarkably, immunization of mice with rMVAs demonstrates that S expressed in trimer elicits higher levels of binding IgG and IgA, as well as neutralizing antibodies for matched and mismatched S proteins than S in the monomer. In addition, trimerized S expressed by rMVA induces enhanced cytotoxic T-cell responses than S in the monomer. Importantly, the rMVA vaccines expressing trimerized S exhibit superior protection against a lethal SARS-CoV-2 challenge as the immunized animals all survive without displaying any pathological conditions. This study suggests that opting for trimerized S may represent a more effective approach and highlights that the MVA platform serves as an ideal foundation to continuously advance SARS-CoV-2 vaccine development. IMPORTANCE: MVA is a promising vaccine vector and has been approved as a vaccine for smallpox and mpox. Our analyses suggested that recombinant MVA expressing S in trimer (rMVA-ST) elicited robust cellular and humoral immunity and was more effective than MVA-S-monomer. Importantly, the rMVA-ST vaccine was able to stimulate decent cross-reactive neutralization against pseudoviruses packaged using S from different sublineages, including Wuhan, Delta, and Omicron. Remarkably, mice immunized with rMVA-ST were completely protected from a lethal challenge of SARS-CoV-2 without displaying any pathological conditions. Our results demonstrated that an MVA vectored vaccine expressing trimerized S is a promising vaccine candidate for SARS-CoV-2 and the strategy might be adapted for future vaccine development for coronaviruses.

Genomic analysis of lumpy skin disease virus asian variants and evaluation of its cellular tropism.

Lumpy skin disease virus (LSDV) is a poxvirus that mainly affects cattle and can lead to symptoms such as severe reduction in milk production as well as infertility and mortality, which has resulted in dramatic economic loss in affected countries in Africa, Europe, and Asia. In this study, we successfully isolated two strains of LSDV from different geographical regions in China. Comparative genomic analyses were performed by incorporating additional LSDV whole genome sequences reported in other areas of Asia. Our analyses revealed that LSDV exhibited an 'open' pan-genome. Phylogenetic analysis unveiled distinct branches of LSDV evolution, signifying the prevalence of multiple lineages of LSDV across various regions in Asia. In addition, a reporter LSDV expressing eGFP directed by a synthetic poxvirus promoter was generated and used to evaluate the cell tropism of LSDV in various mammalian and avian cell lines. Our results demonstrated that LSDV replicated efficiently in several mammalian cell lines, including human A549 cells. In conclusion, our results underscore the necessity for strengthening LSD outbreak control measures and continuous epidemiological surveillance.

The role of ferroptosis in virus infections.

Regulated cell death (RCD) is a strategy employed by host cells to defend invasions of pathogens, such as viruses and bacteria. Ferroptosis is a type of RCD characterized by excessive accumulation of iron and lipid peroxidation. While ferroptosis is primarily considered as a mechanism associated with tumorigenesis, emerging evidence begin to suggest that it may play essential role during virus infections. Recent studies illustrated that activation of ferroptosis could either induce or prohibit various types of RCDs to facilitate virus replication or evade host surveillance. More experimental evidence has demonstrated how viruses regulate ferroptosis to influence replication, transmission, and pathogenesis. This review summarizes ferroptosis-related metabolism, including iron metabolism, lipid peroxidation, and antioxidant metabolism. Furthermore, we discuss the interplay between viral infections and host ferroptosis process, with a focus on the mechanism of how viruses exploit ferroptosis for its own replication. Understanding how ferroptosis impacts virus infection can offer valuable insights into the development of effective therapeutic strategies to combat virus infections.

The FDA-approved anti-amyloid-ß monoclonal antibodies for the treatment of Alzheimer's disease: a systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: Alzheimer's disease (AD) is a worldwide public health problem and is difficult to cure. Drugs aimed at slowing the progression of the disease have been developed, with the Food and Drug Administration (FDA) granting accelerated approval for aducanumab on June 21, 2021 and a new accelerated approval for lecanemab on January 22, 2023. We performed this systematic review and meta-analysis to assess the efficacy and safety of FDA-approved anti-amyloid-ß (anti-Aß) monoclonal antibodies (mabs) for the treatment of AD. METHOD: PubMed, Embase, and Cochrane Library were systematically searched to identify relevant studies published before May 2023. Efficacy outcomes included Aß, neuroimaging, and biomarker outcomes. Safety outcomes included amyloid-related imaging abnormalities with edema or effusions (ARIA-E) and ARIA with cerebral microhemorrhages, cerebral macrohemorrhages, or superficial siderosis (ARIA-H). Review Manager 5.4 software was used to assess the data. The standard mean differences (SMDs) or odds ratio (OR) with 95% confidence interval (95% CI) were analyzed and calculated with a random effect model or a fixed effect model. RESULT: Overall, 4471 patients from 6 randomized controlled trials (RCTs), with 2190 patients in the treatment group and 2281 patients in the placebo group meeting the inclusion criteria. FDA-approved anti-Aß mabs showed statistically significant improvements in clinical outcomes, including CDR-SB (P = 0.01), ADCS-ADL-MCI (P = 0.00003), ADCOMS (P < 0.00001), ADAS-Cog (P < 0.00001). Moreover, FDA-approved anti-Aß mabs increased cerebrospinal fluid (CSF) Aß1-42 (P = 0.002) and plasma Aß42/40 ratios (P = 0.0008). They also decreased CSF P-Tau (P < 0.00001), CSF T-Tau (P < 0.00001), and plasma p-tau181 (P < 0.00001). FDA-approved anti-Aß mabs perform neuroimaging changes in amyloid Positron Emission Tomography Standardized Uptake Value ratio (PET SUVr) (P < 0.00001). However, compared with placebo, FDA-approved anti-Aß mabs had higher risk of ARIA-E (P < 0.00001) and ARIA-H (P < 0001). CONCLUSION: FDA-approved anti-Aß mabs have a role in slowing disease progression in patients with AD, at the cost of an increased probability of side effects.

Construction and Analysis of ceRNA Networks Reveal the Key Genes Associated with Bovine Herpesvirus Type 1 Infection.

Background: Virus infection can cause the changes of lncRNA expression levels to regulate the interaction between virus and host, but the relationship between BHV-1 infection and lncRNA has not been reported. Methods: In this study, in order to reveal the molecular mechanism of RNA in BoHV-1 infection, the Madin-Darby bovine kidney (MDBK) cells were infected with BoHV-1, transcriptome sequencing were performed by next-generation sequencing at 18 h or 24 h or 33 h of viral infection and then based on the competitive endogenous RNA (ceRNA) theory, lncRNA-miRNA-mRNA networks were constructed using these high-throughput sequencing data. The network analysis, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed for functional annotation and exploration of ncRNA ceRNAs in BoHV-1 infection. Results: The results showed that 48 lncRNAs, 123 mRNAs and 20 miRNAs as differentially expressed genes, and the mitogen activated protein kinase (MAPK) pathway and calcium signaling pathway were significantly enriched in the ceRNA network. Some differentially expressed lncRNA genes were randomly selected for verification by RT-qPCR, and the results showed that their expression trend was consistent with the results of transcriptome sequencing data. Conclusion: This study revealed that BoHV-1 infection can affect the expression of RNAs in MDBK cells and the regulation of ceRNA network to carry out corresponding biological functions in the host, but further experimental studies are still necessary to prove the hub genes function in ceRNA network and the molecular mechanism in BoHV-1 infection.

Human FAM111A inhibits vaccinia virus replication by degrading viral protein I3 and is antagonized by poxvirus host range factor SPI-1.

Zoonotic poxviruses such as mpox virus (MPXV) continue to threaten public health safety since the eradication of smallpox. Vaccinia virus (VACV), the prototypic poxvirus used as the vaccine strain for smallpox eradication, is the best-characterized member of the poxvirus family. VACV encodes a serine protease inhibitor 1 (SPI-1) conserved in all orthopoxviruses, which has been recognized as a host range factor for modified VACV Ankara (MVA), an approved smallpox vaccine and a promising vaccine vector. FAM111A (family with sequence similarity 111 member A), a nuclear protein that regulates host DNA replication, was shown to restrict the replication of a VACV SPI-1 deletion mutant (VACV-ΔSPI-1) in human cells. Nevertheless, the detailed antiviral mechanisms of FAM111A were unresolved. Here, we show that FAM111A is a potent restriction factor for VACV-ΔSPI-1 and MVA. Deletion of FAM111A rescued the replication of MVA and VACV-ΔSPI-1 and overexpression of FAM111A significantly reduced viral DNA replication and virus titers but did not affect viral early gene expression. The antiviral effect of FAM111A necessitated its trypsin-like protease domain and DNA-binding domain but not the PCNA-interacting motif. We further identified that FAM111A translocated into the cytoplasm upon VACV infection by degrading the nuclear pore complex via its protease activity, interacted with VACV DNA-binding protein I3, and promoted I3 degradation through autophagy. Moreover, SPI-1 from VACV, MPXV, or lumpy skin disease virus was able to antagonize FAM111A by prohibiting its nuclear export. Our findings reveal the detailed mechanism by which FAM111A inhibits VACV and provide explanations for the immune evasive function of VACV SPI-1.

High Microeukaryotic Diversity in the Cold-Seep Sediment.

Microeukaryotic diversity, community structure, and their regulating mechanisms remain largely unclear in chemosynthetic ecosystems. Here, using high-throughput sequencing data of 18S rRNA genes, we explored microeukaryotic communities from the Haima cold seep in the northern South China Sea. We compared three distinct habitats: active, less active, and non-seep regions, with vertical layers (0-25 cm) from sediment cores. The results showed that seep regions harbored more abundant and diverse parasitic microeukaryotes (e.g., Apicomplexa and Syndiniales) as indicator species, compared to nearby non-seep region. Microeukaryotic community heterogeneity was larger between habitats than within habitat, and greatly increased when considering molecular phylogeny, suggesting the local diversification in cold-seep sediments. Microeukaryotic α-diversity at cold seeps was positively increased by metazoan richness and dispersal rate of microeukaryotes, while its ß-diversity was promoted by heterogeneous selection mainly from metazoan communities (as potential hosts). Their combined effects led to the significant higher γ-diversity (i.e., total diversity in a region) at cold seeps than non-seep regions, suggesting cold-seep sediment as a hotspot for microeukaryotic diversity. Our study highlights the importance of microeukaryotic parasitism in cold-seep sediment and has implications for the roles of cold seep in maintaining and promoting marine biodiversity.

Evolutionary ecology of microbial populations inhabiting deep sea sediments associated with cold seeps.

Deep sea cold seep sediments host abundant and diverse microbial populations that significantly influence biogeochemical cycles. While numerous studies have revealed their community structure and functional capabilities, little is known about genetic heterogeneity within species. Here, we examine intraspecies diversity patterns of 39 abundant species identified in sediment layers down to 430 cm below the sea floor across six cold seep sites. These populations are grouped as aerobic methane-oxidizing bacteria, anaerobic methanotrophic archaea and sulfate-reducing bacteria. Different evolutionary trajectories are observed at the genomic level among these physiologically and phylogenetically diverse populations, with generally low rates of homologous recombination and strong purifying selection. Functional genes related to methane (pmoA and mcrA) and sulfate (dsrA) metabolisms are under strong purifying selection in most species investigated. These genes differ in evolutionary trajectories across phylogenetic clades but are functionally conserved across sites. Intrapopulation diversification of genomes and their mcrA and dsrA genes is depth-dependent and subject to different selection pressure throughout the sediment column redox zones at different sites. These results highlight the interplay between ecological processes and the evolution of key bacteria and archaea in deep sea cold seep extreme environments, shedding light on microbial adaptation in the subseafloor biosphere.

Rational design of near-infrared ratiometric fluorescent probes for real-time tracking of ß-galactosidase in vivo.

ß-Galactosidase (ß-gal) is a hydrolytic enzyme in lysosomes and is also an important biomarker of cellular senescence and primary ovarian cancer. Therefore, real-time non-invasive detection of ß-gal activity in vivo is of great significance for the prevention of cell senescence and early diagnosis of ovarian cancer. We designed an enzyme-activated proportional near-infrared (NIR) probe (Gal-Br-NO2) for real-time fluorescence quantification and capture of ß-gal activity in vivo. The main characteristics of the Gal-Br-NO2 probe include short response time (less than 10 min), large Stokes displacement (155 nm), and near-infrared fluorescence emission (670 nm). The probe has also been successfully used to detect ß-gal in ovarian cancer cells and senile cells and can accurately detect endogenous ß-gal in zebrafish. Our work provides a potential tool for pre-clinical real-time tracking of ß-gal activity in vivo and early diagnosis of disease.

The Efficacy and Safety of Anti-CD20 Antibody Treatments in Relapsing Multiple Sclerosis: A Systematic Review and Network Meta-analysis.

BACKGROUND: Several large randomized controlled trials of anti-CD20 antibodies have been successfully conducted for the treatment of relapsing multiple sclerosis. Despite this, there are few systematic comparisons of different anti-CD20 antibodies and a comprehensive evaluation of their efficacy and safety is yet to be carried out. OBJECTIVE: The objective of this systematic review and network meta-analysis was to evaluate the efficacy and safety of the three approved anti-CD20 antibodies for the treatment of relapsing multiple sclerosis and to aid clinicians in choosing medications. METHODS: MEDLINE, EMBASE, Cochrane Library, and clinicaltrials.gov were all searched for randomized controlled trials conducted to evaluate anti-CD20 antibodies (rituximab, ocrelizumab, ofatumumab) and corresponding controls up to 31 May, 2022. Review Manager 5.3 and R 3.5.2 software were used to assess the data. The risk ratio and mean difference were analyzed and calculated with a random-effects model. RESULTS: We pooled 4181 patients from ten randomized controlled trials. Without increasing the risk of adverse events and serious adverse events, anti-CD20 antibodies were superior to the active control group in all efficacy outcomes (both p < 0.005, certainty of evidence, very low to high). For the comparison between anti-CD20 groups, rituximab was found to be able to significantly increase the number of patients free of relapse more effectively than the other two interventions; however, the surface under curve ranking area values for serious adverse events were also the highest (84.8%). At the same time, ocrelizumab and ofatumumab exhibited satisfactory efficacy without showing a worse safety than any other interventions. CONCLUSIONS: Overall, anti-CD20 antibody treatment is superior to a corresponding control in efficacy and safety measures and ocrelizumab and ofatumumab may be the most suitable anti-CD20 antibodies for treating relapsing multiple sclerosis. Additional large-scale and high-quality studies are still needed to further explore the safety of these therapies.

Efficacy of Neurostimulations for Upper Extremity Function Recovery after Stroke: A Systematic Review and Network Meta-Analysis.

BACKGROUND: Neurostimulations for the post-stroke recovery of upper extremity function has been explored in previous research, but there remains a controversy about the superiority of different neurostimulations. METHODS: Randomized controlled trials (RCTs) were searched in MEDLINE, Embase, Cochrane Library and ClinicalTrials.gov, from 1 January 2000 to 1 June 2022. A conventional pair-wise meta-analysis with a random-effect model was used to evaluate direct evidence. Bayesian random effect models were used for network meta-analysis. The grading of the recommendations assessment, development and evaluation (GRADE) approach was applied to assess the clinical quality of the results. RESULTS: A total of 88 RCTs, which enrolled 3491 participants, were included. For the Fugl-Meyer Assessment-Upper Extremity score change from the baseline to the longest follow-up, the following interventions showed a significant difference: VNS (MD = 4.12, 95%CrI: 0.54 to 7.80, moderate certainty), cNMES (MD = 3.98, 95%CrI: 1.05 to 6.92, low certainty), FES (MD = 7.83, 95%CrI: 4.42 to 11.32, very low certainty), drTMS (MD = 7.94, 95%CrI: 3.71 to 12.07, moderate certainty), LFrTMS (MD = 2.64, 95%CrI: 1.20 to 4.11, moderate certainty), HFrTMS (MD = 6.73, 95%CrI: 3.26 to 10.22, moderate certainty), and iTBS combined with LFrTMS (MD = 5.41, 95%CrI: 0.48 to 10.35, moderate certainty). CONCLUSIONS: The neurostimulations above the revealed significant efficacy for improving the upper limb function after stroke eased the suffering of the patient.

Efficacy and Safety of Different Bioactive Coils in Intracranial Aneurysm Interventional Treatment, a Systematic Review and Bayesian Network Meta-Analysis.

BACKGROUND: Bioactive coils have been used for nearly 20 years to improve aneurysm treatments. Previous studies are inadequate for comparing the efficacy and safety between different coils. The aim of this study was to investigate the safety and efficacy of different coils by comparing the percentage of people with different modified Raymond scale grades, re-rupture rates, and mortality in patients with intracranial aneurysms embolized with different coils. METHOD: Randomized controlled trials (RCTs) containing coils for aneurysm interventional treatment were collected from Web of Science, PubMed, and the Cochrane Library up to December 2021. Bayesian network meta-analysis with a randomized or fixed model was performed to compare the efficacy and safety among different bioactive coils and bare platinum coils. RESULTS: We pooled 3362 patients from eight RCTs. No significant differences were found between coils in the proportion of patients with a three-grade classification assessed with the modified Raymond scale immediately after surgery. Hydrogel coils did not show a significant difference in the percentage of patients with a modified Raymond scale grade I postoperatively compared with bare platinum coils (OR, -0.1080; 95% CI, -0.4201-0.2423), but at follow-up, the percentage of patients with modified Raymond scale grade I was significantly higher with hydrogel coils than with bare platinum coils (OR, 0.4957; 95% CI, 0.0060-0.9442). There were no statistical differences between these four coils in terms of aneurysm rupture or re-rupture rate and mortality. CONCLUSION: Though there was no significant difference in the embolization effect between the several coils in the postoperative period, complete embolization was more likely to be achieved with hydrogel coils compared to bare platinum coils at follow-up. There were no significant differences in safety between the several coil materials.

Performance of deep learning in the detection of intracranial aneurysm: A systematic review and meta-analysis.

PURPOSE: Early detection and diagnosis of intracranial aneurysms (IAs) are particularly critical. Deep learning models (DLMs) are now widely used in the diagnosis of various diseases. Different DLMs have been developed to detect IAs. However, the overall performance of various DLMs for detecting IAs has not been evaluated. We aimed at exploring the performance of DLMs in the detection of IAs and measuring the effect of DLMs in assisting clinicians. METHODS: A diagnostic accuracy meta-analysis using a mixed-effect binary regression model was performed to estimate accuracy in patient-level and lesion-level. Moreover, the effect in assisting clinicians was measured by a random-effect meta-analysis. RESULTS: Twenty cohort studies including a total of 17 DLMs were assessed eligible in the present study. We summarized that DLMs had both high sensitivity (0.92, 95 % CI: 0.85 to 0.96) and specificity (0.96, 95 % CI: 0.94 to 0.97) in the detection of IAs in patient-level. In lesion-level, we also found a high summary sensitivity of 0.92 (95 % CI: 0.87 to 0.95). Moreover, assisted by DLMs, the sensitivity of clinicians became higher (Risk ratio: 1.09, P-value: 0.0006), with no effect on the specificity in diagnosis (Risk ratio: 0.99, P-value: 0.53). The reading time was reduced by the assistance of the deep learning model. (Mean difference: -7.37, P-value: 0.0077) CONCLUSIONS: DLMs have the competence of detecting IAs accurately. Moreover, DLMs can improve clinicians' sensitivity and reduce the reading time without affecting the specificity in diagnosing IAs.

Disentangling the Ecological Processes Shaping the Latitudinal Pattern of Phytoplankton Communities in the Pacific Ocean.

Phytoplankton diversity and community compositions vary across spaces and are fundamentally affected by several deterministic (e.g., environmental selection) and stochastic (e.g., ecological drift) processes. How this suite of different processes regulates the biogeography of phytoplankton remains to be comprehensively explored. Using high-throughput sequencing data and null model analysis, we revealed the ecological processes shaping the latitudinal community structure of three major phytoplankton groups (i.e., diatoms, Synechococcus, and haptophytes) across the Pacific Ocean (70°N, 170°W to 35°S, 170°W). At the basin scale, heterogeneous selection (selection under heterogeneous environmental conditions) dominated the assembly processes of all phytoplankton groups; however, its relative importance varied greatly at the climatic zonal scale, explaining the distinct latitudinal α- and ß-diversity among phytoplankton groups. Assembly processes in Synechococcus and haptophyte communities were mainly controlled by physical and nutrient factors, respectively. High temperature drove Synechococcus communities to be more deterministic with higher diversity, while haptophyte communities were less environmentally selected at low latitudes due to their wide niche breadth and mixotrophic lifestyle. Diatom communities were overwhelmingly dominated by the selection process but with low correlation of measured environmental factors to their community compositions. This could be attributed to the high growth rate of diatoms, as indicated by their lower site occupation frequency than predicted in the neutral community model. Our study showed that heterogeneous selection is the main force that shaped the biogeography of three key phytoplankton groups in the Pacific Ocean, with a latitudinal variation of relative importance due to the distinct traits among phytoplankton. IMPORTANCE Phytoplankton are diverse and abundant as primary producers in the ocean, with diversity and community compositions varying spatially. How fundamental processes (e.g., selection, dispersal, and drift) regulate their global biogeography remains to be comprehensively explored. In this study, we disentangled the ecological processes of three key phytoplankton groups (i.e., diatoms, Synechococcus, and haptophytes) along the same latitudinal gradients in the Pacific Ocean. Heterogeneous selection, by promoting species richness and reducing similarity between communities, was the dominant process shaping the communities of each phytoplankton group at the basin scale. However, its relative importance varied greatly among different phytoplankton groups in different climate zones, explaining the uneven latitudinal α- and ß-diversity. We also highlight the importance of identifying key factors mediating the relative importance of assembly processes in phytoplankton communities, which will enhance our understanding of their biogeography in the ocean and future patterns under climate changes.

Impact of sarcopenia on clinical outcomes of patients with stage I gastric cancer after radical gastrectomy: A prospective cohort study.

BACKGROUND: The relationships between sarcopenia and postoperative outcomes in patients with early-stage gastric cancer who undergo radical gastrectomy is unclear. We aimed to investigate the predictive value of sarcopenia on adverse outcomes for stage I gastric cancer. METHODS: The clinical data of patients who underwent radical gastrectomy for stage I gastric cancer between July 2013 and May 2019 were prospectively collected. Basic sarcopenia components were measured preoperatively. Univariate and multivariate analyses were conducted to evaluate the risk factors for short- and long-term outcomes. RESULTS: A total of 507 patients with early-stage gastric cancer were included in the study, and 73 (14.4%) patients were diagnosed as sarcopenia. Patients with sarcopenia had significantly higher incidence of postoperative complications (32.9% vs. 17.5%, P = 0.002), longer postoperative hospital stays (13 days vs. 12 days, P < 0.001), higher hospitalization costs (65210 yuan vs. 55197 yuan, P < 0.001) and one-year mortality (8.2% vs. 1.8%, P = 0.002). During the median follow-up time of 38.8 months, 12 (16.4%) patients dead in the sarcopenic group and 25 (5.8%) patients dead in the non-sarcopenic group. Sarcopenia was an independent risk factor for both short- and long-term clinical outcomes. Moreover, we found that low muscle quantity and low handgrip strength mediated the adverse impacts of sarcopenia on postoperative complications while low muscle quality mediated the adverse impacts of sarcopenia on overall survival. CONCLUSION: Sarcopenia was strongly associated with worse short- and long-term clinical outcomes in patients with stage I gastric cancer who undergo radical gastrectomy.

Establishment of a Suspension MDBK Cell Line in Serum-Free Medium for Production of Bovine Alphaherpesvirus-1.

The Madin-Darby bovine kidney (MDBK) cell line is currently used for the production of bovine alphaherpesvirus-1 (BoHV-1) vaccine. For the purpose of vaccine manufacturing, suspension cells are preferred over adherent ones due to simplified sub-cultivation and an easier scale-up process, both of which could significantly reduce production cost. This study aimed to establish a procedure for the culture of BoHV-1 in the suspended MDBK cell line in serum-free medium. We screened several commercially available serum-free media and chose ST503 for subsequent experiments. We successfully adapted the adherent MDBK cells to suspended growth in ST503 in the absence of serum. The maximum density of suspension-adapted MDBK cells could reach 2.5 × 107 cells/mL in ST503 medium with optimal conditions. The average size of suspension-adapted cells increased to 18 ± 1 µm from 16 ± 1 µm. Moreover, we examined tumorigenicity of the suspended cells and found no sign of tumorigenicity post adaptation. Next, we developed a protocol for the culture of BoHV-1 in the cell line described above and found that ultrasonic treatment could facilitate virus release and enhance virus yield by 11-fold, with the virus titer reaching 8.0 ± 0.2 log10TCID50/mL. Most importantly, the prototype inactivated BoHV-1 vaccine we generated using the suspension cultures of MDBK cells induced neutralizing antibodies to a titer comparable to that of the commercial inactivated BoHV-1 vaccine. Overall, we established and optimized a protocol for the production of inactivated BoHV-1 vaccine in MDBK cells adapted for suspension culture, which provides insights for future large-scale manufacturing of BoHV-1 vaccine.

A SNP-based GWAS and functional haplotype-based GWAS of flag leaf-related traits and their influence on the yield of bread wheat (Triticum aestivum L.).

KEY MESSAGE: The genetic architecture of five flag leaf morphology traits was dissected by the functional haplotype-based GWAS and a standard SNP-based GWAS in a diverse population consisting of 197 varieties. Flag leaf morphology (FLM) is a critical factor affecting plant architecture and grain yield in wheat. The genetic architecture of FLM traits has been extensively studied with QTL mapping in bi-parental populations, while few studies exploited genome-wide association studies (GWAS) in diverse populations. In this study, a panel of 197 elite and historical varieties from China was evaluated for five FLM traits including the length (FLL), width (FLW), ratio (FLR), area (FLA) and angle (FLANG) as well as yield in nine environments. Based on the phenotypic correlation between yield and FLL (-0.43), FLA (- 0.32) and FLW (0.11), an empirical FLM index combining the three FLM traits proved to be a good predictor for yield. Two GWAS approaches were applied to dissect the genetic architecture of five FLM traits with a Wheat660K SNP array. The functional haplotype-based GWAS revealed 6, 5 and 7 QTL for FLANG, FLL and FLR, respectively, whereas two QTL for FLW and one for FLR were identified by the standard SNP-based GWAS. Due to co-localization, there were 18 independent QTL and 10 of them were close to known ones. One co-localized QTL on chromosome 5A was associated with FLL, FLANG and FLR. Moreover, both GWAS approaches identified a novel QTL for FLR on chromosome 6B which was not reported in previous studies. This study provides new insights into the relationship between FLM and yield and broadens our understanding of the genetic architecture of FLM traits in wheat.

The application of aptamer Apt-236 targeting PvpA protein in the detection of antibodies against Mycoplasma gallisepticum.

Mycoplasma gallisepticum (M. gallisepticum) is the primary agent of chronic respiratory disease causing important economic losses in the poultry industry. Compared to antibodies, aptamers used to diagnose M. gallisepticum have many advantages, such as being chemically, animal-free produced and easily modifiable without affecting their affinity. Herein, a single-stranded DNA (ssDNA) aptamer Apt-236 which can specifically bind to PvpA protein of M. gallisepticum with a Kd of 1.30 ± 0.18 nM was selected successfully. An indirect blocking ELAA (ib-ELAA) for M. gallisepticum antibodies detection was also developed using Apt-236, in which M. gallisepticum antibodies would block the binding-position of aptamers. Therefor positive sera would prevent color development whereas negative sera will allow a strong color reaction. The ib-ELAA was consistent with other three widely used assays in terms of the growth and decline of the antibody response to M. gallisepticum, and showed substantial agreement with the results obtained using a commercial ELISA kit in clinical chicken sera samples. Therefore, the ib-ELAA developed in this study was a new format for aptamer application and would be an alternative method for the surveillance of M. gallisepticum.

MBOVPG45_0375 Encodes an IgG-Binding Protein and MBOVPG45_0376 Encodes an IgG-Cleaving Protein in Mycoplasma bovis.

Mycoplasma bovis is a significant bacterial pathogen which is able to persist in cattle and cause chronic diseases. This phenomenon may relate to M. bovis evading the immune system of the host. Immunoglobulin-binding proteins are widely distributed in a variety of pathogenic bacteria, including some Mycoplasma species. These proteins are considered to help the bacteria evade the immune response of the host. Here we found M. bovis strain PG45 can bind to IgG from several animals. MBOVPG45_0375 encodes a putative membrane protein, has strong amino acid sequence similarity with Immunoglobulin G-binding protein in Mycoplasma mycoides subsp. capri. Hence, we constructed recombinant MBOVPG45_0375 (r0375) in the Escherichia coli expression system and demonstrated that r0375 can bind to IgG non-immunologically rather than specific binding similar to interaction of antigen and antibody. Moreover, r0375 can bind to the Fab fragment of IgG. Also, the binding of r0375 and IgG inhibits the formation of antigen-antibody union. Furthermore, MBOVPG45_0376 encodes an IgG-cleaving protein of M. bovis strain PG45. Nevertheless, r0375 binding to IgG is required for the cleavage activity of recombinant 0376 (r0376). The activity of r0376 is also affected by incubation time and temperature. In addition, we found both MBOVPG45_0375 and MBOVPG45_0376 are membrane proteins of M. bovis strain PG45. These results about MBOVPG45_0375 as an IgG-binding protein and MBOVPG45_0376 as an IgG-cleaving protein offer a new insight into the interaction between M. bovis and its host.

Induction of Robust and Specific Humoral and Cellular Immune Responses by Bovine Viral Diarrhea Virus Virus-Like Particles (BVDV-VLPs) Engineered with Baculovirus Expression Vector System.

Bovine viral diarrhea virus (BVDV) is an important animal pathogen that affects cattle. Infections caused by the virus have resulted in substantial economic losses and outbreaks of BVDV are reported globally. Virus-like particles (VLPs) are promising vaccine technology largely due to their safety and strong ability to elicit robust immune responses. In this study, we developed a strategy to generate BVDV-VLPs using a baculovirus expression vector system (BEVS). We were able to assemble BVDV-VLPs composed of dimerized viral proteins E2 and Erns, and the VLPs were spherical particles with the diameters of about 50 nm. Mice immunized with 15 µg of VLPs adjuvanted with ISA201 elicited higher levels of E2-specific IgG, IgG1, and IgG2a antibodies as well as higher BVDV-neutralizing activity in comparison with controls. Re-stimulation of the splenocytes collected from mice immunized with VLPs led to significantly increased levels of CD3+CD4+T cells and CD3+CD8+T cells. In addition, the splenocytes showed dramatically enhanced proliferation and the secretion of Th1-associated IFN-γ and Th2-associated IL-4 compared to that of the unstimulated control group. Taken together, our data indicate that BVDV-VLPs efficiently induced BVDV-specific humoral and cellular immune responses in mice, showing a promising potential of developing BVDV-VLP-based vaccines for the prevention of BVDV infections.