Search

The promotion and use of liquid feeding face the challenge of insufficiently stable delivery. This issue can be resolved, in part, by using the spiral flow produced by a spiral pipe (SPP). The aim of this study is to investigate how the structural characteristics of the spiral pipe affect the flow state of the liquid feed, and for this purpose, the computational fluid dynamics (CFD) technique has been employed and the liquid feed delivery process has been simulated by means of an Eulerian two-fluid model The results reveal a significant improvement in the slurry’s homogeneity as it traveled through a spiral pipe compared with a straight pipe (STP). The swirl number normally increased with the number, length, height, and angle of the spiral pipe’s guide vanes. The solid-phase distribution was more homogeneous when values of N = 1, L = 1D, H = 3/8R, and θ = 20° were used, respectively, and the COV within 10D downstream of the outlet of the spiral pipe was 3.902% smaller than that of the straight pipe. The results of this study can be used as a reference for the design of liquid feed-conveying pipes.

The plant-specific NAC gene family is one of the largest transcription factor families, participating in plant growth regulation and stress response. Despite extensive characterization in various plants, our knowledge of the NAC family in oat is lacking. Herein, we identified 333 NAC genes from the latest release of the common oat genome. We provide a comprehensive overview of the oat NAC gene family, covering gene structure, chromosomal localization, phylogenetic characteristics, conserved motif compositions, and gene duplications. AsNAC gene expression in different tissues and the response to various abiotic stresses were characterized using RT-qPCR. The main driver of oat NAC gene family expansion was identified as segmental duplication using collinearity analysis. In addition, the functions of AsNAC109 in regulating abiotic stress tolerance in Arabidopsis were clarified. This is the first genome-wide investigation of the NAC gene family in cultivated oat, which provided a unique resource for subsequent research to elucidate the mechanisms responsible for oat stress tolerance and provides valuable clues for the improvement of stress resistance in cultivated oat.

ABSTRACT Candidiasis is a life-threatening disease that increases mortality in critically ill patients. However, such epidemiological data are still lacking in underdeveloped regions of China. A retrospective analysis (2016 to 2021) was conducted in Meizhou People’s Hospital, China to study the burden of candidiasis, particularly candidemia, and antifungal susceptibilities of the species among hospitalized patients. Of the 7,864 candidiasis cases, 461 (5.86%) were candidemia cases. Candida albicans (64.25%) was the most identified species, followed by C. tropicalis (12.61%), C. glabrata (10.79%), and C. parapsilosis (9.79%). In non-C. albicans (NCA) candidemia cases, the number of C. glabrata cases was higher (102/461, 22.37%) than C. tropicalis (64/461, 14.04%). Gastrointestinal pathology, respiratory dysfunctions, septic shock, and malignancies were common underlying comorbidities, respectively. A central venous catheter was an independent risk factor for both C. albicans and NCA candidemia. The mortality rate was not statistically significant for either C. albicans or NCA. Amphotericin B and 5-flucytosine were highly effective (98 to 100%), while azoles were least effective (67.74 to 95.66%). Candidemia cases caused by C. tropicalis and C. glabrata had significantly lower azole susceptibility than non-candidemia-causing isolates. This study provides valuable information for prescribers to choose the right empirical therapy, for researchers to explore different resistance mechanisms, and for health care managers to control candidiasis better. IMPORTANCE This study provides important information on the burden of candidiasis, particularly candidemia, and the antifungal susceptibility of Candida species among hospitalized patients in an underdeveloped region of China. First, the finding that azoles were least effective against Candida species causing candidemia is particularly noteworthy, as it suggests the possibility of resistance to this class of antifungal agents. This information can guide the choice of empirical therapy and help in the selection of appropriate antifungal agents for the treatment of candidemia, thereby reducing the risk of resistance development. Second, the study provides important information for researchers to explore different resistance mechanisms in Candida species. Finally, the study has important implications for health care managers in controlling the spread of candidiasis. The high prevalence of candidemia cases in the study highlights the need for appropriate infection control measures to prevent the spread of the disease.

A novel indoor positioning method for a pigsty based on ultra-wideband (UWB) technology is proposed via reasonably arranging the base stations and positioning tags, which can provide strong support for the accurate positioning of specific pig pens in the pig house. The distribution of Dilution of Precision (DOP) values is analyzed in different deployment schemes which include the amount, the height, the deployment shape, and the distance of the base stations through positioning simulation. Moreover, the positioning effect of dynamic following positioning, the root-mean-square error (RMSE), and the variance of static positioning are measured. It is noteworthy that the Kalman filtering algorithm is adopted to smooth the dynamic following positioning experimental data for reducing the positioning error. The simulated results show that the DOP values decrease with the increase in height and distance, and these results and laws are consistent with the experimental results. However, the DOP values are best reached when the height is from 1.75 m to 2.75 m, and the distance is over 3.00 m. Furthermore, the DOP values of quadrilateral deployment are smaller than the ones of triangular deployment. In addition, the variance of static positioning and dynamic following positioning is decreased by utilizing the Kalman filtering algorithm. Therefore, the excellent positioning system and its solutions in a pigsty can be provided by our proposed indoor positioning method.

This study proposes a GC-YOLOv5s crack-detection network of UAVs to work out several issues, such as the low efficiency, low detection accuracy caused by shadows, occlusions and low contrast, and influences due to road noise in the classic crack-detection methods in the complicated traffic routes. A Focal-GIOU loss function with a focal loss has been introduced in this proposed algorithm, which is applied to address the issue of the imbalance of difficult and easy samples in crack images. Meanwhile, the original localization loss function CIOU is replaced by a GIOU loss function that is more suitable for irregular target (crack) detection. In order to improve the ability of the modified model of representing the features, a Transposed Convolution layer is simultaneously added in place of the original model’s upsampling layer. According to the advantage of computing resources of the Ghost module, the C3Ghost module is applied to decrease the amount of network parameters while maintaining adequate feature representation. Additionally, a lightweight module, CSPCM, is designed with the Conmix module and the Ghost concept, which successfully reduces the model parameters and zooms out the volume. At the same time, this modified module can have enough detection accuracy, and it can satisfy the requirements of UAV detection of small models and rapidity. In order to prove the model’s performance, this study has established a new UAV road-crack-detection dataset (named the UMSC), and has conducted extensive trials. To sum up, the precision of GC-YOLOv5s has increased by 8.2%, 2.8%, and 3.1%, respectively, and has reduced the model parameters by 16.2% in comparison to YOLOv5s. Furthermore, it outperforms previous YOLO comparison models in Precision, Recall, mAP_0.5, mAP_0.5:0.95, and Params.

This paper proposes an improved model based on YOLOv5s, specifically designed to overcome the challenges faced by current target detection algorithms in the field of electric bike helmet detection. In order to enhance the model’s ability to detect small targets and densely populated scenes, a specialized layer dedicated to small target detection and a novel loss function called Normalized Wasserstein Distance (NWD) are introduced. In order to solve the problem of increasing model parameters and complexity due to the inclusion of a small target detection layer, a Cross-Stage Partial Channel Mixing (CSPCM) on top of Convmix is designed. The collaborative fusion of CSPCM and the Deep Feature Consistency (DFC) attention mechanism makes it more suitable for hardware devices. In addition, the conventional Nearest Upsample technology is replaced with the advanced CARAFE Upsample module, further improving the accuracy of the model. Through rigorous experiments on carefully constructed datasets, the results show significant improvements in various evaluation indicators such as precision, recall, mAP.5, and mAP.95. Compared with the unmodified YOLOv5s algorithm, the proposed enhanced model achieves significant improvements of 1.1%, 8.4%, 5.2%, and 8.6% on these indicators, respectively, and these enhancements are accompanied by a reduction of 778,924 parameters. The experimental results on our constructed dataset demonstrate the superiority of the improved model and elucidate its potential applications. Furthermore, promising improvements for future research are suggested. This study introduces an efficient approach for improving the detection of electric bike helmets and verifies the effectiveness and practicality of the model through experiments. Importantly, the proposed scheme has implications for other target detection algorithms, especially in the field of small target detection.

Tobacco dust represents a critical threat to the operators’ respiratory health. Even when wet scrubbers display high purification efficiencies and strong adaptability to dust, their potential applications in the tobacco industry have not been studied. In the present research, removal efficiency of wet scrubber for tobacco dust and fine dust were examined at laboratory and industrial level. Results showed that the dust removal efficiency of the wet scrubber under laboratory conditions reached a maximum value of 99.17%. In addition, the dust discharge concentration was reduced to a minimum value of 1.1 mg/m3. Our data indicated that after using the wet scrubber, large dust particles (≥10 μm) were more effectively removed than fine dust particles (≤5 μm). After the use of the wet scrubber at the industrial facility, the overall dust removal efficiency reached a value of 98.9%, and dust concentration at the discharge point was as low as 1 mg/m3. This value complies with the ultra-low discharge standard and industrial production requirements. Our study demonstrated the feasibility of using wet scrubbers in the removal of tobacco dust and provides new insights for dust suppression in the tobacco processing industry.

In Atlantic salmon (Salmo salar L.), and also in other fish species, certain plant protein ingredients can increase fecal water content creating a diarrhea-like condition which may impair gut function and reduce fish growth. The present study aimed to strengthen understanding of the underlying mechanisms by observing effects of various alternative plant protein sources when replacing fish meal on expression of genes encoding proteins playing key roles in regulation of water transport across the mucosa of the distal intestine (DI). A 48-day feeding trial was conducted with five diets: A reference diet (FM) in which fish meal (72%) was the only protein source; Diet SBMWG with a mix of soybean meal (30%) and wheat gluten (22%); Diet SPCPM with a mix of soy protein concentrate (30%) and poultry meal (6%); Diet GMWG with guar meal (30%) and wheat gluten (14.5%); Diet PM with 58% poultry meal. Compared to fish fed the FM reference diet, fish fed the soybean meal containing diet (SBMWG) showed signs of enteritis in the DI, increased fecal water content of DI chyme and higher plasma osmolality. Altered DI expression of a battery of genes encoding aquaporins, ion transporters, tight junction and adherens junction proteins suggested reduced transcellular transport of water as well as a tightening of the junction barrier in fish fed the SBMWG diet, which may explain the observed higher fecal water content and plasma osmolality. DI structure was not altered for fish fed the other experimental diets but alterations in target gene expression and fecal water content were observed, indicating that alterations in water transport components may take place without clear effects on intestinal structure.

Abstract Influenza remains a severe respiratory illness that poses significant global health threats. Recent studies have identified distinct microbial communities within the respiratory tract, from nostrils to alveoli. This research explores specific anti-influenza respiratory microbes using a mouse model supported by 16S rDNA sequencing and untargeted metabolomics. The study found that transferring respiratory microbes from mice that survived H9N2 influenza to antibiotic-treated mice enhanced infection resistance. Notably, the levels of Aeromicrobium were significantly higher in the surviving mice. Mice pre-treated with antibiotics and then inoculated with Aeromicrobium camelliae showed reduced infection severity, as evidenced by decreased weight loss, higher survival rates, and lower lung viral titres. Metabolomic analysis revealed elevated LysoPE (16:0) levels in mildly infected mice. In vivo and in vitro experiments indicated that LysoPE (16:0) suppresses inducible nitric oxide synthase (INOS) and cyclooxygenase-2 (COX2) expression, enhancing anti-influenza defences. Our findings suggest that Aeromicrobium camelliae could serve as a potential agent for influenza prevention and a prognostic marker for influenza outcomes.

Abstract Background Chicken coccidiosis is a protozoan disease that leads to considerable economic losses in the poultry industry. Live oocyst vaccination is currently the most effective measure for the prevention of coccidiosis. However, it provides limited protection with several drawbacks, such as poor immunological protection and potential reversion to virulence. Therefore, the development of effective and safe vaccines against chicken coccidiosis is still urgently needed. Methods In this study, a novel oral vaccine against Eimeria tenella was developed by constructing a recombinant Lactobacillus plantarum (NC8) strain expressing the E. tenella RON2 protein. We administered recombinant L. plantarum orally at 3, 4 and 5 days of age and again at 17, 18 and 19 days of age. Meanwhile, each chick in the commercial vaccine group was immunized with 3 × 102 live oocysts of coccidia. A total of 5 × 104 sporulated oocysts of E. tenella were inoculated in each chicken at 30 days. Then, the immunoprotection effect was evaluated after E. tenella infection. Results The results showed that the proportion of CD4+ and CD8+ T cells, the proliferative ability of spleen lymphocytes, inflammatory cytokine levels and specific antibody titers of chicks immunized with recombinant L. plantarum were significantly increased (P

Coastal wetlands play a vital role in the migration and transformation of heavy metal pollutants in watersheds. There were 30 surface sediment samples that were analysed to investigate the distribution and ecological risks of heavy metals in the coastal wetlands around the Bohai Sea. Our results showed that the average concentrations of Pb, Cr, Ni, Cu, Zn, and Cd in these wetlands were 17.92 ± 5.81, 50.29 ± 20.50, 31.53 ± 9.71, 25.37 ± 4.29, 80.13 ± 15.11, and 0.92 ± 0.54 mg/kg, respectively. Relative to other wetlands, Pb (25.43 ± 2.68 mg/kg) and Cd (1.67 ± 0.06 mg/kg) contents were higher in the Liaohe Delta wetland (LHDW). Cu (28.44 ± 3.71 mg/kg), Cr (83.11 ± 5.80 mg/kg), and Ni (45.91 ± 3.02 mg/kg) contents were higher in the Yellow River Delta wetland (YRDW). The Zn (120.86 ± 7.41 mg/kg) content was higher in the Qilihai wetland. Heavy metal concentrations in coastal wetland sediments are shown to be positively correlated with organic matter content. Our results showed that the concentration of heavy metals decreases with increasing sediment particle size. In this study, Cd showed the highest pollution index and, therefore, more attention should be paid to the potential ecological risks of Cd in coastal wetlands around the Bohai Sea, especially in the LHDW and YRDW.

Avian influenza virus (AIV) subtype H9N2 still poses a great threat to the poultry farming industry and public health worldwide, and the development of a new influenza vaccine that is safe and conservative and able to address influenza virus mutations is highly promising for application. HA2, the neck of the HA protein, and M2e, the extracellular N-terminal structural domain of the M2 protein, are conserved and effective protective antigens.In this study, the HA2 sequences were fused with three M2e copies (H9N2, H1N1 and H5N1) to the norovirus VP1 protein via the SpyTag-SpyCatcher platform to form self-assembled nanoparticles and display antigenic proteins on its surface, yielding pYL262. The chicken dendritic cells (DCs) targeting the nanobody phage-54 were then fused to HA2-3M2e to yield pYL327. Finally, a synthesized 20-repeat CpG adjuvant gene fragment was inserted into pYL327, resulting in the plasmid pYL331. All the constructed plasmids were then transformed into the sifA gene-deficient Salmonella vector χYL56 for oral immunization.The results showed that sifA-deficient Salmonella could efficiently increase antigen-specific mucosal sIgA antibody titers, especially in alveolar lavage samples, whereas the presence of the phage-54 nanobody could dramatically increase intracellular IFN-γ mRNA levels, indicating its ability to enhance the Th1-type immune response. Finally, the presence of the CpG adjuvant clearly increased T-cell proliferation and promoted DC activation, with elevated splenic TLR21 levels observed. Strikingly, after oral immunization with χYL56 (pYL331), chickens were protected against challenge with the G57 genotype H9N2 virus, which presented similar or even better levels of virus shedding and body weight gain compared with the commercial inactivated vaccine, providing a new option for controlling H9N2 virus infection in the future.

ABSTRACT E3 ubiquitin ligase (E3) plays a vital role in regulating inflammatory responses by mediating ubiquitination. Previous studies have shown that ankyrin repeat and SOCS box-containing protein 3 (ASB3) is involved in immunomodulatory functions associated with cancer. However, the impact of ASB3 on the dynamic interplay of microbiota and inflammatory responses in inflammatory bowel disease (IBD) is unclear. Here, we systematically identify the E3 ligase ASB3 as a facilitative regulator in the development and progression of IBD. We observed that ASB3 exhibited significant upregulation in the lesions of patients with IBD. ASB3−/− mice are resistant to dextran sodium sulfate-induced colitis. IκBα phosphorylation levels and production of proinflammatory factors IL-1β, IL-6, and TNF-α were reduced in the colonic tissues of ASB3−/− mice compared to WT mice. This colitis-resistant phenotype was suppressed after coprophagic microbial transfer and reversed after combined antibiotics removed the gut commensal microbiome. Mechanistically, ASB3 specifically catalyzes K48-linked polyubiquitination of TRAF6 in intestinal epithelial cells. In contrast, in ASB3-deficient organoids, the integrity of the TRAF6 protein is shielded, consequently decelerating the onset of intestinal inflammation. ASB3 is associated with dysregulation of the colitis microbiota and promotes proinflammatory factors’ production by disrupting TRAF6 stability. Strategies to limit the protein level of ASB3 in intestinal epithelial cells may help in the treatment of colitis.IMPORTANCEUbiquitination is a key process that controls protein stability. We determined the ubiquitination of TRAF6 by ASB3 in intestinal epithelial cells during colonic inflammation. Inflammatory bowel disease patients exhibit upregulated ASB3 expression at focal sites, supporting the involvement of degradation of TRAF6, which promotes TLR-Myd88/TRIF-independent NF-κB aberrant activation and intestinal microbiota imbalance. Sustained inflammatory signaling in intestinal epithelial cells and dysregulated protective probiotic immune responses mediated by ASB3 collectively contribute to the exacerbation of inflammatory bowel disease. These findings provide insights into the pathogenesis of inflammatory bowel disease and suggest a novel mechanism by which ASB3 increases the risk of colitis. Our results suggest that future inhibition of ASB3 in intestinal epithelial cells may be a novel clinical strategy.

This paper investigates the odometry drift problem in differential-drive indoor mobile robots and proposes a multi-sensor fusion approach utilizing a Fuzzy Inference System (FIS) within a Wheel-Inertial-Visual Odometry (WIVO) framework to optimize the 6-DoF localization of the robot in unstructured scenes. The structure and principles of the multi-sensor fusion system are developed, incorporating an Iterated Error State Kalman Filter (IESKF) for enhanced accuracy. An FIS is integrated with the IESKF to address the limitations of traditional fixed covariance matrices in process and observation noise, which fail to adapt effectively to complex kinematic characteristics and visual observation challenges such as varying lighting conditions and unstructured scenes in dynamic environments. The fusion filter gains in FIS-IESKF are adaptively adjusted for noise predictions, optimizing the rule parameters of the fuzzy inference process. Experimental results demonstrate that the proposed method effectively enhances the localization accuracy and system robustness of differential-drive indoor mobile robots in dynamically changing movements and environments.

ABSTRACTEnterotoxigenic Escherichia coli (ETEC) is one of the major pathogens contributing to piglet diarrhea, with significant implications for both piglet health and the economic aspects of the livestock industry. SW207 is an isolate of Bacillus halotolerans isolated from the cold- and disease-resistant Leixiang pigs in Northeastern China. We have discovered that SW207 can survive in the pig’s gastrointestinal fluid and under conditions of high bile salt concentration, displaying potent antagonistic activity against ETEC. In this study, we established a weaned piglet diarrhea model infected with ETEC to investigate the role of SW207 in preventing diarrhea and improving intestinal health. Results indicate that SW207 upregulates the expression of tight junction proteins, including claudin-1, occludin, and zonula occludens-1, at both the transcriptional and translational levels. Furthermore, SW207 reduces serum endotoxin, D-lactic acid, and various oxidative stress markers while enhancing piglet mechanical barrier function. In terms of immune barrier, SW207 suppressed the activation of the TLR4/MyD88/NF-κB pathway, reducing the expression of various inflammatory factors and upregulating the expression of small intestine mucosal sIgA. Concerning the biological barrier, SW207 significantly reduces the content of E. coli in the intestines and promotes the abundance of beneficial bacteria, thereby mitigating the microbiota imbalance caused by ETEC. In summary, SW207 has the potential to prevent weaned piglet diarrhea caused by ETEC, alleviate intestinal inflammation and epithelial damage, and facilitate potential beneficial changes in the intestinal microbiota. This contributes to elucidating the potential mechanisms of host-microbe interactions in preventing pathogen infections.IMPORTANCEEnterotoxigenic Escherichia coli (ETEC) has consistently been one of the significant pathogens causing mortality in weaned piglets in pig farming. The industry has traditionally relied on antibiotic administration to control ETEC-induced diarrhea. However, the overuse of antibiotics has led to the emergence of drug-resistant zoonotic bacterial pathogens, posing a threat to public health. Therefore, there is an urgent need to identify alternatives to control pathogens and reduce antibiotic usage. In this study, we assessed the protective effect of a novel probiotic in a weaned piglet model infected with ETEC and analyzed its mechanisms both in vivo and in vitro. The study results provide theoretical support and reference for implementing interventions in the gut microbiota to alleviate early weaned piglet diarrhea and improve intestinal health.

ABSTRACTCanine distemper virus (CDV) poses a severe threat to both domesticated and wild animals, including multiple carnivores. With the continued expansion of its host range, there is an urgent need for the development of a safer and more effective vaccine. In this study, we developed subunit vaccines based on a bacterium-like particle (BLP) delivery platform containing BLPs-F and BLPs-H, which display the CDV F and H glycoprotein antigens, respectively, using the antigen-protein anchor fusions produced by a recombinant baculovirus insect cell expression system. The combination of BLPs-F and BLPs-H (CDV-BLPs), formulated with colloidal manganese salt [Mn jelly (MnJ)] adjuvant, triggered robust CDV-specific antibody responses and a substantial increase in the number of interferon gamma (IFN-γ)-secreting CD4+ and CD8+ T cells in mice. Dogs immunized intramuscularly with this vaccine not only produced CDV-specific IgG but also displayed elevated concentrations of IFN-γ and interleukin 6 in their serum, along with an increase of the CD3+CD4+ and CD3+CD8+ T cell subsets. Consequently, this heightened immune response provided effective protection against disease development and reduced viral shedding levels following challenge with a virulent strain. These findings suggest that this BLP-based subunit vaccine has the potential to become a novel canine distemper vaccine.IMPORTANCEMany sensitive species require a safe and effective distemper vaccine. Non-replicating vaccines are preferred. We constructed subunit particles displaying canine distemper virus (CDV) antigens based on a bacterium-like particle (BLP) delivery platform. The CDV-BLPs formulated with theMn jelly adjuvant induced robust humoral and cell-mediated immune responses to CDV in mice and dogs, thereby providing effective protection against a virulent virus challenge. This work is an important step in developing a CDV subunit vaccine.

Abstract Vaccination is still the most promising strategy for combating influenza virus pandemics. However, the highly variable characteristics of influenza virus make it difficult to develop antibody-based universal vaccines, until now. Lung tissue-resident memory T cells (TRM), which actively survey tissues for signs of infection and react rapidly to eliminate infected cells without the need for a systemic immune reaction, have recently drawn increasing attention towards the development of a universal influenza vaccine. We previously designed a sequential immunization strategy based on orally administered Salmonella vectored vaccine candidates. To further improve our vaccine design, in this study, we used two different dendritic cell (DC)-targeting strategies, including a single chain variable fragment (scFv) targeting the surface marker DC-CD11c and DC targeting peptide 3 (DCpep3). Oral immunization with Salmonella harboring plasmid pYL230 (S230), which displayed scFv-CD11c on the bacterial surface, induced dramatic production of spleen effector memory T cells (TEM). On the other hand, intranasal boost immunization using purified DCpep3-decorated 3M2e-ferritin nanoparticles in mice orally immunized twice with S230 (S230inDC) significantly stimulated the differentiation of lung CD11b+ DCs, increased intracellular IL-17 production in lung CD4+ T cells and elevated chemokine production in lung sections, such as CXCL13 and CXCL15, as determined by RNAseq and qRT‒PCR assays, resulting in significantly increased percentages of lung TRMs, which could provide efficient protection against influenza virus challenge. The dual DC targeting strategy, together with the sequential immunization approach described in this study, provides us with a novel “prime and pull” strategy for addressing the production of protective TRM cells in vaccine design.

Abstract Squeeze casting (SC) is an advanced net manufacturing process with many advantages for which the quality and properties of the manufactured parts depend strongly on the process parameters. Unfortunately, a universal efficient method for the determination of optimal process parameters is still unavailable. In view of the shortcomings and development needs of the current research methods for the setting of SC process parameters, by consulting and analyzing the recent research literature on SC process parameters and using the CiteSpace literature analysis software, manual reading and statistical analysis, the current state and characteristics of the research methods used for the determination of SC process parameters are summarized. The literature data show that the number of publications in the literature related to the design of SC process parameters generally trends upward albeit with significant fluctuations. Analysis of the research focus shows that both “mechanical properties” and “microstructure” are the two main subjects in the studies of SC process parameters. With regard to materials, aluminum alloys have been extensively studied. Five methods have been used to obtain SC process parameters: Physical experiments, numerical simulation, modeling optimization, formula calculation, and the use of empirical values. Physical experiments are the main research methods. The main methods for designing SC process parameters are divided into three categories: Fully experimental methods, optimization methods that involve modeling based on experimental data, and theoretical calculation methods that involve establishing an analytical formula. The research characteristics and shortcomings of each method were analyzed. Numerical simulations and model-based optimization have become the new required methods. Considering the development needs and data-driven trends of the SC process, suggestions for the development of SC process parameter research have been proposed.

The molecular mechanism of broad-spectrum neutralization of a rabbit monoclonal antibody 1H1 against Omicron sublineages is revealed by cryo-EM.

Generally, the interesting objects in aerial images are completely different from objects in nature, and the remote sensing objects in particular tend to be more distinctive in aspect ratio. The existing convolutional networks have equal aspect ratios of the receptive fields, which leads to receptive fields either containing non-relevant information or being unable to fully cover the entire object. To this end, we propose Horizontal and Vertical Convolution, which is a plug-and-play module to address different aspect ratio problems. In our method, we introduce horizontal convolution and vertical convolution to expand the receptive fields in the horizontal and vertical directions, respectively, to reduce redundant receptive fields, so that remote sensing objects with different aspect ratios can achieve better receptive fields coverage, thereby achieving more accurate feature representation. In addition, we design an attention module to dynamically aggregate these two sub-modules to achieve more accurate feature coverage. Extensive experimental results on the DOTA and HRSC2016 datasets show that our HVConv achieves accuracy improvements in diverse detection architectures and obtains SOTA accuracy (mAP score of 77.60% with DOTA single-scale training and mAP score of 81.07% with DOTA multi-scale training). Various ablation studies were conducted as well, which is enough to verify the effectiveness of our model.

Taking the gear assembly of the reducer assembly line as a research background, the hole-finding strategy for robot assembly with keyed circular pegs is proposed to solve the problem of low hole-finding efficiency and success rate. in the circular hole-finding task, deviation domains were divided based on a static mechanism, position vector trajectory optimization in the step distance and direction was designed, and a deviation domain-force mapping relationship was established using a genetic algorithm-support vector machine (GA-SVM) classification algorithm; the accuracy of this algorithm is approximately 90.00%. Thirty groups completed the circular hole-finding task in an average time of 5.4 s. For the square hole-finding task, dual monocular cameras were integrated to identify corner points of the flat key and keyway. Image semantic segmentation based on deep learning was used for the corner-point recognition of the flat key to suppress the effect of changes in light intensity; the recognition has an average error of 0.39 mm. Coarse and fine adjustment circumferential deflection strategies were adopted sequentially. The 30 groups of square hole-finding tasks exhibited a 96.70% success rate in an average time of 10.2 s. The proposed hole-finding strategy improves the efficiency and success rate of the gear assembly.

Abstract In this paper, we tackle the challenging problem of point cloud completion from the perspective of feature learning. Our key observation is that to recover the underlying structures as well as surface details, given partial input, a fundamental component is a good feature representation that can capture both global structure and local geometric details. We accordingly first propose FSNet, a feature structuring module that can adaptively aggregate point-wise features into a 2D structured feature map by learning multiple latent patterns from local regions. We then integrate FSNet into a coarse-to-fine pipeline for point cloud completion. Specifically, a 2D convolutional neural network is adopted to decode feature maps from FSNet into a coarse and complete point cloud. Next, a point cloud upsampling network is used to generate a dense point cloud from the partial input and the coarse intermediate output. To efficiently exploit local structures and enhance point distribution uniformity, we propose IFNet, a point upsampling module with a self-correction mechanism that can progressively refine details of the generated dense point cloud. We have conducted qualitative and quantitative experiments on ShapeNet, MVP, and KITTI datasets, which demonstrate that our method outperforms state-of-the-art point cloud completion approaches.

ABSTRACT The influenza virus continues to pose a great threat to public health due to the frequent variations in RNA viruses. Vaccines targeting conserved epitopes, such as the extracellular domain of the transmembrane protein M2 (M2e), a nucleoprotein, and the stem region of hemagglutinin proteins, have been developed, but more efficient strategies, such as nanoparticle-based vaccines, are still urgently needed. However, the labor-intensive in vitro purification of nanoparticles is still necessary, which could hinder the application of nanoparticles in the veterinary field in the future. To overcome this limitation, we used regulated lysis Salmonella as an oral vector with which to deliver three copies of M2e (3M2e-H1N1)-ferritin nanoparticles in situ and evaluated the immune response. Then, sequential immunization using Salmonella-delivered nanoparticles followed by an intranasal boost with purified nanoparticles was performed to further improve the efficiency. Compared with 3M2e monomer administration, Salmonella-delivered in situ nanoparticles significantly increased the cellular immune response. Additionally, the results of sequential immunization showed that the intranasal boost with purified nanoparticles dramatically stimulated the activation of lung CD11b dendritic cells (DCs) and elevated the levels of effector memory T (TEM) cells in both spleen and lung tissues as well as those of CD4 and CD8 tissue-resident memory T (TRM) cells in the lungs. The increased production of mucosal IgG and IgA antibody titers was also observed, resulting in further improvements to protection against a virus challenge, compared with the pure oral immunization group. Salmonella-delivered in situ nanoparticles efficiently increased the cellular immune response, compared with the monomer, and sequential immunization further improved the systemic immune response, as shown by the activation of DCs, the production of TEM cells and TRM cells, and the mucosal immune response, thereby providing us with a novel strategy by which to apply nanoparticle-based vaccines in the future. IMPORTANCE Salmonella-delivered in situ nanoparticle platforms may provide novel nanoparticle vaccines for oral administration, which would be beneficial for veterinary applications. The combination of administering Salmonella-vectored, self-assembled nanoparticles and an intranasal boost with purified nanoparticles significantly increased the production of effector memory T cells and lung resident memory T cells, thereby providing partial protection against an influenza virus challenge. This novel strategy could open a novel avenue for the application of nanoparticle vaccines for veterinary purposes.

Currently, most routes of the oil and gas pipelines in mountainous areas are selected in a traditional manual manner, with the quality greatly depending on the experience and technical level of the route selection personnel. Thus, the result of route selection is often not scientific and reasonable enough. Therefore, a route selection method of oil and gas pipelines in mountainous areas based on geological information atlas was put forward. Specifically, the regional geological information atlas is established in terms of landform, geological risk, high consequence area and sensitive area, the cost grid map for route selection through the quantification of factor cost is built with consideration to the routing cost of pipeline, and thus the optimal route is obtained through the analysis with optimal route algorithm. In addition, this method was applied to a pipeline in the mountainous area in Southwest China. By comparing the calculated route and the existing route, it is found that the route analyzed by this method of pipelines in mountainous areas based on the geological information atlas could effectively avoid the complex terrain, poor geological environment, high consequence area and environment sensitive area, and the distance cost could be considered comprehensively, thus solving the problem of inadequate consideration to the comprehensive factors in manual route selection. In general, the route selected by this method could be used as a reference for the pipeline route designers.

Global concerns arose as the emerged and rapidly spreading SARS-CoV-2 variants might escape host immunity induced by vaccination. In this study, a heterologous prime-boost immunization strategy for COVID-19 was designed to prime with a DNA vaccine encoding wild type (WT) spike protein receptor-binding domain (RBD) followed by S1 protein-based vaccine in rabbits. Four vaccine-elicited rabbit monoclonal antibodies (RmAbs), including 1H1, 9H1, 7G5, and 5E1, were isolated for biophysical property, neutralization potency and sequence analysis. All RmAbs recognized RBD or S1 protein with KD in the low nM or sub nM range. 1H1 and 9H1, but neither 7G5 nor 5E1, can bind to all RBD protein variants derived from B.1.351. All four RmAbs were able to neutralize wild type (WT) SARS-CoV-2 strain in pseudovirus assay, and 1H1 and 9H1 could neutralize the SARS-CoV-2 WT authentic virus with IC50 values of 0.136 and 0.026 μg/mL, respectively. Notably, 1H1 was able to neutralize all 6 emerging SARS-CoV-2 variants tested including D614G, B.1.1.7, B.1.429, P.1, B.1.526, and B.1.351 variants, and 5E1 could neutralize against the above 5 variants except P.1. Epitope binning analysis revealed that 9H1, 5E1 and 1H1 recognized distinct epitopes, while 9H1 and 7G5 may have overlapping but not identical epitope. In conclusion, DNA priming protein boost vaccination was an effective strategy to induce RmAbs with potent neutralization capability against not only SARS-CoV-2 WT strain but also emergent variants, which may provide a new avenue for effective therapeutics and point-of-care diagnostic measures.

In this article, the tensile strength, melting properties, and fiber-forming properties of natural continuous basalt fibers, such as andesite, andesite basalt, and tholeiitic basalt, are investigated as a function of their chemical and mineral compositions. The results indicate that when the basalt glass melts are homogeneous, the tensile strength is mainly determined by the chemical composition as well as the mineral components, which will affect the melting properties and fiber-forming properties. By optimizing the melting properties and fiber-forming properties, a high filament tensile strength of 3390 MPa was achieved for the basalt andesite with regions of tholeiitic basalt with an SiO2 weight percentage of 49%–57%.

Abstract Background Avian coccidiosis posts a severe threat to poultry production. In addition to commercial attenuated vaccines, other strategies to combat coccidiosis are urgently needed. Lactobacillus plantarum has been frequently used for expression of foreign proteins as an oral vaccine delivery system using traditional erythromycin resistance gene (erm). However, antibiotic selection markers were often used during protein expression and they pose a risk of transferring antibiotic resistance genes to the environment, and significantly restricting the application in field production. Therefore, a food-grade recombinant L. plantarum vaccine candidate would dramatically improve its application potential in the poultry industry. Results In this study, we firstly replaced the erythromycin resistance gene (erm) of the pLp_1261Inv-derived expression vector with a non-antibiotic, asd-alr fusion gene, yielding a series of non-antibiotic and reliable, food grade expression vectors. In addition, we designed a dual-expression vector that displayed two foreign proteins on the surface of L. plantarum using the anchoring sequences from either a truncated poly-γ-glutamic acid synthetase A (pgsA′) from Bacillus subtilis or the L. acidophilus surface layer protein (SlpA). EGFP and mCherry were used as marker proteins to evaluate the surface displayed properties of recombinant L. plantarum strains and were inspected by western blot, flow cytometry and fluorescence microscopy. To further determine its application as oral vaccine candidate, the AMA1 and EtMIC2 genes of E. tenella were anchored on the surface of L. plantarum strain. After oral immunization in chickens, the recombinant L. plantarum strain was able to induce antigen specific humoral, mucosal, and T cell-mediated immune responses, providing efficient protection against coccidiosis challenge. Conclusions The novel constructed food grade recombinant L. plantarum strain with double surface displayed antigens provides a potential efficient oral vaccine candidate for coccidiosis.

Unmanned surface vehicles (USV) is the development trend of future ships, and it will be widely used in various kinds of marine tasks. Obstacle avoidance is one key technology for autonomous navigation of USV. Convolutional neural network based obstacle classification and detection method is applied to USV visual images in environment sensing task. To solve the problem of low detection and classification accuracy of obstacles in the visual inspection of USV, a bidirectional feature pyramid networks is proposed combining hybrid network architecture of ResNet and improved DenseNet. The proposed method can further enhance the detection and classification some types of obstacles by using the underlying multi-layer detail features and high-level strong semantic features in the network architecture. The detection and classification performance of the proposed method is evaluated on a self built dataset. Ablation experiments and performance tests on open datasets are also employed. The experimental results show that the proposed algorithm has best performance for obstacles detection, and it is more suitable for autonomous navigation of USV.

The impact of the sintering temperature on the microstructure and characteristics of Al–Si composite was investigated in this study. Al-60Si (wt%) composites were sintered at different temperatures, and the microstructural features were evaluated. Optimal microstructural characteristics were observed when the composites were sintered at 850 °C for 2 h. The resulting material exhibited a thermal conductivity, coefficient of thermal expansion, and relative density of 131.4 W/(m·K), 10.02 × 10 ^–6 K ^–1 , and 99.87%, respectively. Elevating the temperature facilitated better wettability of the material, which in turn aided the filling of voids within the material by liquid Al. In addition, a decrease in the total number of interfaces within the material and the establishment of a connected network by the Al matrix contributed to enhanced thermal conductivity and coefficient of thermal expansion.

In this study, we electroplated Co and Cu on nano-spiked silicon substrates that were treated with femtosecond laser irradiations. With energy-dispersive X-ray (EDX) analysis by a scanning electron microscope (SEM), it was found that both Co and Cu are primarily coated on the spike surfaces without changing the morphology of the nanospikes. We also found that nanoscale bridges were formed, connecting the Co-coated silicon spikes. The formation of these bridges was studied and optimized through a series of time-controlled electroplating and oxidizing processes. The bridges are related to the oxidation of Co in the air. When it is irradiated with visible light, this special structure has shown a capability of interactions with carbon monoxide and carbon dioxide molecules. The electroplated cobalt may be used for gas sensors.

In order to study the vibration influence of tunnel drilling and blasting method on the built tunnel with small clear distance, taking the intersection of Zhuhai Dahengqinshan No. 1 tunnel and Zhuji urban rail tunnel as the engineering background, we used ABAQUS finite element software to conduct numerical simulation analysis on the influence of different blasting loads on existing tunnels with small clear distance in Zhuji tunnel construction. The following conclusions were drawn: the blasting construction of the tunnel under construction had the greatest impact on the vault of the existing tunnel; when the peak load was reduced by half, the stress value, vertical displacement, and resultant velocity of Mises were also reduced by half, which indicates that reducing the peak value of blasting load appropriately can ensure the safety of tunnel construction. When the peak load is 2.7 MPa, the measured and simulated values were less than the resultant velocity limit required by the specification. In addition, the relative error between the measured value and the simulated value was less than 5%, indicating the accuracy of the numerical simulation.

The Cre-recombinase mediated in vivo minicircle DNA vaccine platform (CRIM) provided a novel option to replace a traditional DNA vaccine. To further improve the immune response of our CRIM vaccine, we designed a dual promoter expression plasmid named pYL87 which could synthesize short HN protein under a prokaryotic in vivo promoter PpagC and full length HN protein of genotype VII Newcastle disease virus (NDV) under the previous eukaryotic CMV promoter at the same time. Making use of the self-lysed Salmonella strain as a delivery vesicle, chickens immunized with the pYL87 construction showed an increased serum haemagglutination inhibition antibody response, as well as an increased cell proliferation level and cellular IL-4 and IL-18 cytokines, compared with the previous CRIM vector pYL47. After the virus challenge, the pYL87 vector could provide 80% protection compared to 50% protection against genotype VII NDV in pYL47 immunized chickens, indicating a promising dual promoter strategy used in vaccine design.

ABSTRACT Development of a vaccine to protect against cryptococcosis is a priority given the enormous global burden of disease in at-risk individuals. Using glucan particles (GPs) as a delivery system, we previously demonstrated that mice vaccinated with crude Cryptococcus-derived alkaline extracts were protected against lethal challenge with Cryptococcus neoformans and Cryptococcus gattii. The goal of the present study was to identify protective protein antigens that could be used in a subunit vaccine. Using biased and unbiased approaches, six candidate antigens (Cda1, Cda2, Cda3, Fpd1, MP88, and Sod1) were selected, recombinantly expressed in Escherichia coli, purified, and loaded into GPs. Three mouse strains (C57BL/6, BALB/c, and DR4) were then vaccinated with the antigen-laden GPs, following which they received a pulmonary challenge with virulent C. neoformans and C. gattii strains. Four candidate vaccines (GP-Cda1, GP-Cda2, GP-Cda3, and GP-Sod1) afforded a significant survival advantage in at least one mouse model; some vaccine combinations provided added protection over that seen with either antigen alone. Vaccine-mediated protection against C. neoformans did not necessarily predict protection against C. gattii. Vaccinated mice developed pulmonary inflammatory responses that effectively contained the infection; many surviving mice developed sterilizing immunity. Predicted T helper cell epitopes differed between mouse strains and in the degree to which they matched epitopes predicted in humans. Thus, we have discovered cryptococcal proteins that make promising candidate vaccine antigens. Protection varied depending on the mouse strain and cryptococcal species, suggesting that a successful human subunit vaccine will need to contain multiple antigens, including ones that are species specific. IMPORTANCE The encapsulated fungi Cryptococcus neoformans and Cryptococcus gattii are responsible for nearly 200,000 deaths annually, mostly in immunocompromised individuals. An effective vaccine could substantially reduce the burden of cryptococcosis. However, a major gap in cryptococcal vaccine development has been the discovery of protective antigens to use in vaccines. Here, six cryptococcal proteins with potential as vaccine antigens were expressed recombinantly and purified. Mice were then vaccinated with glucan particle preparations containing each antigen. Of the six candidate vaccines, four protected mice from a lethal cryptococcal challenge. However, the degree of protection varied as a function of mouse strain and cryptococcal species. These preclinical studies identify cryptococcal proteins that could serve as candidate vaccine antigens and provide a proof of principle regarding the feasibility of protein antigen-based vaccines to protect against cryptococcosis.

Aspergillus fumigatus causes invasive pulmonary disease in immunocompromised hosts and allergic asthma in atopic individuals. We studied the contribution of lung eosinophils to these fungal diseases. By in vivo intracellular cytokine staining and confocal microscopy, we observed that eosinophils act as local sources of IL-23 and IL-17. Remarkably, mice lacking eosinophils had a >95% reduction in the percentage of lung IL-23p19+ cells as well as markedly reduced IL-23 heterodimer in lung lavage fluid. Eosinophils killed A. fumigatus conidia in vivo. Eosinopenic mice had higher mortality rates, decreased recruitment of inflammatory monocytes, and decreased expansion of lung macrophages after challenge with conidia. All of these functions underscore a potential protective role for eosinophils in acute aspergillosis. Given the postulated role for IL-17 in asthma pathogenesis, we assessed whether eosinophils could act as sources of IL-23 and IL-17 in models where mice were sensitized to either A. fumigatus antigens or ovalbumin (OVA). We found IL-23p19+ IL-17AF+ eosinophils in both allergic models. Moreover, close to 95% of IL-23p19+ cells and >90% of IL-17AF+ cells were identified as eosinophils. These data establish a new paradigm in acute and allergic aspergillosis whereby eosinophils act not only as effector cells but also as immunomodulatory cells driving the IL-23/IL-17 axis and contributing to inflammatory cell recruitment.

A series of silicate glass fibers with different ratios of Fe2O3/ZrO2 were prepared, and their corrosion resistance, mass loss, and strength loss were characterized. The crystallization and melting properties of the fibers were analyzed by differential scanning calorimetry (DSC), high temperature viscometer, and high temperature microscope. The results show that the deformation temperature, sphere temperature, hemisphere temperature, and crystallization temperature of the fiber initially decrease and then increase with the increase of Fe2O3/ZrO2 ratio, while the molding temperature decreases with the increase of the ratio of Fe2O3/ZrO2. When the ratio is close to 1 : 1, its alkali resistance is almost same as that of AR-glass fiber, and the drawing process performance is better. However, with the increase of the ratio, its alkali resistance continues to decline and the poor wire drawing performance is not conducive to the drawing operation.

A novel low-temperature SnO2 gas sensor was prepared and studied on silicon nanostructures formed by femtosecond laser irradiation. By applying a bias voltage on the silicon substrate to alter the charge distribution on the surface of the SnO2, carbon monoxide (CO), and ammonia (NH3) gas can be distinguished by the same sensor at room temperature. The experimental results are explained with a mechanism that the sensor works at low temperature because of adsorption of gas molecules that trap electrons to the surface of the SnO2.

A novel fabrication method for surface-enhanced Raman scattering (SERS) sensors that used a fast femtosecond (fs) laser scanning process to etch uniform patterns and structures on the endface of a fused silica optical fiber, which is then coated with a thin layer of silver through thermal evaporation is presented. A high quality SERS signal was detected on the patterned surface using a Rhodamine 6G (Rh6G) solution. The uniform SERS sensor built on the tip of the optical fiber tip was small, light weight, and could be especially useful in remote sensing applications.

ABSTRACT A vaccine capable of protecting at-risk persons against infections due to Cryptococcus neoformans and Cryptococcus gattii could reduce the substantial global burden of human cryptococcosis. Vaccine development has been hampered though, by lack of knowledge as to which antigens are immunoprotective and the need for an effective vaccine delivery system. We made alkaline extracts from mutant cryptococcal strains that lacked capsule or chitosan. The extracts were then packaged into glucan particles (GPs), which are purified Saccharomyces cerevisiae cell walls composed primarily of β-1,3-glucans. Subcutaneous vaccination with the GP-based vaccines provided significant protection against subsequent pulmonary infection with highly virulent strains of C. neoformans and C. gattii. The alkaline extract derived from the acapsular strain was analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS), and the most abundant proteins were identified. Separation of the alkaline extract by size exclusion chromatography revealed fractions that conferred protection when loaded in GP-based vaccines. Robust Th1- and Th17-biased CD4+ T cell recall responses were observed in the lungs of vaccinated and infected mice. Thus, our preclinical studies have indicated promising cryptococcal vaccine candidates in alkaline extracts delivered in GPs. Ongoing studies are directed at identifying the individual components of the extracts that confer protection and thus would be promising candidates for a human vaccine. IMPORTANCE The encapsulated yeast Cryptococcus neoformans and its closely related sister species, Cryptococcus gattii, are major causes of morbidity and mortality, particularly in immunocompromised persons. This study reports on the preclinical development of vaccines to protect at-risk populations from cryptococcosis. Antigens were extracted from Cryptococcus by treatment with an alkaline solution. The extracted antigens were then packaged into glucan particles, which are hollow yeast cell walls composed mainly of β-glucans. The glucan particle-based vaccines elicited robust T cell immune responses and protected mice from otherwise-lethal challenge with virulent strains of C. neoformans and C. gattii. The technology used for antigen extraction and subsequent loading into the glucan particle delivery system is relatively simple and can be applied to vaccine development against other pathogens.

Based on experimental results, we propose a mechanism that allows the use of metal nanostructures to synthesize hydrocarbons and carbohydrates from carbon dioxide, water and sunlight. When sunlight impinges on cobalt nanostructures in a glass chamber, its intensity is greatly enhanced around the tips of the nanostructures through surface plasmon excitations focusing effect, and it then photodissociates the water and carbon dioxide molecules through enhanced photon absorptions of ions around the tips of the nanostructures. The photodissociated molecules in excited states remain on the cobalt nanostructure surfaces and various hydrocarbons and carbohydrates then will be formed around the surfaces at temperatures much lower than 100 oC.

ABSTRACT Fungal cell walls are predominantly composed of glucans, mannans, and chitin. Recognition of these glycans by the innate immune system is a critical component of host defenses against the mycoses. Complement, an important arm of innate immunity, plays a significant role in fungal pathogenesis, especially the alternative pathway (AP). Here we determine that the glycan monosaccharide composition and glycosidic linkages affect AP activation and C3 deposition. Furthermore, properdin, a positive regulator of the AP, contributes to these functions. AP activation by glycan particles that varied in composition and linkage was measured by C3a generation in serum treated with 10 mM EGTA and 10 mM Mg2+ (Mg-EGTA-treated serum) (AP specific; properdin functional) or Mg-EGTA-treated serum that lacked functional properdin. Particles that contained either β1→3 or β1→6 glucans or both generated large and similar amounts of C3a when the AP was intact. Blocking properdin function resulted in 5- to 10-fold-less C3a production by particulate β1→3 glucans. However, particulate β1→6 glucans generated C3a via the AP only in the presence of intact properdin. Interestingly, zymosan and glucan-mannan particles (GMP), which contain both β-glucans and mannans, also required properdin to generate C3a. The β1→4 glycans chitin and chitosan minimally activated C3 even when properdin was functional. Finally, properdin binding to glucan particles (GP) and zymosan in serum required active C3. Properdin colocalized with bound C3, suggesting that in the presence of serum, properdin bound indirectly to glycans through C3 convertases. These findings provide a better understanding of how properdin facilitates AP activation by fungi through interaction with the cell wall components. IMPORTANCE Invasive fungal infections have increased in incidence with the widespread use of immunosuppressive therapy and invasive procedures. Activation of the complement system contributes to innate immunity against fungi by generating chemoattractants that recruit white blood cells and by coating the pathogen with complement fragments that “mark” them for phagocytosis. The fungal cell wall activates complement in an antibody-independent manner through the alternative pathway (AP). Properdin is a positive regulator of the AP. This study elucidates how the specificity of cell wall glycan linkages affects AP activation and the role properdin plays in this process. Particulate β1→3 glucans activated the AP even in the absence of properdin, while β1→6 glucans required properdin for AP activation. In contrast, the β1→4 glycans chitin and chitosan failed to activate the AP. These findings enhance our mechanistic understanding of how fungi activate complement and have implications for the use of glycans in biomedical applications.

ABSTRACT β-Glucan particles (GPs) are purified Saccharomyces cerevisiae cell walls treated so that they are primarily β1,3-d-glucans and free of mannans and proteins. GPs are phagocytosed by dendritic cells (DCs) via the Dectin-1 receptor, and this interaction stimulates proinflammatory cytokine secretion by DCs. As the hollow, porous GP structure allows for high antigen loading, we hypothesized that antigen-loaded GPs could be exploited as a receptor-targeted vaccine delivery system. Ovalbumin (OVA) was electrostatically complexed inside the hollow GP shells (GP-OVA). Incubation of C57BL/6J mouse bone marrow-derived DCs with GP-OVA resulted in phagocytosis, upregulation of maturation markers, and rapid proteolysis of OVA. Compared with free OVA, GP-OVA was >100-fold more potent at stimulating the proliferation of OVA-reactive transgenic CD8+ OT-I and CD4+ OT-II T cells, as measured by in vitro [3H]thymidine incorporation using DCs as antigen-presenting cells. Next, immune responses in C57BL/6J mice following subcutaneous immunizations with GP-OVA were compared with those in C57BL/6J mice following subcutaneous immunizations with OVA absorbed onto the adjuvant alum (Alum/OVA). Vaccination with GP-OVA stimulated substantially higher antigen-specific CD4+ T-cell lymphoproliferative and enzyme-linked immunospot (ELISPOT) responses than that with Alum/OVA. Moreover, the T-cell responses induced by GP-OVA were Th1 biased (determined by gamma interferon [IFN-γ] ELISPOT assay) and Th17 biased (determined by interleukin-17a [IL-17a] ELISPOT assay). Finally, both the GP-OVA and Alum/OVA formulations induced strong secretions of IgG1 subclass anti-OVA antibodies, although only GP-OVA induced secretion of Th1-associated IgG2c antibodies. Thus, the GP-based vaccine platform combines adjuvanticity and antigen delivery to induce strong humoral and Th1- and Th17-biased CD4+ T-cell responses. IMPORTANCE Most licensed vaccines work by promoting protective antibody responses. However, for many infectious diseases, antibody-mediated protection appears to play a relatively minor role, and vaccination has met with limited success. While live-attenuated organisms generally elicit T-cell responses, their use in vaccines is limited by the potential for causing disease. Thus, there is an urgent need for new vaccine platforms that deliver antigens in such a manner as to promote strong T-cell-mediated responses. Here we designed a novel vaccine platform consisting of yeast-derived β-glucan particles (GPs) that combines antigen delivery and adjuvant activity. GPs loaded with the model antigen ovalbumin (OVA) stimulated robust humoral and T-cell responses in mice. In addition, the cellular response was Th1 and Th17 biased. This work has implications for the design of vaccines that stimulate biased T-cell responses as well as for understanding how immunity to fungal pathogens develops.

An experimental approach for improving vaccine efficacy involves targeting antigens to mannose receptors (MRs) on dendritic cells (DCs) and other professional antigen presenting cells. Previously, we demonstrated that mannosylated Pichia pastoris-derived recombinant proteins exhibited increased immunogenicity compared to proteins lacking mannosylation. In order to gain insight into the mechanisms responsible for this observation, the present study examined the cellular uptake of the mannosylated and deglycosylated recombinant proteins.Utilizing transfected cell lines, roles for the macrophage mannose receptor (MMR, CD206) and DC-SIGN (CD209) in the recognition of the mannosylated, but not deglycosylated, antigens were demonstrated. The uptake of mannosylated antigens into murine bone marrow-derived DCs (BMDCs) was inhibited by yeast mannans (YMs), suggesting a mannose-specific C-type lectin receptor-dependent process, while the uptake of deglycosylated antigens remained unaffected. In particular, antigens with both N-linked and extensive O-linked mannosylation showed the highest binding and uptake by BMDCs. Finally, confocal microscopy studies revealed that both mannosylated and deglycosylated P. pastoris-derived recombinant proteins localized in MHC class II+ compartments within BMDCs.Taken together with our previous results, these data suggest that increased uptake by mannose-specific C-type lectin receptors is the major mechanism responsible for the enhanced antigenicity seen with mannosylated proteins. These findings have important implications for vaccine design and contribute to our understanding of how glycosylation affects the immune response to eukaryotic pathogens.