Multi-isotopes (δD, δ18Owater, 87Sr/86Sr, δ34S and δ18Osulfate) as indicators for groundwater salinization genesis and evolution of a large agricultural drainage lake basin in Inner Mongolia, Northwest China.

Groundwater salinization, a major eco-environmental problem in arid and semi-arid areas, can accelerate soil salinization, reducing crop productivity and imbalances in ecosystem diversity. This study classified water samples collected from the Ulansuhai Lake basin into five clusters using self-organizing maps (SOM). On this basis, multiple isotopes (δ18Owater, δD, 87Sr/86Sr, δ18Osulfate and δ34S) and isotopic models (Rayleigh fractionation and Bayesian isotope mixing models) were used to identify and quantify the genesis and evolution of groundwater salinization. The results showed that the samples were brackish or saline water, and the hydrochemical types were dominated by Na + K-Cl (SO4). It has been proved that the processes associated with groundwater salinization in the Ulansuhai Lake basin were dominated by water-rock interaction and human inputs. Among them, evaporite dissolution contributed substantially to groundwater salinity. Furthermore, salt inputs from human activities cannot be negligible. Based on the model calculations, evaporite dissolution accounted for the most significant proportion of all sources, with a mean value of 53 %. In addition, human inputs from regular agricultural activities (28 % from sewage and manure and 8 % from fertilizers) constituted another vital source of groundwater salinization associated with extensive agricultural activities in the study area. This study's results can deepen our understanding of the genesis of groundwater salinization and the evolution of the agricultural drainage lake basin. This knowledge will assist the Environmental Protection Department in developing effective policies for groundwater management in the Yellow River Basin.

Foxo1 is an iron-responsive transcriptional factor regulating systemic iron homeostasis.

The liver plays a crucial role in maintaining systemic iron homeostasis by secreting hepcidin, which is essential for coordinating iron levels in the body. Imbalances in iron homeostasis are associated with various clinical disorders related to iron deficiency or iron overload. Despite the clinical significance, the mechanisms underlying how hepatocytes sense extracellular iron levels to regulate hepcidin synthesis and iron storage are not fully understood. In this study, we identified Foxo1, a well-known regulator of macronutrient metabolism, that translocates to the nucleus of hepatocytes in response to high-iron feeding, holo-transferrin, and BMP6 treatment. Furthermore, Foxo1 plays a crucial role in mediating hepcidin induction in response to both iron and BMP signals by directly interacting with evolutionally conserved Foxo binding sites within the hepcidin promoter region. These binding sites were found to colocalize with Smad-binding sites. To investigate the physiological relevance of Foxo1 in iron metabolism, we generated mice with hepatocyte-specific deletion of Foxo1. These mice exhibited reduced hepatic hepcidin expression and serum hepcidin levels, accompanied by elevated serum iron and liver non-heme iron concentrations. Moreover, high-iron diet further exacerbated these abnormalities in iron metabolism in mice lacking hepatic Foxo1. Conversely, hepatocyte-specific Foxo1 overexpression increased hepatic hepcidin expression and serum hepcidin levels, thereby ameliorating iron overload in a murine model of hereditary hemochromatosis (Hfe-/- mice). In summary, our study identifies Foxo1 is a critical regulator of hepcidin and systemic iron homeostasis. Targeting Foxo1 may offer therapeutic opportunities for managing conditions associated with aberrant iron metabolism.

Heterologous expression of P9 from Akkermansia muciniphila increases the GLP-1 secretion of intestinal L cells.

Glucagon-like peptide-1(GLP-1) is an incretin hormone secreted primarily from the intestinal L-cells in response to meals. GLP-1 is a key regulator of energy metabolism and food intake. It has been proven that P9 protein from A. muciniphila could increase GLP-1 release and improve glucose homeostasis in HFD-induced mice. To obtain an engineered Lactococcus lactis which produced P9 protein, mature polypeptide chain of P9 was codon-optimized, fused with N-terminal signal peptide Usp45, and expressed in L. lactis NZ9000. Heterologous secretion of P9 by recombinant L. lactis NZP9 were successfully detected by SDS-PAGE and western blotting. Notably, the supernatant of L. lactis NZP9 stimulated GLP-1 production of NCI-H716 cells. The relative expression level of GLP-1 biosynthesis gene GCG and PCSK1 were upregulated by 1.63 and 1.53 folds, respectively. To our knowledge, this is the first report on the secretory expression of carboxyl-terminal processing protease P9 from A. muciniphila in L. lactis. Our results suggest that genetically engineered L. lactis which expressed P9 may have therapeutic potential for the treatment of diabetes, obesity and other metabolic disorders.

DNA vaccine prime and replicating vaccinia vaccine boost induce robust humoral and cellular immune responses against MERS-CoV in mice.

As of December 2022, 2603 laboratory-identified Middle East respiratory syndrome coronavirus (MERS-CoV) infections and 935 associated deaths, with a mortality rate of 36%, had been reported to the World Health Organization (WHO). However, there are still no vaccines for MERS-CoV, which makes the prevention and control of MERS-CoV difficult. In this study, we generated two DNA vaccine candidates by integrating MERS-CoV Spike (S) gene into a replicating Vaccinia Tian Tan (VTT) vector. Compared to homologous immunization with either vaccine, mice immunized with DNA vaccine prime and VTT vaccine boost exhibited much stronger and durable humoral and cellular immune responses. The immunized mice produced robust binding antibodies and broad neutralizing antibodies against the EMC2012, England1 and KNIH strains of MERS-CoV. Prime-Boost immunization also induced strong MERS-S specific T cells responses, with high memory and poly-functional (CD107a-IFN-γ-TNF-α) effector CD8+ T cells. In conclusion, the research demonstrated that DNA-Prime/VTT-Boost strategy could elicit robust and balanced humoral and cellular immune responses against MERS-CoV-S. This study not only provides a promising set of MERS-CoV vaccine candidates, but also proposes a heterologous sequential immunization strategy worthy of further development.

Homologous Overexpression of Tyrosinase in Trichoderma reesei and Its Application in Glycinin Cross-Linking.

Tyrosinase is capable of oxidizing tyrosine residues in proteins, leading to intermolecular protein cross-linking, which could modify the protein network of food and improve the texture of food. To obtain the recombinant tyrosinase with microbial cell factory instead of isolation tyrosinase from the mushroom Agaricus bisporus, a TYR expression cassette was constructed in this study. The expression cassette was electroporated into Trichoderma reesei Rut-C30 and integrated into its genome, resulting in a recombinant strain C30-TYR. After induction with microcrystalline cellulose for 7 days, recombinant tyrosinase could be successfully expressed and secreted by C30-TYR, corresponding to approximately 2.16 g/L tyrosinase in shake-flask cultures. The recombinant TYR was purified by ammonium sulfate precipitation and gel filtration, and the biological activity of purified TYR was 45.6 U/mL. The purified TYR could catalyze the cross-linking of glycinin, and the emulsion stability index of TYR-treated glycinin emulsion was increased by 30.6% compared with the untreated one. The cross-linking of soy glycinin by TYR resulted in altered properties of oil-in-water emulsions compared to emulsions stabilized by native glycinin. Therefore, cross-linking with this recombinant tyrosinase is a feasible approach to improve the properties of protein-stabilized emulsions and gels.

Faecalibacterium prausnitzii Supplementation Prevents Intestinal Barrier Injury and Gut Microflora Dysbiosis Induced by Sleep Deprivation.

Sleep deprivation (SD) leads to impaired intestinal barrier function and intestinal flora disorder, especially a reduction in the abundance of the next generation of probiotic Faecalibacterium prausnitzii (F. prausnitzii). However, it remains largely unclear whether F. prausnitzii can ameliorate SD-induced intestinal barrier damage. A 72 h SD mouse model was used in this research, with or without the addition of F. prausnitzii. The findings indicated that pre-colonization with F. prausnitzii could protect against tissue damage from SD, enhance goblet cell count and MUC2 levels in the colon, boost tight-junction protein expression, decrease macrophage infiltration, suppress pro-inflammatory cytokine expression, and reduce apoptosis. We found that the presence of F. prausnitzii helped to balance the gut microbiota in SD mice by reducing harmful bacteria like Klebsiella and Staphylococcus, while increasing beneficial bacteria such as Akkermansia. Ion chromatography analysis revealed that F. prausnitzii pretreatment increased the fecal butyrate level in SD mice. Overall, these results suggested that incorporating F. prausnitzii could help reduce gut damage caused by SD, potentially by enhancing the intestinal barrier and balancing gut microflora. This provides a foundation for utilizing probiotics to protect against intestinal illnesses.

Identification of an IGHV3-53-Encoded RBD-Targeting Cross-Neutralizing Antibody from an Early COVID-19 Convalescent.

Since November 2021, Omicron has emerged as the dominant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant, and its sublineages continue to appear one after another, significantly reducing the effectiveness of existing therapeutic neutralizing antibodies (NAbs). It is urgent to develop effective NAbs against circulating Omicron variants. Here, we isolated receptor binding domain (RBD)-specific single memory B cells via flow cytometry from a COVID-19 convalescent. The antibody variable region genes of the heavy chain (VHs) and light chain (VLs) were amplified and cloned into expression vectors. After antibody expression, ELISA screening and neutralizing activity detection, we obtained an IGHV3-53-encoded RBD-targeting cross-neutralizing antibody D6, whose VL originated from the IGKV1-9*01 germlines. D6 could potently neutralize circulating Omicron variants (BA.1, BA.2, BA.4/5 and BF.7), with IC50 values of less than 0.04 µg/mL, and the neutralizing ability against XBB was reduced but still effective. The KD values of D6 binding with RBD of the prototype and BA.1 were both less than 1.0 × 10-12 M. The protein structure of the D6-RBD model indicates that D6 interacts with the RBD external subdomain and belongs to the RBD-1 community. The sufficient contact and deep interaction of D6 HCDR3 and LCDR3 with RBD may be the crucial reason for its cross-neutralizing activity. The sorting and analysis of mAb D6 will provide important information for the development of anti-COVID-19 reagents.

Amino acid substitution of the membrane-proximal external region alter neutralization sensitivity in a chronic HIV-1 clade B infected patient.

The membrane-proximal external region (MPER) represents a highly conserved region of the Human Immunodeficiency Virus (HIV) envelope glycoprotein (env) targeted by several broadly neutralizing antibodies (bnAbs). In this study, we employed single genome amplification to amplify 34 full-length env sequences from the 2005 plasma sample of CBJC504, a chronic HIV-1 clade B infected individual. We identified three amino acid changes (N671S, D674N, and K677R) in the MPER. A longitudinal analysis revealed that the proportion of env sequences with MPER mutations increased from 26.5 % in 2005 to 56.0 % in 2009, and the sequences with the same mutation clustered together. Nine functional pseudoviruses were generated from the 34 env sequences to examine the effect of these mutations on neutralizing activity. Pseudoviruses carrying N674 or R677 mutations demonstrate increased sensitivity to autologous plasma and monoclonal antibodies 2F5, 4E10, and 10E8. Reverse mutations were performed in env including N674, R677, D659, and S671/N677 mutations, to validate the impact of the mutations on neutralizing sensitivity. Neutralization assays indicated that the N671S mutation increased neutralization sensitivity to 2F5 and 10E8. The amino acid R at position 677 increased viral resistance to 10E8, whereas N enhanced viral resistance to 4E10 and 10E8. It has been proposed that critical amino acids in the extra-MPER and the number of potential N-like glycosylation sites (PNGSs) in the V1 loop may have an impact on neutralizing activity. Understanding the mutations and evolution of MPER in chronically infected patients with HIV-1 is crucial for the design and development of vaccines that trigger bnAbs against MPER.

Ameliorative Effect of Coenzyme Q10 on Phenotypic Transformation in Human Smooth Muscle Cells with FBN1 Knockdown.

Mutations of the FBN1 gene lead to Marfan syndrome (MFS), which is an autosomal dominant connective tissue disorder featured by thoracic aortic aneurysm risk. There is currently no effective treatment for MFS. Here, we studied the role of mitochondrial dysfunction in the phenotypic transformation of human smooth muscle cells (SMCs) and whether a mitochondrial boosting strategy can be a potential treatment. We knocked down FBN1 in SMCs to create an MFS cell model and used rotenone to induce mitochondrial dysfunction. Furthermore, we incubated the shFBN1 SMCs with Coenzyme Q10 (CoQ10) to assess whether restoring mitochondrial function can reverse the phenotypic transformation. The results showed that shFBN1 SMCs had decreased TFAM (mitochondrial transcription factor A), mtDNA levels and mitochondrial mass, lost their contractile capacity and had increased synthetic phenotype markers. Inhibiting the mitochondrial function of SMCs can decrease the expression of contractile markers and increase the expression of synthetic genes. Imposing mitochondrial stress causes a double-hit effect on the TFAM level, oxidative phosphorylation and phenotypic transformation of FBN1-knockdown SMCs while restoring mitochondrial metabolism with CoQ10 can rapidly reverse the synthetic phenotype. Our results suggest that mitochondria function is a potential therapeutic target for the phenotypic transformation of SMCs in MFS.

Up-regulation of sortase-dependent pili in Bifidobacterium longum BBMN68 in response to bile stress enhances its adhesion to HT-29 cells.

Adhesion to gastrointestinal tract is crucial for bifidobacteria to exert their probiotic effects. Our previous work found that bile salts significantly enhance the adhesion ability of Bifidobacterium longum BBMN68 to HT-29 cells. In this study, trypsin-shaving and LC-MS/MS-based surface proteomics were employed to identify surface proteins involved in bile stress response. Among the 829 differentially expressed proteins, 56 up-regulated proteins with a fold change >1.5 were subjected to further analysis. Notably, the minor pilin subunit FimB was 4.98-fold up-regulated in response to bile stress. In silico analysis and RT-PCR confirmed that gene fimB, fimA and srtC were co-transcribed and contributed to the biosynthesis of sortase-dependent pili Pil1. Moreover, scanning electron microscopy and immunogold electron microscopy assays showed increased abundance and length of Pil1 on BBMN68 under bile stress. As the major pilin subunit FimA serves as adhesion component of Pil1, an inhibition assay using anti-FimA antibodies further confirmed the critical role of Pil1 in mediating the adhesion of BBMN68 to HT-29 cells under bile stress. Our findings suggest that the up-regulation of Pil1 in response to bile stress enhances the adhesion of BBMN68 to intestinal epithelial cells, highlighting a novel mechanism of gut persistence in B. longum strains.

A Ropy Exopolysaccharide-Producing Strain Bifidobacterium pseudocatenulatum Bi-OTA128 Alleviates Dextran Sulfate Sodium-Induced Colitis in Mice.

Inflammatory bowel disease (IBD) is a chronic disease associated with overactive inflammation and gut dysbiosis. Owing to the beneficial effects of bifidobacteria on IBD treatment, this study aimed to investigate the anti-inflammation effects of an exopolysaccharide (EPS)-producing strain Bifidobacterium pseudocatenulatum Bi-OTA128 through a dextran sulfate sodium (DSS)-induced colitis mice model. B. pseudocatenulatum treatment improved DSS-induced colitis symptoms and maintained intestinal barrier integrity by up-regulating MUC2 and tight junctions' expression. The oxidative stress was reduced after B. pseudocatenulatum treatment by increasing the antioxidant enzymes of SOD, CAT, and GSH-Px in colon tissues. Moreover, the overactive inflammatory responses were also inhibited by decreasing the pro-inflammatory cytokines of TNF-α, IL-1ß, and IL-6, but increasing the anti-inflammatory cytokine of IL-10. The EPS-producing strain Bi-OTA128 showed better effects than that of a non-EPS-producing stain BLYR01-7 in modulating DSS-induced gut dysbiosis. The Bi-OTA128 treatment increased the relative abundance of beneficial bacteria Bifidobacterium and decreased the maleficent bacteria Escherichia-Shigella, Enterorhabuds, Enterobacter, and Osillibacter associated with intestinal inflammation. Notably, the genera Clostridium sensu stricto were only enriched in Bi-OTA128-treated mice, which could degrade polysaccharides to produce acetic acid and butyrate in the gut. This finding demonstrated a cross-feeding effect induced by the EPS-producing strain in gut microbiota. Collectively, these results highlighted the anti-inflammatory effects of the EPS-producing strain B. pseudocatenulatum Bi-OTA128 on DSS-induced colitis, which could be used as a candidate probiotic supporting recovery from ongoing colitis.

bglG Regulates the Heterogeneity Driven by the Acid Tolerance Response in Lacticaseibacillus paracasei L9.

The acid tolerance of lactic acid bacteria is crucial for their fermentation and probiotic functions. Acid adaption significantly enhances the acid tolerance of strains, and the phenotypic heterogeneity driven by the acid tolerance response (ATR) contributes to this process by providing a selective advantage in harsh environments. The mechanism of heterogeneity under the ATR is not yet clear, but individual gene expression differences are recognized as the cause. In this study, we observed four heterogeneous subpopulations (viable, injured, dead, and unstained) of Lacticaseibacillus paracasei L9 (L9) induced by acid adaption (pH 5.0, 40 min) using flow cytometry. The viable subpopulation represented a significantly superior acid tolerance to the injured subpopulation or total population. Different subpopulations were sorted and transcriptomic analysis was performed. Five genes were found to be upregulated in the viable subpopulation and downregulated in the injured subpopulation, and bglG (LPL9_RS14735) was identified as having a key role in this process. Using salicin (glucoside)-inducing gene expression and gene insertion mutagenesis, we verified that bglG regulated the heterogeneity of the acid stress response and that the relevant mechanisms might be related to activating hsp20. This study provides new evidence for the mechanism of the ATR and may contribute to the theoretical basis of improving the acid tolerance of Lacticaseibacillus paracasei L9.

Lactobacillus plantarum Zhang-LL Inhibits Colitis-Related Tumorigenesis by Regulating Arachidonic Acid Metabolism and CD22-Mediated B-Cell Receptor Regulation.

Colorectal cancer (CRC) is a significant health concern and is the third most commonly diagnosed and second deadliest cancer worldwide. CRC has been steadily increasing in developing countries owing to factors such as aging and epidemics. Despite extensive research, the exact pathogenesis of CRC remains unclear, and its causes are complex and variable. Numerous in vitro, animal, and clinical trials have demonstrated the efficacy of probiotics such as Lactobacillus plantarum in reversing the adverse outcomes of CRC. These findings suggest that probiotics play vital roles in the prevention, adjuvant treatment, and prognosis of CRC. In this study, we constructed a mouse model of CRC using an intraperitoneal injection of azomethane combined with dextran sodium sulfate, while administering 5-fluorouracil as well as high- and low-doses of L. plantarum Zhang-LL live or heat-killed strains. Weight changes and disease activity indices were recorded during feeding, and the number of polyps and colon length were measured after euthanasia. HE staining was used to observe the histopathological changes in the colons of mice, and ELISA was used to detect the expression levels of IL-1ß, TNF-α, and IFN-γ in serum. To investigate the specific mechanisms involved in alleviating CRC progression, gut microbial alterations were investigated using 16S rRNA amplicon sequencing and non-targeted metabolomics, and changes in genes related to CRC were assessed using eukaryotic transcriptomics. The results showed that both viable and heat-killed strains of L. plantarum Zhang-LL in high doses significantly inhibited tumorigenesis, colon shortening, adverse inflammatory reactions, intestinal tissue damage, and pro-inflammatory factor expression upregulation. Specifically, in the gut microbiota, the abundance of the dominant flora Acutalibacter muris and Lactobacillus johnsonii was regulated, PGE2 expression was significantly reduced, the arachidonic acid metabolism pathway was inhibited, and CD22-mediated B-cell receptor regulation-related gene expression was upregulated. This study showed that L. plantarum Zhang-LL live or heat-inactivated strains alleviated CRC progression by reducing the abundance of potentially pathogenic bacteria, increasing the abundance of beneficial commensal bacteria, mediating the arachidonic acid metabolism pathway, and improving host immunogenicity.

Dandelion Polysaccharides Ameliorate High-Fat-Diet-Induced Atherosclerosis in Mice through Antioxidant and Anti-Inflammatory Capabilities.

Dandelion (Taraxacum officinale), a member of the Asteraceae (Compositae) family, is well known as the traditional medical plant. Dandelion polysaccharides, a natural active ingredient extracted from the dandelion, possess immune regulation, anti-inflammatory, antioxidant, and anti-aggregation properties. These properties suggest that dandelion polysaccharides might alleviate atherosclerosis. Using an ApoE-/- atherosclerotic mice model fed a high-fat diet, we investigated the impact and potential mechanism of dandelion polysaccharides on atherosclerosis. We observed that dandelion polysaccharides significantly reduced the levels of triglyceride, total cholesterol, and low-density lipoprotein-cholesterol in serum, while elevated the high-density lipoprotein-cholesterol level. Concomitantly, dandelion polysaccharides reduced the area of atherosclerotic lesions and necrotic core of the aortic sinus, and increased the collagen content. Mechanistic studies showed that dandelion polysaccharides were effective in reducing serum malondialdehyde levels while elevating the enzymatic activities of superoxide dismutase and glutathione peroxidase. Furthermore, dandelion polysaccharides reduced the expression of chemotactic factor Mcp-1 and pro-inflammatory cytokines (Tnf-α, Il-1ß, and Il-6) in atherosclerotic lesions. Overall, these results indicated that dandelion polysaccharides may take an important part in the attenuation of atherosclerosis via its antioxidant and anti-inflammatory properties.

An underlying mechanism for MleR activating the malolactic enzyme pathway to enhance acid tolerance in Lacticaseibacillus paracasei L9.

Tolerance to acid stress is a crucial property of probiotics against gastric acids. The malolactic enzyme pathway is one of the most important acid resistance systems in lactic acid bacteria. It has been reported that the malolactic enzyme pathway was regulated by the transcriptional regulator, MleR. However, regulatory mechanisms underlying malolactic enzyme pathway to cope with acid stress remain unknown. In this study, the acid tolerance ability of the ΔmleR deletion strain was significantly lower than that of the wild-type strain, and the complementation of the mleR gene into the ΔmleR strain restored the acid tolerance of the ΔmleR strain, indicating that MleR was involved in acid tolerance response of Lacticaseibacillus paracasei L9. Real-time quantitative PCR and transcriptional fusion experiments confirmed MleR-activated transcription of the mleST gene cluster. Furthermore, MleR was confirmed to directly bind to the promoter region of the mleST operon using ChIP assays and EMSAs. The transcription start site G of the mleST operon was located at position -198 relative to the start codon of the mleS gene. The region from -80 to -61 upstream of the transcription start site was determined to be essential for MleR binding. Moreover, L-malic acid acted as an effector for MleR to activate the transcription of the mleST operon in a dose-dependent manner. These results revealed the regulatory mechanism behind MleR-mediated activation of the malolactic enzyme pathway to enhance acid tolerance in Lc. paracasei L9. IMPORTANCE Lacticaseibacillus paracasei is extensively used as probiotics in human health and fermented dairy production. Following consumption, Lc. paracasei is exposed to a variety of physico-chemical stresses, such as low pH in the stomach and bile salts in the intestines. The high acidity of the stomach severely inhibits bacterial metabolism and growth. Therefore, the acid tolerance response is critical for Lc. paracasei to survive. It has been reported that the malolactic enzyme (MLE) pathway plays an important role for LAB to resist acid stress. However, the regulatory mechanism has not yet been investigated. In this study, we determined that the LysR-type regulator MleR positively regulated the MLE pathway to enhance acid tolerance by binding -80 to -61 upstream of the transcription start site of the mleST operon. Further, L-malic acid acts as a co-inducer for MleR transcriptional regulation. Our study provides novel insights into acid tolerance mechanisms in LAB.

Sequential heterologous immunization with COVID-19 vaccines induces broader neutralizing responses against SARS-CoV-2 variants in comparison with homologous boosters.

The recently prevalent variants of concerns (VOCs) of SARS-CoV-2 belong to Omicron variants which display increased transmissibility and evade from immune protection generated by vaccines and/or natural infections. Better immunization strategies should be explored to induce broader immune responses against evolving SARS-CoV-2 variants. Here, we used inactivated vaccines derived from ancestral (Wu), Delta (Del) and Omicron (Omi) strains to immunize mice with homologous booster (3 × Wu, 3 × Del and 3 × Omi) or heterologous sequential booster (Wu/Del/Omi and Omi/Wu/Del) to evaluate their responses against two pre-Omicron (Wu and Del) and four Omicron variants. Even though neutralization responses against Wu and Del variants were similar in heterologous and homologous immunization groups, heterologous immunization groups induced significantly stronger neutralizing antibody against BA.1 (4.1-11 folds higher) and BA.2 (4.7-14.2 folds higher) than those of homologous immunization groups. While homologous immunization only induced strong neutralizing responses to either pre-Omicron variants (Wu and Del) in 3 × Wu and 3 × Del groups or to Omicron variants (BA.1 and BA.2) in 3 × Omi group, heterologous immunization groups induced strong and broader neutralizing responses to both pre-Omicron (Wu, Del) and Omicron variants (BA.1 and BA.2). Homologous and heterologous immunization groups elicited similar antigen-specific T cell (IFN-γ+) and B cell responses. Compared with homologous immunization, heterologous immunization could induce stronger plasma cell responses, which have the potential to generate broader and stronger neutralizing antibodies. However, neither heterologous nor homologous immunization groups induced strong neutralizing antibody against variants with bigger genetic deviation, such as BA.4/5 or BF.7, only weak neutralizing responses were induced. Surveillance on SARS-CoV2 variants evolution and immunization strategy are needed to explore better vaccines with broader and stronger neutralizing antibodies against post pandemic COVID-19.

Structures and immune recognition of Env trimers from two Asia prevalent HIV-1 CRFs.

Structure-guided immunofocusing HIV-1 vaccine design entails a comprehensive understanding of Envs from diverse HIV-1 subtypes, including circulating recombinant forms (CRFs). Here, we present the cryo-EM structures of Envs from two Asia prevalent CRFs (CRF01_AE and CRF07_BC) at 3.0 and 3.5 Å. We compare the structures and glycosylation patterns of Envs from different subtypes and perform cross-clade statistical analyses to reveal the unique features of CRF01_AE V1 region, which are associated with the resistance to certain bNAbs. We also solve a 4.1 Å cryo-EM structure of CRF01_AE Env in complex with F6, the first bNAb from CRF01_AE-infected individuals. F6 recognizes a gp120-gp41 spanning epitope to allosterically destabilize the Env trimer apex and weaken inter-protomer packing, which in turn hinders the receptor binding and induces Env trimer disassembly, demonstrating a dual mechanism of neutralization. These findings broaden our understanding of CRF Envs and shed lights on immunofocusing HIV-1 vaccine design.

Characterization of RBD-specific cross-neutralizing antibodies responses against SARS-CoV-2 variants from COVID-19 convalescents.

Introduction: The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has been posing a severe threat to global public health. Although broadly neutralizing antibodies have been used to prevent or treat corona virus disease 2019 (COVID-19), new emerging variants have been proven resistant to these antibodies. Methods: In this study, we isolated receptor binding domain (RBD)-specific memory B cells using single-cell sorting method from two COVID-19 convalescents and expressed the antibody to test their neutralizing activity against diverse SARS-CoV-2 variants. Then, we resolved antibody-RBD complex structures of potent RBD-specific neutralizing antibodies by X-ray diffraction method. Finally, we analyzed the whole antibody repertoires of the two donors and studied the evolutionary pathway of potent neutralizing antibodies. Results and discussion: We identified three potent RBD-specific neutralizing antibodies (1D7, 3G10 and 3C11) from two COVID-19 convalescents that neutralized authentic SARS-CoV-2 WH-1 and Delta variant, and one of them, 1D7, presented broadly neutralizing activity against WH-1, Beta, Gamma, Delta and Omicron authentic viruses. The resolved antibody-RBD complex structures of two antibodies, 3G10 and 3C11, indicate that both of them interact with the external subdomain of the RBD and that they belong to the RBD-1 and RBD-4 communities, respectively. From the antibody repertoire analysis, we found that the CDR3 frequencies of the light chain, which shared high degrees of amino acid identity with these three antibodies, were higher than those of the heavy chain. This research will contribute to the development of RBD-specific antibody-based drugs and immunogens against multiple variants.

Mucin Binding Protein of Lactobacillus casei Inhibits HT-29 Colorectal Cancer Cell Proliferation.

Many Lactobacillus casei strains are reported to exhibit anti-proliferative effects on colorectal cancer cells; however, the mechanism remains largely unknown. While there has been considerable interest in bacterial small metabolites such as short chain fatty acids, prior reports suggested that larger-sized molecules mediate the anti-proliferative effect of L. casei. Here, other possible ways of communication between gut bacteria and its host are investigated. LevH1 is a protein displayed on the surface of L. casei, and its mucin binding domain is highly conserved. Based on previous reports that the cell-free supernatant fractions decreased colorectal cell proliferation, we cloned the mucin binding domain of the LevH1 protein, expressed and purified this mucin binding protein (MucBP). It has a molecular weight of 10 kDa, is encoded by a 250 bp gene, and is composed primarily of a ß-strand, ß-turns, and random coils. The amino acid sequence is conserved while the 36th amino acid residue is arginine in L. casei CAUH35 and serine in L. casei IAM1045, LOCK919, 12A, and Zhang. MucBP36R exhibited dose-dependent anti-proliferative effects against HT-29 cells while a mutation of 36S abolished this activity. Predicted structures suggest that this mutation slightly altered the protein structure, thus possibly affecting subsequent communication with HT-29 cells. Our study identified a novel mode of communication between gut bacteria and their host.

Sequential Administration of SARS-CoV-2 Strains-Based Vaccines Effectively Induces Potent Immune Responses against Previously Unexposed Omicron Strain.

In the past few years, the continuous pandemic of COVID-19 caused by SARS-CoV-2 has placed a huge burden on public health. In order to effectively deal with the emergence of new SARS-CoV-2 variants, it becomes meaningful to further enhance the immune responses of individuals who have completed the first-generation vaccination. To understand whether sequential administration using different variant sequence-based inactivated vaccines could induce better immunity against the forthcoming variants, we tried five inactivated vaccine combinations in a mouse model and compared their immune responses. Our results showed that the sequential strategies have a significant advantage over homologous immunization by inducing robust antigen-specific T cell immune responses in the early stages of immunization. Furthermore, the three-dose vaccination strategies in our research elicited better neutralizing antibody responses against the BA.2 Omicron strain. These data provide scientific clues for finding the optimal strategy within the existing vaccine platform in generating cross-immunity against multiple variants including previously unexposed strains.