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Proceedings of the National Academy of... Feb 2024The Ebola virus causes hemorrhagic fever in humans and poses a significant threat to global public health. Although two viral vector vaccines have been approved to...
The Ebola virus causes hemorrhagic fever in humans and poses a significant threat to global public health. Although two viral vector vaccines have been approved to prevent Ebola virus disease, they are distributed in the limited ring vaccination setting and only indicated for prevention of infection from (EBOV)-one of three species that have caused previous outbreaks. Ebola virus glycoprotein GP mediates viral infection and serves as the primary target of neutralizing antibodies. Here, we describe a universal Ebola virus vaccine approach using a structure-guided design of candidates with hyperglycosylation that aims to direct antibody responses away from variable regions and toward conserved epitopes of GP. We first determined the hyperglycosylation landscape on Ebola virus GP and used that to generate hyperglycosylated GP variants with two to four additional glycosylation sites to mask the highly variable glycan cap region. We then created vaccine candidates by displaying wild-type or hyperglycosylated GP variants on ferritin nanoparticles (Fer). Immunization with these antigens elicited potent neutralizing antisera against EBOV in mice. Importantly, we observed consistent cross-neutralizing activity against Bundibugyo virus and Sudan virus from hyperglycosylated GP-Fer with two or three additional glycans. In comparison, elicitation of cross-neutralizing antisera was rare in mice immunized with wild-type GP-Fer. These results demonstrate a potential strategy to develop universal Ebola virus vaccines that confer cross-protective immunity against existing and emerging filovirus species.
Topics: Humans; Animals; Mice; Hemorrhagic Fever, Ebola; Ebola Vaccines; Antibodies, Viral; Ebolavirus; Antibodies, Neutralizing; Viral Vaccines; Immune Sera
PubMed: 38319964
DOI: 10.1073/pnas.2316960121 -
Journal of Virology Oct 2022Mutations at spike protein L452 are recurrently observed in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOC), including omicron...
Mutations at spike protein L452 are recurrently observed in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOC), including omicron lineages. It remains elusive how amino acid substitutions at L452 are selected in VOC. Here, we characterized all 19 possible mutations at this site and revealed that five mutants expressing the amino acids Q, K, H, M, and R gained greater fusogenicity and pseudovirus infectivity, whereas other mutants failed to maintain steady-state expression levels and/or pseudovirus infectivity. Moreover, the five mutants showed decreased sensitivity toward neutralization by vaccine-induced antisera and conferred escape from T cell recognition. Contrary to expectations, sequence data retrieved from the Global Initiative on Sharing All Influenza Data (GISAID) revealed that the naturally occurring L452 mutations were limited to Q, M, and R, all of which can arise from a single nucleotide change. Collectively, these findings highlight that the codon base change mutational barrier is a prerequisite for amino acid substitutions at L452, in addition to the phenotypic advantages of viral fitness and decreased sensitivity to host immunity. In a span of less than 3 years since the declaration of the coronavirus pandemic, numerous SARS-CoV-2 variants of concern have emerged all around the globe, fueling a surge in the number of cases and deaths that caused severe strain on the health care system. A major concern is whether viral evolution eventually promotes greater fitness advantages, transmissibility, and immune escape. In this study, we addressed the differential effect of amino acid substitutions at a frequent mutation site, L452 of SARS-CoV-2 spike, on viral antigenic and immunological profiles and demonstrated how the virus evolves to select one amino acid over the others to ensure better viral infectivity and immune evasion. Identifying such virus mutation signatures could be crucial for the preparedness of future interventions to control COVID-19.
Topics: Humans; SARS-CoV-2; Spike Glycoprotein, Coronavirus; COVID-19; Amino Acid Substitution; Immune Sera; Amino Acids; Nucleotides; Mutation
PubMed: 36214577
DOI: 10.1128/jvi.01162-22 -
Toxicon : Official Journal of the... Oct 2020Antivenoms are currently the most effective medication used in the treatment of snakebites. However, there were relatively few studies on preparation of antivenoms...
Antivenoms are currently the most effective medication used in the treatment of snakebites. However, there were relatively few studies on preparation of antivenoms targeting sea snakes, especially common sea snakes in China. In this study, we sought to prepare and detect mono- and bispecific antisera raised in rabbits against venoms of two sea snakes, Hydrophis cyanocinctus and H. curtus. The results of enzyme-linked immunosorbent assay showed that the rabbit antisera generally showed clearly detectable immunological cross-reactions after the third immunization and indicated that the strength of cross-reactions increased with an increase in the immunizing dose. Proteins within the H. cyanocinctus and H. curtus venoms showed similar profiles and were mainly concentrated in the low-molecular-weight region (8-25 kDa). Western blotting results revealed that the bands of these low-molecular weight proteins were dense and showed strong immunogenicity. Although we detected comparatively few bands of the high-molecular-weight proteins, these also showed strong immunogenicity. Our results indicate that both mono- and bispecific antisera both can neutralize H. cyanocinctus and H. curtus venoms, and in this regard, the monospecific H. curtus and bispecific antiserum were found to be superior to the H. cyanocinctus antiserum. Given the increasing frequency of snakebites worldwide, we believe that the findings of this study will have high practical applicability.
Topics: Animals; Antivenins; China; Cross Reactions; Elapid Venoms; Elapidae; Hydrophiidae; Immune Sera; Rabbits; Snake Bites; Snake Venoms
PubMed: 32828954
DOI: 10.1016/j.toxicon.2020.08.018 -
ACS Chemical Biology Nov 2021is a significant cause of human gastroenteritis worldwide, and all strains express an N-glycan that is added to at least 80 different proteins. We characterized 98...
is a significant cause of human gastroenteritis worldwide, and all strains express an N-glycan that is added to at least 80 different proteins. We characterized 98 isolates from infants from 7 low- and middle-income countries and identified 4 isolates unreactive with our N-glycan-specific antiserum that was raised against the heptasaccharide composed of GalNAc-GalNAc-GalNAc(Glc)-GalNAc-GalNAc-diNAcBac. Mass spectrometric analyses indicated these isolates express a hexasaccharide lacking the glucose branch. Although all 4 strains encode the PglI glucosyltransferase (GlcTF), one aspartate in the DXDD motif was missing, an alteration also present in ∼4% of all available PglI sequences. Deleting this residue from an active PglI resulted in a nonfunctional GlcTF when the protein glycosylation system was reconstituted in , while replacement with Glu/Ala was not deleterious. Molecular modeling proposed a mechanism for how the DXDD residues and the structure/length beyond the motif influence activity. Mouse vaccination with an strain expressing the full-length heptasaccharide produced N-glycan-specific antibodies and a corresponding reduction in colonization and weight loss following challenge. However, the antibodies did not recognize the hexasaccharide and were unable to opsonize isolates lacking glucose, suggesting this should be considered when designing N-glycan-based vaccines to prevent campylobacteriosis.
Topics: Amino Acid Sequence; Animals; Aspartic Acid; Campylobacter jejuni; Carbohydrate Conformation; Carbohydrate Sequence; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Glucose; Glycosylation; Immune Sera; Mice; Phagocytosis; Polysaccharides; Sequence Alignment
PubMed: 34726367
DOI: 10.1021/acschembio.1c00498 -
Virulence Dec 2023Brucellosis is a major threat to public health and animal husbandry. Several vertebrate models, such as mice, guinea pigs, and nonhuman primates, have been used to...
Brucellosis is a major threat to public health and animal husbandry. Several vertebrate models, such as mice, guinea pigs, and nonhuman primates, have been used to study pathogenesis, bacteria-host interactions, and vaccine efficacy. However, these models have limitations whereas the invertebrate model is a cost-effective and ethical alternative. The aim of the present study was to examine the invertebrate as an in vivo infection model for . Infection assays were employed to validate the fitness of the larval model for infection and virulence evaluation. The protective efficacy of immune sera was evaluated by pre-incubated with a lethal dose of bacteria before infection. The consistency between the mouse model and the larval model was confirmed by assessing the protective efficacy of two vaccine strains. The results show that could be infected by strains, in a dose- and temperature-dependent way. Moreover, this larval model can effectively evaluate the virulence of strains in a manner consistent with that of mammalian infection models. Importantly, this model can assess the protective efficacy of vaccine immune sera within a day. Further investigation implied that haemolymph played a crucial role in the protective efficacy of immune sera. In conclusion, could serve as a quick, efficient, and reliable model for evaluating the virulence of strains and efficacy of immune sera in an ethical manner.
Topics: Animals; Mice; Guinea Pigs; Brucella; Moths; Larva; Virulence; Immune Sera; Disease Models, Animal; Mammals
PubMed: 37817444
DOI: 10.1080/21505594.2023.2268496 -
Methods in Molecular Biology (Clifton,... 2019Immunoelectrophoresis can be used for analysis of individual proteins in complex mixtures. The conditions involved in immunoelectrophoresis are mild, avoiding the risk...
Immunoelectrophoresis can be used for analysis of individual proteins in complex mixtures. The conditions involved in immunoelectrophoresis are mild, avoiding the risk of denaturation, and it is possible to perform relative quantification of individual components. The fundamental disadvantage is the dependence on rabbit antisera as reagents. The usefulness of immunoelectrophoresis in allergy research is greatly enhanced by the possibility of identification of allergens to which the individual in question has IgE.The common principle is characterized by two independent electrophoreses having direction of current perpendicular to each other, i.e., crossed immunoelectrophoresis (CIE). This ultimately results in the formation of characteristic bell-shaped precipitates, each precipitate representing one antigen. There is a linear relationship between the amount of antigen and size of precipitate for a given antibody concentration for each precipitate and so relative quantification can be performed. The sensitivity and resolution power of CIE are very high and there are multiple variations of the technique, some of which will be illustrated in this chapter.
Topics: Allergens; Animals; Cats; Humans; Immune Sera; Immunoelectrophoresis, Two-Dimensional; Rabbits
PubMed: 31177491
DOI: 10.1007/978-1-4939-9591-2_4 -
The American Journal of Tropical... Jun 2020
Topics: Antibodies, Monoclonal; Antiviral Agents; Betacoronavirus; COVID-19; COVID-19 Testing; COVID-19 Vaccines; Clinical Laboratory Techniques; Coronavirus Infections; Humans; Immune Sera; Immunotherapy; Pandemics; Pneumonia, Viral; SARS-CoV-2; United States; Viral Vaccines
PubMed: 32274985
DOI: 10.4269/ajtmh.20-0239 -
Theriogenology Sep 2019Superovulation technique is important to improve the efficiency of oocyte and animal production and reduce the number of oocyte donors. Previously, we have reported that...
Superovulation technique is important to improve the efficiency of oocyte and animal production and reduce the number of oocyte donors. Previously, we have reported that the coadministration of inhibin antiserum (IAS) and equine chorionic gonadotropin (eCG) results in the production of >100 oocytes in a 4-week-old female C57BL/6 mice. It is well established that superovulation depends on the age of the female mice. However, detailed data regarding the ovulation of juvenile, mature, and aged female mice following the administration of IAS and eCG as well as the performance of reproductive technologies using oocytes have not yet been investigated. In the present study, we examined the effect of the age of female mice (3-50 weeks old) on the number of ovulated oocytes via the coadministration of IAS and eCG or eCG alone. Treatment with IAS plus eCG produced the maximum number of oocytes at 4 weeks of age. Moreover, IAS plus eCG produced more oocytes than eCG alone in mice aged between 3 and 5 weeks or 7 and 30 weeks. The fertilization and birth rates were similar between the two treatments at any age. Moreover, after vitrifying and warming the embryos, the survival and birth rates of two-cell embryos were similar between the two treatments. Subsequently, we examined the optimal ages of female mice (between 24 and 34 days) to obtain a high and stable number of oocytes. In mice aged between 24 and 32 days, IAS plus eCG induced the production of more eggs than eCG alone. Notably, the coadministration of IAS and eCG in mice aged between 25 and 31 days resulted in stable ovulation and high number of oocytes. Using the tip of the optimal female aged between 25 and 31 days old, we demonstrated an efficient production of embryos and offspring between homozygous knockout males and few females aged 26-28 days via in-vitro fertilization and embryo transfer. In summary, the coadministration of IAS and eCG resulted in a higher number of oocytes in juvenile, mature, and aged female mice. This treatment may be useful for the efficient production of homozygous mutant mice from a limited number of female mice.
Topics: Aging; Animals; Chorionic Gonadotropin; Cryopreservation; Drug Therapy, Combination; Embryo Culture Techniques; Female; Fertilization in Vitro; Immune Sera; Inhibins; Male; Mice; Mice, Inbred C57BL; Oocytes; Superovulation
PubMed: 31185424
DOI: 10.1016/j.theriogenology.2019.05.028 -
Journal of Bacteriology Oct 2019Capsular polysaccharides (CPS) are crucial virulence factors of The previously unknown CPS structures of the pneumococcal serogroup 16 (serotypes 16F and 16A) were...
Capsular polysaccharides (CPS) are crucial virulence factors of The previously unknown CPS structures of the pneumococcal serogroup 16 (serotypes 16F and 16A) were thoroughly elucidated by nuclear magnetic resonance (NMR) spectroscopy and verified by chemical analysis. The following repeat unit structures were determined: 16F, -3)-α-l-Rha-[4-P-1-Gro]-(1-3)-α-d-Glc-[(6-P-1)-Gro]-(1-3)-β-l-Rha-[2-OAc]-(1-4)-β-d-Glc-(1-; 16A, -3)-β-d-Gal-[2-OAc (70%)]-(1-3)-α-l-Rha-(1-2)-α-l-Rha-(1-3)-α-d-Gal-[(6-P-1)-Gro]-(1-3)-β-d-Gal-(1-4)-β-d-Glc-(1- (OAc, O-acetyl substitution; P-1-Gro, glycerol-1-phosphate substitution) A further analysis of CPS biosynthesis of serotypes 16F and 16A, in conjunction with published gene bioinformatics analysis and structures of related serotypes, revealed presumable specific function of glycosyltransferase, acetyltransferase, phosphotransferase, and polymerase. The functions of glycosyltransferases WcxN and WcxT were proposed for the first time, and they were assigned to catalyze linkage of α-l-Rha-(1-3)-α-d-Glc and α-l-Rha-(1-2)-α-l-Rha, respectively. Furthermore, since serotype 16F was genetically close to serogroup 28, cross-reactions between serogroup 16 and serogroup 28 were studied using diagnostic antisera, which provided further understanding of antigenic properties of CPS and diagnostic antisera. Interestingly, serotype 16F cross-reacted with factor antisera 28b and 11c. Meanwhile, serotype 16A cross-reacted with factor antiserum 11c. The vaccine pressure against could result in a change of prevalence in carriage and invasive serotypes. As such, it is necessary to monitor the distribution to achieve successful vaccination of the population, and similarly, it is important to increase the knowledge of even the currently less prevalent serotypes. The CPS are vital for the virulence of the pathogen, and antigenic properties of CPS are based on the structure. Consequently, a better understanding of the structure, biosynthesis, and serology of the capsular polysaccharides can be of great importance toward developing future diagnostic tools and vaccines.
Topics: Animals; Bacterial Capsules; Bacterial Proteins; Carbohydrate Sequence; Cross Reactions; Glycosyltransferases; Immune Sera; Magnetic Resonance Spectroscopy; Mutation; Polysaccharides, Bacterial; Rabbits; Serogroup; Streptococcus pneumoniae
PubMed: 31383737
DOI: 10.1128/JB.00453-19 -
Vaccine Mar 2021Zika virus(ZIKV) is primarily spread by Aedes. aegyptimosquitoes. Infection with ZIKV can result in diverse clinical symptoms in humans, ranging from mild to severe....
Zika virus(ZIKV) is primarily spread by Aedes. aegyptimosquitoes. Infection with ZIKV can result in diverse clinical symptoms in humans, ranging from mild to severe. Previously, we demonstrated that passive immunization against A. aegypti AgBR1 or NeSt1 antiserum, two mosquito saliva proteins that are transmitted with the virus, conferred partial protection against ZIKV in mice. Each individual antiserum altered the early host response in the skin and reduced viremia. Here, we show that passive immunization with a combination of AgBR1- and NeSt1-specific antibodies enhanced survival and reduced the viral burden in blood, thereby protecting mice from mosquito-borne ZIKV infection. This finding suggests that targeting a combination of mosquito saliva proteins, with AgBR1 and NeSt1 as model antigens, may be used as a vaccine strategy to help prevent mosquito-borne ZIKV infection.
Topics: Aedes; Animals; Immune Sera; Mice; Mosquito Vectors; Saliva; Zika Virus; Zika Virus Infection
PubMed: 33622591
DOI: 10.1016/j.vaccine.2021.01.072