Antigenic Cross-reactivity Research Articles

Genetic Susceptibility to Fungal Infections and Links to Human Ancestry.

Over the ages, fungi have associated with different parts of the human body and established symbiotic associations with their host. They are mostly commensal unless there are certain not so well-defined factors that trigger the conversion to a pathogenic state. Some of the factors that induce such transition can be dependent on the fungal species, environment, immunological status of the individual, and most importantly host genetics. In this review, we discuss the different aspects of how host genetics play a role in fungal infection since mutations in several genes make hosts susceptible to such infections. We evaluate how mutations modulate the key recognition between the pathogen associated molecular patterns (PAMP) and the host pattern recognition receptor (PRR) molecules. We discuss the polymorphisms in the genes of the immune system, the way it contributes toward some common fungal infections, and highlight how the immunological status of the host determines fungal recognition and cross-reactivity of some fungal antigens against human proteins that mimic them. We highlight the importance of single nucleotide polymorphisms (SNPs) that are associated with several of the receptor coding genes and discuss how it affects the signaling cascade post-infection, immune evasion, and autoimmune disorders. As part of personalized medicine, we need the application of next-generation techniques as a feasible option to incorporate an individual’s susceptibility toward invasive fungal infections based on predisposing factors. Finally, we discuss the importance of studying genomic ancestry and reveal how genetic differences between the human race are linked to variation in fungal disease susceptibility.

A Defined Antigen Skin Test for Diagnosis of Bovine Tuberculosis in Domestic Water Buffaloes (Bubalus bubalis)

Bovine tuberculosis (bTB) remains endemic in domestic water buffaloes (Bubalus bubalis) in India and elsewhere, with limited options for control other than testing and slaughter. The prescribed tuberculin skin tests with purified protein derivative (PPD) for diagnosis of bTB preclude the use of Bacille Calmette-Guérin (BCG)-based vaccination because of the antigenic cross-reactivity of vaccine strains with Mycobacterium bovis and related pathogenic members of the M. tuberculosis complex (MTBC). For the diagnosis of bTB in domestic water buffaloes, we here assessed a recently described defined-antigen skin test (DST) that comprises overlapping peptides representing the ESAT-6, CFP-10 and Rv3615c antigens, present in disease-causing members of the MTBC but missing in BCG strains. The performance characteristics of three doses (5, 10 or 20 μg/peptide) of the DST were assessed in natural tuberculin skin test reactor (n = 11) and non-reactor (n = 35) water buffaloes at an organized dairy farm in Hisar, India, and results were compared with the single intradermal skin test (SIT) using standard bovine tuberculin (PPD-B). The results showed a dose-dependent response of DST in natural reactor water buffaloes, although the SIT induced a significantly greater (P < 0.001) skin test response than the highest dose of DST used. However, using a cut-off of 2 mm or greater, the 5, 10, and 20 μg DST cocktail correctly classified eight, 10 and all 11 of the SIT-positive reactors, respectively, suggesting that the 20 μg DST cocktail has a diagnostic sensitivity (Se) of 1.0 (95% CI: 0.72–1.0) identical to that of the SIT. Importantly, none of the tested DST doses induced any measurable skin induration responses in the 35 SIT-negative animals, suggesting a specificity point estimate of 1.0 (95% CI: 0.9–1.0), also identical to that of the SIT and compares favorably with that of the comparative cervical test (Se = 0.85; 95% CI: 0.55–0.98). Overall, the results suggest that similar to tuberculin, the DST enables sensitive and specific diagnosis of bTB in water buffaloes. Future field trials to explore the utility of DST as a defined antigen replacement for tuberculin in routine surveillance programs and to enable BCG vaccination of water buffaloes are warranted.

Immunogenicity and Antigenicity of Rabbit Hepatitis E Virus-Like Particles Produced by Recombinant Baculoviruses.

Rabbit hepatitis E virus (HEV) is a novel HEV belonging to genotype 3 (HEV-3) in the Orthohepevirus A species of the genus Hepevirus, family Hepeviridae. Rabbit HEV was originally isolated from rabbits and found to cause zoonotic infection. Although rabbit HEV can be successfully grown in culture with several cell lines, including the human carcinoma cell line PLC/PRF/5, it is difficult to obtain the large amounts of viral antigen required for diagnosis and vaccine development. In this study, we expressed N-terminal 13 and 111 aa-truncated rabbit HEV ORF2 proteins using recombinant baculoviruses and obtained two types of virus-like particles (VLPs), RnVLPs and RsVLPs with ~35 and 24 nm diameter, respectively. Anti-rabbit HEV IgG antibodies were induced in high titer by immunizing rabbits with RnVLPs or RsVLPs. The antibody secretion in the serum persisted more than three years. RsVLPs showed stronger antigenic cross-reactivity against HEV-1, HEV-3 and HEV-4 than rat HEV. Moreover, anti-RsVLPs antibodies neutralized not only the cognate virus but also HEV-1, HEV-3 and HEV-4 ex vivo, indicating that rabbit HEV had the same serotype as human HEVs. In contrast, the antibody did not block rat HEV infection, demonstrating that rat HEV belonged to a different serotype. Animal experiments indicated that immunization with either RnVLPs or RsVLPs completely protected the rabbits from challenge by rabbit HEV, suggesting that the VLPs are candidates for rabbit HEV vaccine development.

Novel Surrogate Neutralizing Assay Supports Parvovirus B19 Vaccine Development for Children with Sickle Cell Disease.

Children with sickle cell disease (SCD) suffer life-threatening transient aplastic crisis (TAC) when infected with parvovirus B19. In utero, infection of healthy fetuses may result in anemia, hydrops, and death. Unfortunately, although promising vaccine candidates exist, no product has yet been licensed. One barrier to vaccine development has been the lack of a cost-effective, standardized parvovirus B19 neutralization assay. To fill this void, we evaluated the unique region of VP1 (VP1u), which contains prominent targets of neutralizing antibodies. We discovered an antigenic cross-reactivity between VP1 and VP2 that, at first, thwarted the development of a surrogate neutralization assay. We overcame the cross-reactivity by designing a mutated VP1u (VP1uAT) fragment. A new VP1uAT ELISA yielded results well correlated with neutralization (Spearman’s correlation coefficient = 0.581; p = 0.001), superior to results from a standard clinical diagnostic ELISA or an ELISA with virus-like particles. Virus-specific antibodies from children with TAC, measured by the VP1uAT and neutralization assays, but not other assays, gradually increased from days 0 to 120 post-hospitalization. We propose that this novel and technically simple VP1uAT ELISA might now serve as a surrogate for the neutralization assay to support rapid development of a parvovirus B19 vaccine.

Polyclonal antibody-based immunohistochemical detection of intraleukocytic Theileria parasites in roan and sable antelopes.

Theileria parasites commonly infect African wild artiodactyls. In rare roan (Hippotragus equinus) and sable (H. niger) antelopes, Theileria sp. (sable)-associated calf mortalities constrain breeding programs. The pathogenicity of most leukocyte-transforming Theileria spp. originates in their invasion of and multiplication in various mononuclear leukocytes, the transformation of both infected and uninfected leukocytes, and their infiltration of multiple organs. Understanding the pathogenesis of theileriosis can be improved by the use of immunohistochemistry (IHC) to identify the localization of the parasites in tissue sections. Our aim was to develop a reproducible IHC assay to detect leukocyte-associated Theileria parasites in formalin-fixed, paraffin-embedded roan and sable tissues. Polyclonal antibodies were purified from the sera of 5 roans from an area endemic for Theileria sp. (sable) and tested for IHC reactivity in 55 infected and 39 control roan and sable antelopes, and for antigen and species cross-reactivity in an additional 58 cases. The 3 strongest antibodies consistently detected intraleukocytic theilerial antigens in known positive cases in roan and sable antelopes, and also detected other Theileria spp. in non-hippotraginid wild artiodactyl tissues. The antibodies did not cross-react with other apicomplexan protozoa, with the exception of Cryptosporidium. Given that PCR on its own cannot determine the significance of theilerial infection in wild ruminants, IHC is a useful laboratory test with which to confirm the diagnosis in these species.

Identification of evolutionarily stable functional and immunogenic sites across the SARS-CoV-2 proteome and greater coronavirus family.

MotivationSince the first recognized case of COVID-19, more than 100 million people have been infected worldwide. Global efforts in drug and vaccine development to fight the disease have yielded vaccines and drug candidates to cure COVID-19. However, the spread of SARS-CoV-2 variants threatens the continued efficacy of these treatments. In order to address this, we interrogate the evolutionary history of the entire SARS-CoV-2 proteome to identify evolutionarily conserved functional sites that can inform the search for treatments with broader coverage across the coronavirus family.ResultsCombining coronavirus family sequence information with the mutations observed in the current COVID-19 outbreak, we systematically and comprehensively define evolutionarily stable sites that may provide useful drug and vaccine targets and which are less likely to be compromised by the emergence of new virus strains. Several experimentally validated effective drugs interact with these proposed target sites. In addition, the same evolutionary information can prioritize cross reactive antigens that are useful in directing multi-epitope vaccine strategies to illicit broadly neutralizing immune responses to the betacoronavirus family. Although the results are focused on SARS-CoV-2, these approaches stem from evolutionary principles that are agnostic to the organism or infective agent.Availability and implementationThe results of this work are made interactively available at http://cov.lichtargelab.org.Supplementary information Supplementary data are available at Bioinformatics online.

Electrochemical immunosensor platform based on gold-clusters, cysteamine and glutaraldehyde modified electrode for diagnosing COVID-19

Amid the global threat caused by the coronavirus disease 2019 (COVID-19) pandemic, developing sufficiently rapid, accurate, sensitive and selective methods of diagnosing both symptomatic and asymptomatic cases is essential to alleviating and controlling the pandemic’s effects. This article describes an electrochemical immunoassay platform developed to determine the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) spike antibody by using gold-clusters capped with cysteamine, glutaraldehyde, the spike protein of the SARS-CoV-2 antigen and bovine serum albumin on a glassy carbon electrode. The electrochemical oxidation signal of the antigen-based immunosensor at 0.9 V was used to detect the SARS-CoV-2 spike antibody. When saliva and oropharyngeal swab samples were analysed, the recovery and relative standard deviation values were 96.97%–101.99% and 4.99%–5.74%, respectively. The method’s limit of detection relative to the SARS-CoV-2 spike antibody in synthetic media and in saliva or oropharyngeal swab samples was 0.01 ag/mL, while the immunosensor’s linear response to the SARS-CoV-2 spike antibody varied from 0.1 to 1000 ag/mL. The cross-reactivity of the Middle East respiratory syndrome-coronavirus spike antigen was evaluated after being immobilised onto the functionalised gold-cluster based sensor, indicated that the good specifity of the produced immunosensor.

Characterization of T-Cell Responses to SMX and SMX-NO in Co-Trimoxazole Hypersensitivity Patients Expressing HLA-B*13:01.

HLA-B*13:01-positive patients in Thailand can develop frequent co-trimoxazole hypersensitivity reactions. This study aimed to characterize drug-specific T cells from three co-trimoxazole hypersensitive patients presenting with either Stevens-Johnson syndrome or drug reaction with eosinophilia and systemic symptoms. Two of the patients carried the HLA allele of interest, namely HLA-B*13:01. Sulfamethoxazole and nitroso sulfamethoxazole specific T cell clones were generated from T cell lines of co-trimoxazole hypersensitive HLA-B*13:01-positive patients. Clones were characterized for antigen specificity and cross-reactivity with structurally related compounds by measuring proliferation and cytokine release. Surface marker expression was characterized via flow cytometry. Mechanistic studies were conducted to assess pathways of T cell activation in response to antigen stimulation. Peripheral blood mononuclear cells from all patients were stimulated to proliferate and secrete IFN-γ with nitroso sulfamethoxazole. All sulfamethoxazole and nitroso sulfamethoxazole specific T cell clones expressed the CD4+ phenotype and strongly secreted IL-13 as well as IFN-γ, granzyme B and IL-22. No secretion of IL-17 was observed. A number of nitroso sulfamethoxazole-specific clones cross-reacted with nitroso dapsone but not sulfamethoxazole whereas sulfamethoxazole specific clones cross-reacted with nitroso sulfamethoxazole only. The nitroso sulfamethoxazole specific clones were activated in both antigen processing-dependent and -independent manner, while sulfamethoxazole activated T cell responses via direct HLA binding. Furthermore, activation of nitroso sulfamethoxazole-specific, but not sulfamethoxazole-specific, clones was blocked with glutathione. Sulfamethoxazole and nitroso sulfamethoxazole specific T cell clones from hypersensitive patients were CD4+ which suggests that HLA-B*13:01 is not directly involved in the iatrogenic disease observed in co-trimoxazole hypersensitivity patients.

Different Cross-Reactivities of IgM Responses in Dengue, Zika and Tick-Borne Encephalitis Virus Infections.

Flaviviruses circulate worldwide and cause a number of medically relevant human diseases, such as dengue, Zika, yellow fever, and tick-borne encephalitis (TBE). Serology plays an important role in the diagnosis of flavivirus infections, but can be impeded by antigenic cross-reactivities among flaviviruses. Therefore, serological diagnosis of a recent infection can be insufficiently specific, especially in areas where flaviviruses co-circulate and/or vaccination coverage against certain flaviviruses is high. In this study, we developed a new IgM assay format, which is well suited for the specific diagnosis of TBE, Zika and dengue virus infections. In the case of TBE and Zika, the IgM response proved to be highly specific for the infecting virus. In contrast, primary dengue virus infections induced substantial amounts of cross-reactive IgM antibodies, which is most likely explained by structural peculiarities of dengue virus particles. Despite the presence of cross-reactive IgM, the standardized nature and the quantitative read-out of the assay even allowed the serotype-specific diagnosis of recent dengue virus infections in most instances.

Comparative venomics and preclinical efficacy evaluation of a monospecific Hemachatus antivenom towards sub-Saharan Africa cobra venoms

Cobras are the most medically important elapid snakes in Africa. The African genera Naja and Hemachatus include snakes with neurotoxic and cytotoxic venoms, with shared biochemical, toxinological and antigenic characteristics. We have studied the antigenic cross-reactivity of four sub-Saharan Africa cobra venoms against an experimental monospecific Hemachatus haemachatus antivenom through comparative proteomics, preclinical assessment of neutralization, and third generation antivenomics. The venoms of H. haemachatus, N. annulifera, N. mossambica and N. nigricollis share an overall qualitative family toxin composition but depart in their proportions of three-finger toxin (3FTxs) classes, phospholipases A2 (PLA2s), snake venom metalloproteinases (SVMPs), and cysteine-rich secretory proteins (CRISPs). A monospecific anti-Hemachatus antivenom produced by Costa Rican Instituto Clodomiro Picado neutralized the lethal activity of the homologous and heterologous neuro/cytotoxic (H. haemachatus) and cyto/cardiotoxic (N. mossambica and N. nigricollis) venoms of the three spitting cobras sampled, while it was ineffective against the lethal and toxic activities of the neurotoxic venom of the non-spitting snouted cobra N. annulifera. The ability of the anti-Hemachatus-ICP antivenom to neutralize toxic (dermonecrotic and anticoagulant) and enzymatic (PLA2) activities of spitting cobra venoms suggested a closer kinship of H. haemachatus and Naja subgenus Afrocobra spitting cobras than to Naja subgenus Uraeus neurotoxic taxa. These results were confirmed by third generation antivenomics. Biological significanceAfrican Naja species represent the most widespread medically important elapid snakes across Africa. To gain deeper insight into the spectrum of medically relevant toxins, we compared the proteome of three spitting cobras (Hemachatus haemachatus, Naja mossambica and N. nigricollis) and one non-spitting cobra (N. annulifera). Three finger toxins and phospholipases A2 are the two major protein families among the venoms analyzed. The development of antivenoms of broad species coverage is an urgent need in sub-Saharan Africa. An equine antivenom raised against H. haemachatus venom showed cross-reactivity with the venoms of H. haemachatus, N. mossambica and N. nigricollis, while having poor recognition of the venom of N. annulifera. This immunological information provides clues for the design of optimum venom mixtures for the preparation of broad spectrum antivenoms.

The Case for S2: The Potential Benefits of the S2 Subunit of the SARS-CoV-2 Spike Protein as an Immunogen in Fighting the COVID-19 Pandemic.

As COVID-19 cases continue to rise, it is imperative to learn more about antibodies and T-cells produced against the causative virus, SARS-CoV-2, in order to guide the rapid development of therapies and vaccines. While much of the current antibody and vaccine research focuses on the receptor-binding domain of S1, a less-recognized opportunity is to harness the potential benefits of the more conserved S2 subunit. Similarities between the spike proteins of both SARS-CoV-2 and HIV-1 warrant exploring S2. Possible benefits of employing S2 in therapies and vaccines include the structural conservation of S2, extant cross-reactive neutralizing antibodies in populations (due to prior exposure to common cold coronaviruses), the steric neutralization potential of antibodies against S2, and the stronger memory B-cell and T-cell responses. More research is necessary on the effect of glycans on the accessibility and stability of S2, SARS-CoV-2 mutants that may affect infectivity, the neutralization potential of antibodies produced by memory B-cells, cross-reactive T-cell responses, antibody-dependent enhancement, and antigen competition. This perspective aims to highlight the evidence for the potential advantages of using S2 as a target of therapy or vaccine design.

Antibodies against HLA cross-reactivity groups: From single antigen bead assay to immunoinformatics interpretation of epitopes

Identification of anti-human leukocyte antigen (HLA) antibodies (Abs) is based on Luminex™ technology. We used bioinformatics to (i) study the correlations of mean fluorescence intensities (MFIs) for all the possible allele pairs, and (ii) determine the degree of epitope homology between HLA antigens.Using MFI data on anti-HLA Abs from 6000 Luminex™ assays, we provide an updated overview of class I and II HLA antigen cross-reactivity in which each node corresponded to an allele and each link corresponded to a strong correlation between two alleles (Spearman’s ρ > 0.8). We compared these correlations with the serological groups and the results of an epitope analysis.The strongest correlations concerned allele-specific Abs directed against the same antigen. For the HLA-A locus, the highest values of Spearman’s ρ reflected broad specificity. For the HLA-B locus, graphs defined the HLA-Bw4 public epitope, and correlations between HLA-A and -B alleles were only present for beads with the same Bw4 public epitope. For the HLA-C locus, we identified two groups that differed with regard to their KIR ligand subclassification. Lastly, the HLA-DRB1 subgroups were part of a network. In the epitope analysis, Spearman’s ρ was related to the number of matched epitopes within pairs of alleles.The combination of Spearman’s ρ with simple, undirected graphing constitutes an effective tool for understanding routinely encountered cross-reactivity profiles. Based on this model, we have implemented an online data visualization tool available at http://cusureau.pythonanywhere.com/.

Does Bordetella pertussis vaccine offer any cross-protection against Bordetella bronchiseptica? Implications for pet owners with cystic fibrosis.

The Gram-negative bacterium, Bordetella bronchiseptica, causes lower airway respiratory disease in people with cystic fibrosis (CF), as well as in companion animals, especially dogs. Presently, there are several acellular vaccines available for B.pertussis but no vaccine available for B.bronchiseptica. However given the shared protein homology between these two closely related species, we wished to explore whether pertussis vaccines may offer some cross-protection against B.bronchiseptica. Bordetella pertussis and B.bronchiseptica are closely related phylogenetically, as well as sharing protein homology in several pertussis vaccine components, including (i) pertussis toxin (PT), (ii) filamentous haemagglutinin (FHA), (iii) pertactin and (iv) fimbriae (types 2 and 3). Given that pertussis vaccine contains cross-reactive antigens with B.bronchiseptica, licensed pertussis vaccines may therefore offer cross-protection against B.bronchiseptica. Cystic fibrosis pet owners should ensure that they have an up-to-date vaccination record relating to their pertussis vaccine. Although no monovalent human pertussis vaccines are currently available, licensed non-live booster vaccines for B.pertussis are available for individuals in the age range >10years old. People with CF should ensure that they are adequately and currently protected against pertussis, to avoid whooping cough, which may also offer some cross-protection against B.bronchiseptica and therefore help further mitigate the risk of zoonotic infection of this organism from pets to their owners.

Prior infection with antigenically heterologous low pathogenic avian influenza viruses interferes with the lethality of the H5 highly pathogenic strain in domestic ducks.

Low and highly pathogenic avian influenza viruses (LPAIVs and HPAIVs, respectively) have been co-circulating in poultry populations in Asian, Middle Eastern, and African countries. In our avian-flu surveillance in Vietnamese domestic ducks, viral genes of LPAIV and HPAIV have been frequently detected in the same individual. To assess the influence of LPAIV on the pathogenicity of H5 HPAIV in domestic ducks, an experimental co-infection study was performed. One-week-old domestic ducks were inoculated intranasally and orally with phosphate-buffered saline (PBS) (control) or 106 EID50 of LPAIVs (A/duck/Vietnam/LBM678/2014 (H6N6) or A/Muscovy duck/Vietnam/LBM694/2014 (H9N2)). Seven days later, these ducks were inoculated with HPAIV (A/Muscovy duck/Vietnam/LBM808/2015 (H5N6)) in the same manner. The respective survival rates were 100% and 50% in ducks pre-infected with LBM694 or LBM678 strains and both higher than the survival of the control group (25%). The virus titers in oral/cloacal swabs of each LPAIV pre-inoculation group were significantly lower at 3–5 days post-HPAIV inoculation. Notably, almost no virus was detected in swabs from surviving individuals of the LBM678 pre-inoculation group. Antigenic cross-reactivity among the viruses was not observed in the neutralization test. These results suggest that pre-infection with LPAIV attenuates the pathogenicity of HPAIV in domestic ducks, which might be explained by innate and/or cell-mediated immunity induced by the initial infection with LPAIV.

Cross-reactive antibody against human coronavirus OC43 spike protein correlates with disease severity in COVID-19 patients: a retrospective study

ABSTRACT Seasonal human coronaviruses (HCoVs) including HCoV-229E, -OC43, -NL63, and -HKU1 widely spread in global human populations. However, the relevance of humoral response against seasonal HCoVs to COVID-19 pathogenesis is elusive. In this study, we profiled the temporal changes of IgG antibody against spike proteins (S-IgG) of SARS-CoV-2 and seasonal HCoVs in 838 plasma samples collected from 344 COVID-19 patients. We tested the antigenic cross-reactivities of S protein between SARS-CoV-2 and seasonal HCoVs and evaluated the correlations between the levels of HCoV-OC43 S-IgG and the disease severity in COVID-19 patients. We found that SARS-CoV-2 S-IgG titres mounted until days 22–28, whereas HCoV-OC43 antibody titres increased until days 15–21 and then plateaued until day 46. However, IgG titres against HCoV-NL63, −229E, and -HKU1 showed no significant increase. A two-way cross-reactivity was identified between SARS-CoV-2 and HCoV-OC43. Neutralizing antibodies against SARS-CoV-2 were not detectable in healthy controls who were positive for HCoV-OC43 S-IgG. HCoV-OC43 S-IgG titres were significantly higher in patients with severe disease than those in mild patients at days 1–21 post symptom onset (PSO). Higher levels of HCoV-OC43 S-IgG were also observed in patients requiring mechanical ventilation. At days 1–10 PSO, HCoV-OC43 S-IgG titres correlated to disease severity in the age group over 60. Our data indicate that there is a correlation between cross-reactive antibody against HCoV-OC43 spike protein and disease severity in COVID-19 patients.

Immunogenicity of antigens from outer membrane vesicles of Neisseria meningitidis associated with bilayer fragment of dioctadecyldimethylammonium in Swiss adult mice.

PurposeIn the present study, meningococcal serogroup B outer membrane vesicles (OMVs) were associated with bilayer fragments of a cationic lipid, dioctadecyldimethylammonium (DDA-BF), used as adjuvant, in an antigenic preparation tested in adult female outbred mice. This adjuvant was compared to the traditional adjuvant aluminum hydroxide.Materials and MethodsThe potential in generating humoral response was evaluated by enzyme-linked immunosorbent assay (ELISA). Individual serum was collected and immunoglobulin G (IgG), IgG1, IgG2a, and IgG2b were quantified. Analyses were carried out 15 and 60 days after immunization. Antibodies avidity index were also analyzed by ELISA. Immunoblot and dot-ELISA were carried out to evaluate specific reaction for homologous strains and cross-reactive antigens present in other meningococcal strains isolated in 2011–2012 year, in Brazil. Delayed type hypersensitivity was used as indicative of cellular immunity and compared between two experimental groups, 24 hours after homologous strain challenge.ResultsThe OMVs of Neisseria meningitidis, and N. lactamica (related species) were characterized by electrophoretic separation of proteins in 13% polyacrylamide gel. The strains presented antigens in the range of 8 to 130 kDa, showing a heterogeneous protein migration pattern. In the group immunized with OMVs/DDA-BF, we found no significant production of total IgG 15 days after the first immunization. On the other hand, 60 days after first immunization both adjuvants act benefiting total IgG production similarly. The antibodies of the IgG isotype produced by animals immunized after one or two doses after first immunization, showed intermediate and high avidity, independent on the adjuvant used. In both experimental groups the swelling of the footpads was significantly higher than those of the controls, suggesting that only one dose was enough to stimulate the generation of cellular immunity.ConclusionThe use of this cationic adjuvant for N. meningitidis OMVs preparation revealed good potential for future new antigen preparation for N. meningitidis vaccine.

Bet v 1 potential allergens are involved in anthracnose resistance of strawberry varieties

BACKGROUND: Bet v 1 family identified as one major plant food allergen class, is highly homologous to pathogenesis-related protein 10 (PR-10), but its biological function involved in disease resistance is still unclear. OBJECTIVE: This study aims to investigate whether Bet v 1 potential allergens participate in the resistance of berry crops against fungal pathogen. METHODS: Allergenicity of Bet v 1 proteins in strawberry (Fragaria) was evaluated by bioinformatics methods. Their expression in response to anthracnose and between susceptible and resistance varieties was analyzed. RESULTS: 19 Bet v 1 homologous proteins were identified and 15 of them were considered as allergen candidates. RNA-seq analysis indicated most of these Fra a 1s expressed in fruits could be largely induced by the invasion of anthracnose pathogen Colletotrichum. The mRNA level of fruit major allergen Fra a 1.05 in the resistant variety Shenyang (SY) was 20∼50 fold higher compared with those in the susceptible cultivar and two diploid wild species. Immunoblotting using Birch (Betula pendula) Bet v 1 allergen-specific IgG antibody confirmed the large-scale accumulation of potential cross-reactive antigens in SY fruit. CONCLUSIONS: Strawberry Bet v 1 potential allergens exhibit their correspondence with anthracnose resistance that might be instructive to future breeding strategies.

Post-Vaccination Yellow Fever Antiserum Reduces Zika Virus in Embryoid Bodies When Placental Cells are Present.

Zika virus (ZIKV) is a flavivirus that originated in Africa but emerged in Latin America in 2015. In this region, other flaviviruses such as Dengue (DENV), West Nile, and Yellow Fever virus (YFV) also circulate, allowing for possible antigenic cross-reactivity to impact viral infections and immune responses. Studies have found antibody-mediated enhancement between DENV and ZIKV, but the impact of YFV antibodies on ZIKV infection has not been fully explored. ZIKV infections cause congenital syndromes, such as microcephaly, necessitating further research into ZIKV vertical transmission through the placental barrier. Recent advancements in biomedical engineering have generated co-culture methods that allow for the in vitro recapitulation of the maternal–fetal interface. This study utilized a transwell assay, which was a co-culture model utilizing human placental syncytiotrophoblasts, fetal umbilical cells, and a differentiating embryoid body, to replicate the maternal–fetal axis. To determine if cross-reactive YFV vaccine antibodies impacted the pathogenesis of ZIKV across the maternal–fetal axis, syncytiotrophoblasts were inoculated with ZIKV or ZIKV incubated with YFV vaccine antisera, and the viral load was measured 72 h post-inoculation. Here, we report that BeWo and HUVEC cells were permissive to ZIKV and that the impact of YFV post-vaccination antibodies on ZIKV replication was cell line-dependent. Embryoid bodies were also permissive to ZIKV, and the presence of YFV antibodies collected 4–14 months post-vaccination reduced ZIKV infection when placental cells were present. However, when directly infected with ZIKV, the embryoid bodies displayed significantly increased viral loads in the presence of YFV antiserum taken 30 days post-vaccination. The data show that each of the cell lines and EBs have a unique response to ZIKV complexed with post-vaccination serum, suggesting there may be cell-specific mechanisms that impact congenital ZIKV infections. Since ZIKV infections can cause severe congenital syndromes, it is crucial to understand any potential enhancement or protection offered from cross-reactive, post-vaccination antibodies.

A PIONEER USAGE OF GOLD NANOPARTICLES IN SANDWICH ELISA BASED ON CIRCULATING HYDATID ANTIGEN AS A TOOL FOR IMPROVEMENT OF SERODIAGNOSTIC TESTING

Human cystic echinococcosis is a zoonosis occurring due to dogs handling and exposure toEchinococcus granulosus ova in their stools being an accidental intermediate host. Serologyremains the only sure and important tool to diagnose this disease. We aimed to improve thedetection of circulating antigen by sandwich ELISA through its binding to gold nanoparticleand proving the role of minute nanoparticles in serodiagnosis. Gold nanoparticles (AuNPs)were employed for the capture of the antigens of protoscolices (pAg) in the patient’s serum bysandwich ELISA. Cross-reactivity for antigens from Hymenolepis nana, Entrobius vermicularis,and Fasciola gigantica was ruled out by using anti-protoscolices polyclonal IgG antibodies(ppAb). Antigen of sonicated protoscolices which were removed from camel lungcysts, was purified and injected to a New-zealand white rabbit giving ppAb which then loadedon AuNPs being used as a diagnostic indicator for circulating pAg by both sandwich ELISAand nanogold sandwich ELISA techniques. Nanogold sandwich ELISA was able to give positiveresults with 96.3% of hydatid patients and 5% of non-hydatid patients while sandwichELISA showed 81.4% and 20% positive cases of the same groups respectively. The sensitivityand specificity of nanogold sandwich ELISA were 96.3% & 95% against 81.5% & 80% forthose of sandwich ELISA respectively. The conjugation of AuNPs to anti-Echinococcus IgGantibodies seems to be beneficial to increase PPV, NPV, and efficiency and reduce the overallcost of the assay as less antibody was needed.

Serological methods for detection of infection with shrew-borne hantaviruses: Thottapalayam, Seewis, Altai, and Asama viruses

The infectivity of shrew-borne hantaviruses to humans is still unclear because of the lack of a serodiagnosis method for these viruses. In this study, we prepared recombinant nucleocapsid (rN) proteins of Seewis orthohantavirus, Altai orthohantavirus (ALTV), Thottapalayam thottimvirus (TPMV), and Asama orthohantavirus. Using monospecific rabbit sera, no antigenic cross-reactivity was observed. In a serosurvey of 104 samples from renal patients and 271 samples from heathy controls from Sri Lanka, one patient serum and two healthy control sera reacted with rN proteins of ALTV and TPMV, respectively. The novel assays should be applied to investigate potential infectivity of shrew-borne hantaviruses to humans.