Molecular and Antigenic Characterization of Avian H9N2 Viruses in Southern China

The H9N2 low pathogenicity avian influenza (LPAI) virus has become endemic in poultry globally. In several Asian countries, vaccination against H9N2 avian influenza virus (AIV) was approved to reduce economic losses in the poultry industry.

Identification and Characterization of P0 Protein as a Vaccine Candidate Against Babesia divergens, Blood Parasite of Veterinary and Zoonotic Importance

The molecular identification and antigenic characterization of P0 protein in Babesia divergens, a blood parasite of veterinary and zoonotic importance, were carried out in this study for use in developing subunit vaccines against B. divergens infection. Recombinant protein encoding P0 (BdP0) was developed in Escherichia coli, and its antiserum was generated in mice for further molecular characterization. Anti-rBdP0 serum had a specific interaction with the corresponding legitimate B. divergens protein, as confirmed by Western blotting and indirect fluorescent antibody tests. ELISA was used to assess the immunogenicity of BdP0 in a group of 68 bovine field samples, and significant immunological reactivity was found in 19 and 20 positive samples of rBdp0 and B. divergens lysate, respectively. The in vitro growth of B. divergens cultures treated with anti-rBdP0 serum was significantly inhibited (p < 0.05). Furthermore, after 6 h of incubation with 2 mg/ml anti-rBdP0 serum, the ability of pre-incubated free merozoites to invade bovine erythrocytes was reduced by 59.88%. The obtained data suggest the possible use of rBdP0 as diagnostic antigen and may serve as a vaccine candidate against babesiosis caused by B. divergens either in animal or human.

ArVirInd - a database of arboviral antigenic proteins from the Indian subcontinent

Studies on antigenic proteins for arboviruses are important for providing diagnostics and vaccine development. India and its neighbouring countries have huge burden of arboviral diseases. Data mining for country-specific sequences from existing databases is cumbersome and time-consuming. This necessitated the development of a database of antigenic proteins from arbo-viruses isolated from the countries of the Indian subcontinent. Arboviral antigenic protein sequences were obtained from the NCBI and other databases. In silico antigenic characterization was performed (Epitope predictions) and data incorporated in the database. The front end is designed and developed using HTML, CSS and PHP. For the backend of the database, we have used MySQL. A database, named ArVirInd, is created as a repository of information on antigenic proteins. This enlists sequences by country and year of outbreak or origin of the viral strain. For each entry antigenic information is provided along with functional sites, etc. Researchers can search this database by virus/protein name, country and year of collection (or in combination). It is available publicly via Internet at http://www.arvirind.co.in. ArVirInd will be useful in the study of immuno-informatics, diagnostics and vaccinology for arboviruses.

Glycan remodeled erythrocytes facilitate antigenic characterization of recent A/H3N2 influenza viruses

During circulation in humans and natural selection to escape antibody recognition for decades, A/H3N2 influenza viruses emerged with altered receptor specificities. These viruses lost the ability to agglutinate erythrocytes critical for antigenic characterization and give low yields and acquire adaptive mutations when cultured in eggs and cells, contributing to recent vaccine challenges. Examination of receptor specificities of A/H3N2 viruses reveals that recent viruses compensated for decreased binding of the prototypic human receptor by recognizing α2,6-sialosides on extended LacNAc moieties. Erythrocyte glycomics shows an absence of extended glycans providing a rationale for lack of agglutination by recent A/H3N2 viruses. A glycan remodeling approach installing functional receptors on erythrocytes, allows antigenic characterization of recent A/H3N2 viruses confirming the cocirculation of antigenically different viruses in humans. Computational analysis of HAs in complex with sialosides having extended LacNAc moieties reveals that mutations distal to the RBD reoriented the Y159 side chain resulting in an extended receptor binding site.

Epidemiology of Pestivirus H in Brazil and Its Control Implications

Along with viruses in the Pestivirus A (Bovine Viral Diarrhea Virus 1, BVDV1) and B species (Bovine Viral Diarrhea Virus 2, BVDV2), members of the Pestivirus H are mainly cattle pathogens. Viruses belonging to the Pestivirus H group are known as HoBi-like pestiviruses (HoBiPev). Genetic and antigenic characterization suggest that HoBiPev are the most divergent pestiviruses identified in cattle to date. The phylogenetic analysis of HoBiPev results in at least five subgroups (a–e). Under natural or experimental conditions, calves infected with HoBiPev strains typically display mild upper respiratory signs, including nasal discharge and cough. Although BVDV1 and BVDV2 are widely distributed and reported in many South American countries, reports of HoBiPev in South America are mostly restricted to Brazil. Despite the endemicity and high prevalence of HoBiPev in Brazil, only HoBiPev-a was identified to date in Brazil. Unquestionably, HoBiPev strains in BVDV vaccine formulations are required to help curb HoBiPev spread in endemic regions. The current situation in Brazil, where at this point only HoBiPev-a seems present, provides a more significant opportunity to control these viruses with the use of a vaccine with a single HoBiPev subtype. Despite the lack of differentiation among bovine pestiviruses by current BVDV tests, the reduced genetic variability of HoBiPev in Brazil may allow reliable identification of cases within the region. On the other hand, introducing foreign ruminants, biologicals, and genetic material to South America, especially if it originated from other HoBiPev-endemic countries, should consider the risk of introducing divergent HoBiPev subtypes.

Antigenic Characterization of ORF2 and ORF3 Proteins of Hepatitis E Virus (HEV)

To evaluate the antigenic properties of Hepatitis E Virus (HEV) Open Reading Frame 2 and 3 (ORF2 and ORF3) codified proteins, we expressed different portions of ORF2 and the entire ORF3 in E. coli, a truncated ORF2, was also expressed in baculovirus. A panel of 37 monoclonal antibodies (MAbs) was raised against ORF2 (1–660 amino acids) and MAbs were mapped and characterized using the ORF2 expressed portions. Selected HEV positive and negative swine sera were used to evaluate ORF2 and ORF3 antigens’ immunogenicity. The MAbs were clustered in six groups identifying six antigenic regions along the ORF2. Only MAbs binding to the sixth ORF2 antigenic region (394–608 aa) were found to compete with HEV positive sera and efficiently catch the recombinant antigen expressed in baculovirus. The ORF2 portion from 394–608 aa demonstrated to include most immunogenic epitopes with 85% of HEV positive swine sera reacting against the region from 461–544 aa. Only 5% of the selected HEV sera reacted against the ORF3 antigen.

Genetic and Antigenic Characterization of Avian Avulavirus Type 6 (AAvV-6) Circulating in Canadian Wild Birds (2005–2017)

We describe for the first time the genetic and antigenic characterization of 18 avian avulavirus type-6 viruses (AAvV-6) that were isolated from wild waterfowl in the Americas over the span of 12 years. Only one of the AAvV-6 viruses isolated failed to hemagglutinate chicken red blood cells. We were able to obtain full genome sequences of 16 and 2 fusion gene sequences from the remaining 2 isolates. This is more than double the number of full genome sequences available at the NCBI database. These AAvV-6 viruses phylogenetically grouped into the 2 existing AAvV-6 genotype subgroups indicating the existence of an intercontinental epidemiological link with other AAvV-6 viruses isolated from migratory waterfowl from different Eurasian countries. Antigenic maps made using HI assay data for these isolates showed that the two genetic groups were also antigenically distinct. An isolate representing each genotype was inoculated in specific pathogen free (SPF) chickens, however, no clinical symptoms were observed. A duplex fusion gene based real-time assay for the detection and genotyping of AAvV-6 to genotype 1 and 2 was developed. Using the developed assay, the viral shedding pattern in the infected chickens was examined. The chickens infected with both genotypes were able to shed the virus orally for about a week, however, no significant cloacal shedding was detected in chickens of both groups. Chickens in both groups developed detectable levels of anti-hemagglutinin antibodies 7 days after infection.

Antigenic cartography reveals complexities of genetic determinants that lead to antigenic differences among pandemic GII.4 noroviruses

Noroviruses are the predominant cause of acute gastroenteritis, with a single genotype (GII.4) responsible for the majority of infections. This prevalence is characterized by the periodic emergence of new variants that present substitutions at antigenic sites of the major structural protein (VP1), facilitating escape from herd immunity. Notably, the contribution of intravariant mutations to changes in antigenic properties is unknown. We performed a comprehensive antigenic analysis on a virus-like particle panel representing major chronological GII.4 variants to investigate diversification at the inter- and intravariant level. Immunoassays, neutralization data, and cartography analyses showed antigenic similarities between phylogenetically related variants, with major switches to antigenic properties observed over the evolution of GII.4 variants. Genetic analysis indicated that multiple coevolving amino acid changes—primarily at antigenic sites—are associated with the antigenic diversification of GII.4 variants. These data highlight complexities of the genetic determinants and provide a framework for the antigenic characterization of emerging GII.4 noroviruses.