Does immune recognition of SARS-CoV2 epitopes vary between different ethnic groups?

The SARS-CoV2 mediated Covid-19 pandemic has impacted humankind at an unprecedented scale. While substantial research efforts have focused towards understanding the mechanisms of viral infection and developing vaccines/ therapeutics, factors affecting the susceptibility to SARS-CoV2 infection and manifestation of Covid-19 remain less explored. Given that the Human Leukocyte Antigen (HLA) system is known to vary among ethnic populations, it is likely to affect the recognition of the virus, and in turn, the susceptibility to Covid-19. To understand this, we used bioinformatic tools to probe all SARS-CoV2 peptides which could elicit T-cell response in humans. We also tried to answer the intriguing question of whether these potential epitopes were equally immunogenic across ethnicities, by studying the distribution of HLA alleles among different populations and their share of cognate epitopes. Results indicate that the immune recognition potential of SARS-CoV2 epitopes tend to vary between different ethnic groups. While the South Asians are likely to recognize higher number of CD8-specific epitopes, Europeans are likely to identify higher number of CD4-specific epitopes. We also hypothesize and provide clues that the newer mutations in SARS-CoV2 are unlikely to alter the T-cell mediated immunogenic responses among the studied ethnic populations. The work presented herein is expected to bolster our understanding of the pandemic, by providing insights into differential immunological response of ethnic populations to the virus as well as by gaging the possible effects of mutations in SARS-CoV2 on efficacy of potential epitope-based vaccines through evaluating ∼40,000 viral genomes.

Complexity of type-specific 56 kDa antigen CD4 T-cell epitopes of Orientia tsutsugamushi strains causing scrub typhus in India.

Orientia tsutsugamushi (Ots) is an obligate, intracellular, mite-transmitted human pathogen which causes scrub typhus. Understanding the diversity of Ots antigens is essential for designing specific diagnostic assays and efficient vaccines. The protective immunodominant type-specific 56 kDa antigen (TSA) of Ots varies locally and across its geographic distribution. TSA contains four hypervariable domains. We bioinformatically analyzed 345 partial sequences of TSA available from India, most of which contain only the three variable domains (VDI-III) and three spacer conserved domains (SVDI, SVDII/III, SVDIII). The total number (152) of antigenic types (amino acid variants) varied from 14-36 in the six domains of TSA that we studied. Notably, 55% (787/1435) of the predicted CD4 T-cell epitopes (TCEs) from all the six domains had high binding affinities (HBA) to at least one of the prevalent Indian human leukocyte antigen (HLA) alleles. A surprisingly high proportion (61%) of such TCEs were from spacer domains; indeed 100% of the CD4 TCEs in the SVDI were HBA. TSA sequences from India had more antigenic types (AT) than TSA from Korea. Overall, >90% of predicted CD4 TCEs from spacer domains were predicted to have HBA against one or more prevalent HLA types from Indian, Korean, Asia-Pacific region or global population data sets, while only <50% of CD4 TCEs in variable domains exhibited such HBA. The phylogenetically and immunologically important amino acids in the conserved spacer domains were identified. Our results suggest that the conserved spacer domains are predicted to be functionally more important than previously appreciated in immune responses to Ots infections. Changes occurring at the TCE level of TSA may contribute to the wide range of pathogenicity of Ots in humans and mouse models. CD4 T-cell functional experiments are needed to assess the immunological significance of these HBA spacer domains and their role in clearance of Ots from Indian patients.

The impact of amino acid variability on alloreactivity defines a functional distance predictive of permissive HLA-DPB1 mismatches in hematopoietic stem cell transplantation.

A major challenge in unrelated hematopoietic stem cell transplantation (HSCT) is the prediction of permissive HLA mismatches, ie, those associated with lower clinical risks compared to their nonpermissive counterparts. For HLA-DPB1, a clinically prognostic model has been shown to be matching for T cell epitope (TCE) groups assigned by cross reactivity of T cells alloreactive to HLA-DPB1∗09:01; however, the molecular basis of this observation is not fully understood. Here, we have mutated amino acids (aa) in 10 positions of HLA-DPB1∗09:01 to other naturally occurring variants, expressed them by lentiviral vectors in B cell lines, and quantitatively measured allorecognition by 17 CD4(+) T cell effectors from 6 unrelated individuals. A significant impact on the median alloresponse was observed for peptide contact positions 9, 11, 35, 55, 69, 76, and 84, but not for positions 8, 56, and 57 pointing away from the groove. A score for the "functional distance" (FD) from HLA-DPB1∗09:01 was defined as the sum of the median impact of polymorphic aa in a given HLA-DPB1 allele on T cell alloreactivity. Established TCE group assignment of 23 alleles correlated with FD scores of ≤0.5, 0.6 to 1.9 and ≥2 for TCE groups 1, 2, and 3, respectively. Based on this, prediction of TCE group assignment will be possible for any given HLA-DPB1 allele, including currently 367 alleles encoding distinct proteins for which T cell cross reactivity patterns are unknown. Experimental confirmation of the in silico TCE group classification was successfully performed for 7 of 7 of these alleles. Our findings have practical implications for the applicability of TCE group matching in unrelated HSCT and provide new insights into the molecular mechanisms underlying this model. The innovative concept of FD opens new potential avenues for risk prediction in unrelated HSCT.

Analysis of the effects of polymorphism on pollen profilin structural functionality and the generation of conformational, T- and B-cell epitopes.

An extensive polymorphism analysis of pollen profilin, a fundamental regulator of the actin cytoskeleton dynamics, has been performed with a major focus in 3D-folding maintenance, changes in the 2-D structural elements, surface residues involved in ligands-profilin interactions and functionality, and the generation of conformational and lineal B- and T-cell epitopes variability. Our results revealed that while the general fold is conserved among profilins, substantial structural differences were found, particularly affecting the special distribution and length of different 2-D structural elements (i.e. cysteine residues), characteristic loops and coils, and numerous micro-heterogeneities present in fundamental residues directly involved in the interacting motifs, and to some extension these residues nearby to the ligand-interacting areas. Differential changes as result of polymorphism might contribute to generate functional variability among the plethora of profilin isoforms present in the olive pollen from different genetic background (olive cultivars), and between plant species, since biochemical interacting properties and binding affinities to natural ligands may be affected, particularly the interactions with different actin isoforms and phosphoinositides lipids species. Furthermore, conspicuous variability in lineal and conformational epitopes was found between profilins belonging to the same olive cultivar, and among different cultivars as direct implication of sequences polymorphism. The variability of the residues taking part of IgE-binding epitopes might be the final responsible of the differences in cross-reactivity among olive pollen cultivars, among pollen and plant-derived food allergens, as well as between distantly related pollen species, leading to a variable range of allergy reactions among atopic patients. Identification and analysis of commonly shared and specific epitopes in profilin isoforms is essential to gain knowledge about the interacting surface of these epitopes, and for a better understanding of immune responses, helping design and development of rational and effective immunotherapy strategies for the treatment of allergy diseases.

Characterization of the allergic T-cell response to Pru p 3, the nonspecific lipid transfer protein in peach.

BACKGROUND: Pru p 3, the nonspecific lipid transfer protein from peach, is an important plant food allergen that frequently induces systemic reactions. OBJECTIVE: We sought to analyze the allergic T-cell response to Pru p 3. METHODS: PBMCs from Italian and Spanish patients with peach allergy were stimulated with purified natural Pru p 3. Allergen-specific T-cell lines were used to identify T-cell epitopes of Pru p 3. Pru p 3-specific T-cell clones (TCCs) were analyzed for allergen-induced secretion of IL-4, IFN-gamma, and IL-10 and expression of the integrin beta7, a receptor critical for gut homing. RESULTS: No difference in T-cell responses of Italian and Spanish patients was found. Among several T cell-activating regions, Pru p 3(13-27), Pru p 3(34-48), Pru p 3(43-57), and Pru p 3(61-75) were most frequently recognized in 18 Pru p 3-specific T-cell lines. The majority of 32 Pru p 3-specific TCCs belonged to the T(H)2 subset. In contrast to TCCs specific for other plant food and pollen allergens, only a limited number of Pru p 3-specific TCCs produced significant amounts of IL-10. The expression of integrin beta7 on Pru p 3-specific TCCs was comparable with that observed on peanut-specific TCCs and higher compared with that seen in different pollen-specific TCCs. CONCLUSION: The T-cell response to Pru p 3 is dominated by T(H)2 cells presumably primed in the gut. The identification of relevant T cell-activating regions provides a basis for engineering hypoallergenic variants of Pru p 3 with less IgE binding and retained T-cell stimulatory capacity for safe immunotherapy of peach allergy.

Inter-clade cross-reactivity of HIV-1-specific T cell responses in human immunodeficiency virus type 1 infection in China.

To determine the degree of HIV-1-specific cytotoxic-T-lymphocyte (CTL) cross-responses to the clade B and C consensus sequences at the single peptide level. We assessed CTL responses in 46 HIV-1 clade B chronically infected individuals using an interferon-gamma Elispot assay with a total of 826 overlapping peptides spanning HIV-1 clade B and C consensus sequences. In general, 583 peptides were recognized by HIV-1-specific T cells in the study subjects (292 clade B, 291 clade C respectively), of which 204 peptides in both clades were recognized simultaneously. The HIV-1-specific CTL responses to both clade peptides contributed 54.23% (954/1759) to the total responses. No significant difference was observed between the overall magnitude or frequency of CTL responses to clade B proteins and those to clade C proteins. According to the profiles of CTL magnitude and CTL frequency, the top 44 and 35 synthetic peptides were identified as immunodominant regions in the clade B and C consensus sequences respectively and 27 corresponding peptides in two immunodominant regions were cross-reactive. These peptides with cross-reactivity had a significantly higher ability to elicit CTL responses (P< 0.01) and preferentially had a trend of lower entropy and higher inter-clade homology. A wide degree of cross-clade reactivity of HIV-1-specific T cells exist in clade B and clade C variants. Most of immunodominant peptides with cross-reactivity are vigorous to elicit CTL responses and preferentially be conservative. This result may make future HIV-1 vaccines including multiple copies of CTL epitopes in these immunodominant peptides effective for this population.

Towards the in silico identification of class II restricted T-cell epitopes: a partial least squares iterative self-consistent algorithm for affinity prediction.

MOTIVATION: The immunogenicity of peptides depends on their ability to bind to MHC molecules. MHC binding affinity prediction methods can save significant amounts of experimental work. The class II MHC binding site is open at both ends, making epitope prediction difficult because of the multiple binding ability of long peptides. RESULTS: An iterative self-consistent partial least squares (PLS)-based additive method was applied to a set of 66 peptides no longer than 16 amino acids, binding to DRB1*0401. A regression equation containing the quantitative contributions of the amino acids at each of the nine positions was generated. Its predictability was tested using two external test sets which gave r(pred) = 0.593 and r(pred) = 0.655, respectively. Furthermore, it was benchmarked using 25 known T-cell epitopes restricted by DRB1*0401 and we compared our results with four other online predictive methods. The additive method showed the best result finding 24 of the 25 T-cell epitopes. AVAILABILITY: Peptides used in the study are available from http://www.jenner.ac.uk/JenPep. The PLS method is available commercially in the SYBYL molecular modelling software package. The final model for affinity prediction of peptides binding to DRB1*0401 molecule is available at http://www.jenner.ac.uk/MHCPred. Models developed for DRB1*0101 and DRB1*0701 also are available in MHCPred.

A study of genetic diversity in the gene encoding the circumsporozoite protein (CSP) of Plasmodium falciparum from different transmission areas--XVI. Asembo Bay Cohort Project.

We have investigated the genetic diversity of the gene encoding the CS protein. A total of 75 complete and 96 partial sequences are studied. We find high levels of genetic polymorphisms as evidenced by 50 and 24 alleles at the Th2R and Th3R epitopes, respectively. Overall, we find that African isolates are more polymorphic as compared with parasites from other geographic regions. We conclude that the uneven geographic polymorphism may have an adverse impact on the effectiveness of vaccines based on this antigen alone. We find extensive polymorphism in the repeat allotypes, or RATs. In order to explore how the protein structure may impose restrictions in the number of repeats, we have simulated the stability of the structure of the tandem repeat region. Our analysis suggests that the protein structure may play an important role in the observed polymorphism in the number of CS repeats in Plasmodium falciparum. We explored the linkage and recombination events among the polymorphic sites. We found that putative recombination events overlap with linked sites. We discuss how this pattern is explained by the action of positive natural selection, where the recombination events detected are convergent mutations. We conclude that it is inappropriate to use linkage-recombination patterns on genes under positive selection for assessing the structure of parasite populations.

Ribosomal scanning past the primary initiation codon as a mechanism for expression of CTL epitopes encoded in alternative reading frames.

An increasing amount of evidence has shown that epitopes restricted to MHC class I molecules and recognized by CTL need not be encoded in a primary open reading frame (ORF). Such epitopes have been demonstrated after stop codons, in alternative reading frames (RF) and within introns. We have used a series of frameshifts (FS) introduced into the Influenza A/PR/8 /34 nucleoprotein (NP) gene to confirm the previous in vitro observations of cryptic epitope expression, and show that they are sufficiently expressed to prime immune responses in vivo. This presentation is not due to sub-dominant epitopes, transcription from cryptic promoters beyond the point of the FS, or internal initiation of translation. By introducing additional mutations to the construct exhibiting the most potent presentation, we have identified initiation codon readthrough (termed scanthrough here, where the scanning ribosome bypasses the conventional initiation codon, initiating translation further downstream) as the likely mechanism of epitope production. Further mutational analysis demonstrated that, while it should operate during the expression of wild-type (WT) protein, scanthrough does not provide a major source of processing substrate in our system. These findings suggest (i) that the full array of self- and pathogen-derived epitopes available during thymic selection and infection has not been fully appreciated and (ii) that cryptic epitope expression should be considered when the specificity of a CTL response cannot be identified or in therapeutic situations when conventional CTL targets are limited, as may be the case with latent viral infections and transformed cells. Finally, initiation codon readthrough provides a plausible explanation for the presentation of exocytic proteins by MHC class I molecules.

Comparison of peptides eluted from the groove of HLA-B27 from Salmonella infected and non-infected cells.

Reactive arthritis (ReA) is associated with the MHC class-I molecule HLA-B27 and caused by certain Gram-negative bacteria. The mechanism by which HLA-B27 confers a higher susceptibility for this disease compared to other MHC Class-I alleles is still not known. We investigated whether infection of human HLA-B27+ cells is able to change the peptide repertoire presented by these HLA-B27 molecules. To this end large quantities of a B-cell line (C1R-B27) transfected with HLA-B2705 were infected with S. typhimurium. Peptides were eluted from the B27 molecules and separated by Reversed Phase Chromatography (RPC). We then compared the peptide profiles obtained from S. typhimurium infected CIR B-cells with that obtained from non infected cells. Apart from a few additional peaks present in the profile derived from the infected batch the peptide profiles were almost identical. A few fractions were subjected to sequencing by Edman degradation. All peptides found were nonameres with arginine (Arg) at position 2 which is in agreement with the previously described HLA-B27 peptide binding motif. The majority of peaks expressed a mixture of at least four different peptides. The analysis of differences between HLA-B27 bound peptides from Salmonella infected and non infected cells might lead to the identification of T-cell epitopes shared by Salmonella and autoantigens.