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Molecular HLA typing techniques are currently undergoing a rapid evolution. While real‐time PCR is established as the standard method in tissue typing laboratories regarding allocation of solid organs, next generation sequencing (NGS) for high‐resolution HLA typing is becoming indispensable but is not yet suitable for deceased donors. By contrast, high‐resolution typing is essential for stem cell transplantation and is increasingly required for questions relating to various disease associations. In this multicentre clinical study, the TGS technique using nanopore sequencing is investigated applying NanoTYPE™ kit and NanoTYPER™ software (Omixon Biocomputing Ltd., Budapest, Hungary) regarding the concordance of the results with NGS and its practicability in diagnostic laboratories. The results of 381 samples show a concordance of 99.58% for 11 HLA loci, HLA‐A, ‐B, ‐C, ‐DRB1, ‐DRB3, ‐DRB4, ‐DRB5, ‐DQA1, ‐DQB1, ‐DPA1 and ‐DPB1. The quality control (QC) data shows a very high quality of the sequencing performed in each laboratory, 34,926 (97.15%) QC values were returned as 'passed', 862 (2.4%) as 'inspect' and 162 (0.45%) as 'failed'. We show that an 'inspect' or 'failed' QC warning does not automatically lead to incorrect HLA typing. The advantages of nanopore sequencing are speed, flexibility, reusability of the flow cells and easy implementation in the laboratory. There are challenges, such as exon coverage and the handling of large amounts of data. Finally, nanopore sequencing presents potential for applications in basic research within the field of epigenetics and genomics and holds significance for clinical concerns. [ABSTRACT FROM AUTHOR]

Objectives To quantitatively profile the T-cell repertoire in the peripheral blood of individuals genetically at risk for RA, namely first-degree relatives of RA patients (RA-FDR) at different phases of disease development. Methods Next-generation sequencing of the TCR CDR3β repertoire was performed on genomic DNA isolated from whole blood samples of RA-FDR selected at three different pre-clinical stages and of matched RA patients (n  = 20/group). T-cell clones were identified by their unique sequence and their degree of expansion (frequency) within each sample was characterized. Clones with a frequency over 0.5% were considered highly expanded clones (HEC). Results The absolute number of HEC was significantly higher in established RA patients (mean 4.65) and tended to be higher in symptomatic RA-FDR (mean 3.4) compared with asymptomatic RA-FDR (mean 1.55, P  =0.003 and P  =0.07, respectively). Asymptomatic individuals with high levels of ACPA did not differ from asymptomatic RA-FDR in terms of absolute number and frequency of clones. The number of HEC tended to be slightly higher at the time of RA onset (P  =0.055). Neither clones shared by several patients, nor clones previously associated with RA, were preferentially present within or between the different groups. Finally, a longitudinal analysis did not allow to uncover a kinetic expansion of RA-specific clones closely correlated with disease development. Conclusions HEC were detected in the peripheral blood before the clinical onset of RA, in particular in the later pre-clinical phase of RA development, and their presence increased over time. [ABSTRACT FROM AUTHOR]

The novel HLA‐B*15:514 allele was characterized using two next‐generation sequencing technologies. [ABSTRACT FROM AUTHOR]

Background & Aims The interaction of killer cell immunoglobulin-like receptors with their human leucocyte antigen ligands drives the activation and inhibition of natural killer cells. Natural killer cells could be implicated in the development of liver fibrosis in chronic hepatitis C. Methods We analysed 206 non-transplanted and 53 liver transplanted patients, selected according to their Metavir fibrosis stage. Several variables such as the number of activator killer cell immunoglobulin-like receptors or the human leucocyte antigen ligands were considered in multinomial and logistic regression models. Possible confounding variables were also investigated. Results The killer cell immunoglobulin-like receptors were not significant predictors of the fibrosis stage. Conversely, a significant reduction of the human leucocyte antigen-C1C2 genotype was observed in the most advanced fibrosis stage group (F4) in both cohorts. Furthermore, the progression rate of fibrosis was almost 10 times faster in the subgroup of patients after liver transplantation, and human leucocyte antigen-C1C2 was significantly reduced in this cohort compared with non-transplanted patients. Conclusion This study suggests a possible role of killer cell immunoglobulin-like receptors and their ligands in the development of liver damage. The absence of C1 and C2 ligands heterozygosity could lead to less inhibition of natural killer cells and a quicker progression to a high level of fibrosis in patients infected with hepatitis C virus, especially following liver transplantation. [ABSTRACT FROM AUTHOR]

Supertypes are groups of human leukocyte antigen (HLA) alleles which bind overlapping sets of peptides due to sharing specific residues at the anchor positions-the B and F pockets-of the peptide-binding region (PBR). HLA alleles within the same supertype are expected to be functionally similar, while those from different supertypes are expected to be functionally distinct, presenting different sets of peptides. In this study, we applied the supertype classification to the HLA-A and HLA-B data of 55 worldwide populations in order to investigate the effect of natural selection on supertype rather than allelic variation at these loci. We compared the nucleotide diversity of the B and F pockets with that of the other PBR regions through a resampling procedure and compared the patterns of within-population heterozygosity ( He) and between-population differentiation ( G) observed when using the supertype definition to those estimated when using randomized groups of alleles. At HLA-A, low levels of variation are observed at B and F pockets and randomized He and G do not differ from the observed data. By contrast, HLA-B concentrates most of the differences between supertypes, the B pocket showing a particularly high level of variation. Moreover, at HLA-B, the reassignment of alleles into random groups does not reproduce the patterns of population differentiation observed with supertypes. We thus conclude that differently from HLA-A, for which supertype and allelic variation show similar patterns of nucleotide diversity within and between populations, HLA-B has likely evolved through specific adaptations of its B pocket to local pathogens. [ABSTRACT FROM AUTHOR]

The above article originally published with an incomplete bibliographic information for Bitarello et al. (2016) and presented correctly in this article. [ABSTRACT FROM AUTHOR]

HLA-C locus mismatches (MMs) are the most frequent class I disparities in unrelated hematopoietic stem cell transplantation (HSCT) and have a detrimental impact on clinical outcome. Recently, a few retrospective clinical studies have reported some variability in the immunogenicity of HLA-C incompatibilities. To get better insight into presumably permissive HLA-C MMs, we have developed a one-way in vitro mixed lymphocyte reaction (MLR) assay allowing to quantify activated CD56 - CD137 + CD8 + lymphocytes in HLA-C incompatible combinations. T cell-mediated alloresponses were correlated with genetic markers such as HLA-C mRNA expression and the number of amino acid (aa) MMs in the α1/α2 domains (peptide-binding region). Because of the high rate of HLA-DPB1 incompatibilities in HLA-A-, B-, C-, DRB1-, and DQB1-matched unrelated HSCT patient/donor pairs, the impact of HLA-DPB1 mismatching, a potential bystander of CD4 + T cell activation, was also considered. Heterogeneous alloresponses were measured in 63 HLA-C-mismatched pairs with a positive assay in 52% of the combinations (2.3-18.6% activated CTLs), representing 24 different HLA-A~B~DRB1~DQB1 haplotypes. There was no correlation between measured alloresponses and mRNA expression of the mismatched HLA-C alleles. The HLA-C*03:03/03:04 MM did not induce any positive alloresponse in five MLRs. We also identified HLA-C*02:02 and HLA-C*06:02 as mismatched alleles with lower immunogenicity, and HLA-C*14:02 as a more immunogenic MM. A difference of at least 10 aa residues known to impact peptide/T cell receptor (TCR) binding and a bystander HLA-DPB1 incompatibility had a significant impact on CTL alloreactivity ( p  = 0.021). The same HLA-C MM, when recognized by two different responders with the same HLA haplotypes, was recognized differently, emphasizing the role of the T-cell repertoire of responding cells. In conclusion, mismatched HLA-C alleles differing by 10 or more aas in the peptide/TCR-binding region, when occurring together with HLA-DPB1 incompatibilities, should be considered as high-risk MMs in unrelated HSCT.