Reanalysis of sequence‐based HLA‐A, ‐B and ‐Cw typings: how ambiguous is today’s SBT typing tomorrow

Abstract

The permanently increasing number of human leukocyte antigen (HLA)-alleles and the growing list of ambiguities require continuous updating of high-resolution HLA typing results. Two different kinds of ambiguities exist: the first, when two or more allele combinations have identical heterozygous sequences, and the second, when differences are located outside the analyzed region. The number of HLA-A, B and C alleles recognized in 1999 was almost tripled in 2006. Two hundred individuals, sequence-based typing (SBT) typed in the period from 1999 to 2002, were reanalyzed using the 2006 database. A final allele typing result of at least four digits was obtained for HLA-A, -B and -C by heterozygous sequencing of exons 2 and 3 and, if necessary, additional exons and/or allele-specific sequencing. Storage of the individual sequences in a specially developed database enabled reanalysis with all present and future HLA releases. In the 5-year period HLA-A, -B and -C typing results became ambiguous in 37%, 46% and 41% of the cases. Most were because of differences outside the analyzed region; ambiguities because of different allele combinations with identical heterozygous sequences were present in 7%, 8% and 13% of the HLA-A, -B and -C typings. These results indicate that within 5 years, approximately half of the HLA SBT typings become ambiguous.

Estimation of high-resolution HLA-A, -B, -C, -DRB1 allele and haplotype frequencies based on 8862 German stem cell donors and implications for strategic donor registry planning

“…Donor registry HLA data, however, are generally heterogeneous with respect to typed HLA genes (only HLA-A and HLA-B or HLA-A, HLA-B, and HLA-DRB1 or more complete typing), typing resolution (high or low/intermediate resolution), and typing method (molecular methods or serology). Also, the discovery of new allelic variants compromises high-resolution typing data over time [11]. Furthermore, HLA data of registered stem cell donors often accumulate over several years.…”

Rapid, scalable and highly automated HLA genotyping using next-generation sequencing: a transition from research to diagnostics

“…Nevertheless, due to the ever increasing numbers of HLA alleles in the International ImMunoGeneTics (IMGT) HLA database [8] (release 3.9.0 includes 8,159 alleles) [9], histocompatibility testing is getting complex even if sequence-based typing (SBT) is used. Defining the phase of sequence motifs becomes more difficult and therefore genotype ambiguity increases exponentially through every new database release [10]. Therefore new typing strategies such as group-specific PCR [11] or allele separation by group-specific sequencing primers [12] prior to sequencing were developed.…”

Allele Frequency Estimation from Ambiguous Data: Using Resampling Schema in Validating Frequency Estimates and in Selective Neutrality Testing

“…The questions raised by the existence of ambiguous typings are well documented in the literature (Adams et al 2004;Leffell 2002;Lind et al 2010;Scott et al 1998;Swelsen et al 2004;Voorter et al 2007), but the approach of improving the resolution is simply not feasible in general for population-based studies. As the vast majority, if not almost all, of the computer programs are not able to handle frequency estimation for arbitrarily ambiguous data [e.g., ARLEQUIN (Excoffier et al 2005), PYPOP (Lancaster et al 2003), PHASE (Stephens and Donnelly 2003)]), we have developed an extension of the EM algorithm (Nunes 2005) capable of dealing with them.…”