Exploring the ancestry and admixture of Mexican Oaxaca Mestizos from Southeast Mexico using next-generation sequencing of 11 HLA loci.

The Mestizos of Oaxaca resulted from the admixture of Zapotecan Natives with Spaniards and Africans. We selected 112 donors from Oaxaca and applied next-generation sequencing to characterize exon and intron variants in complete or extended HLA genes. Some alleles found, are unique to Mexican Natives and most likely will be absent in most major ethnicities, namely: Caucasians, Africans or Asians. Among these are HLA-A*68:03:01, HLA-A*68:05:01, HLA-C*03:04:01:02, HLA-C*15:09, HLA-C*3:05, HLA-C*03:06:01, HLA-B*39:05:01, HLA-B*35:14:01, HLA-B*35:12:01, HLA-B*35:43:01, HLA-B*40:05, HLA-B:40:08, HLA-B*51:02:01, HLA-B*35:24:01 and HLA-B*39:08. HLA-DQA1*05:05:01:05 and some HLA-DRB1 alleles were only present in Amerindians/Mestizos. Three haplotypes are unique to Mexican Natives, five to Middle-Eastern and Sephardi-Jews. We detected a novel HLA-DQA1*04:01:01 exon 4 variant. Any novel allele may have been positively selected to enlarge the peptide-binding repertoire, and some, like HLA-B*39:02:02 and HLA-B*39:05:01 were found with unique haplotype associations, suggesting convergent evolution events and/or allele lineage diversification. The allele frequencies were fairly evenly distributed in most HLA loci with the exception of HLA-DPB1. The application of NGS in Oaxaca is novel and will lead to better use in the clinical setting. It offers deep knowledge on the population structure, origins, migration, and discovery of new alleles and haplotypes that other techniques did not achieve.

Strong linkage disequilibrium between HLA-B*3913 and DRB1*0807 in Brazilians.

The purpose of this research was to study the HLA-B39 distribution in 2560 healthy, unrelated, randomly selected individuals living in the southeastern region of Brazil (the states of Rio de Janeiro and São Paulo). Molecular methods were used to type HLA class I and II polymorphism: PCR-SSP, PCR-SSO, and PCR-SBT. HLA-B*39 was found in 7% (n = 182) of these individuals. HLA-B*3901, B*3906, and B*3913 were the most common alleles in this group (n = 57, 36, and 24, respectively). B*3913 was found associated with DRB1*0807 and DQB1*0402 in 16 of the 24 individuals and 13 of these were also associated with A*31012. This haplotype segregation was confirmed by family studies. Furthermore, in 5 of the 13 individuals carrying the A*31012, B*3913, DRB1*0807, and DQB1*0402 haplotype, HLA-DPB1*2701 was also present, suggesting that these alleles were found preferentially in cis association. DRB1-DPB1 linkage disequilibrium analysis was performed in 420 of the 2560 individuals and the association of DRB1*0807 with the uncommon DPB1*2701 was found to be highly significant (P <.0001). Because HLA-B*3913 and HLA-DRB1*0807 have been observed only in South American populations, it is possible that interlocus association has been selected to act on the same haplotype to collaborate in the class I and II restricted immune response to local pathogens and functional adaptation. Although numbers are small to predict which ethnic groups of the Brazilian population display this haplotype prevalently, it is possible to speculate that these data may have clinical application, such as in the selection of unrelated donors for bone marrow transplantation.

HLA diversity, differentiation, and haplotype evolution in Mesoamerican Natives.

Genetic variation of the Human Leukocyte Antigen region (HLA) in three Amerindian populations from the Southern Mexican state of Oaxaca, the Zapotec, Mixtec and the Mixe is examined. Individuals were typed using PCR-SSOP for four class II loci (DRB1, DQA1, DQB1, DPB1) and three class I loci (HLA-A, -B, and -C). Based on known HLA distributions, European admixture ranged from 1% to 10%. Individuals with European alleles were excluded from subsequent analysis. New alleles were revealed at each of the class I loci. In general, genotype frequencies were in Hardy-Weinberg equilibrium, although some deviations were detected. Allele frequency distributions at the DRB1, DQA1, DQB1 and HLA-A loci in all populations were more even than expected under neutrality, supporting a model of balancing selection at these loci. A history of directional selection for DPB1 in all three populations was indicated, as homozygosity values were significantly above expected values. Allele frequency distributions at HLA-B and HLA-C did not differ significantly from neutrality expectations. The data also provide evidence from linkage disequilibrium that strong haplotypic associations are present across the entire HLA region in each of the populations. Significant overall linkage disequilibrium exists between all pairs of loci typed in these populations, except those which include the DPB1 locus. These associations exist despite the fact that the recombination fraction between HLA-A, in the class I region, and DQB1, in the class II region, may exceed 0.02. One explanation is that selective pressures are maintaining the relationships between particular alleles at these loci in these populations. These relationships are maintained in general across the entire HLA region in the Oaxacan Amerindians, with the exception of DPB1.

Evolution of HLA-class I compared to HLA-class II polymorphism in Terena, a South-American Indian tribe.

We have studied the HLA alleles of 60 unrelated healthy Terena and 10 Terena families. They are members of an isolated Brazilian tribe located in Mato Grosso do Sul (South Central Brazil). Six novel alleles were found in this population: HLA-A*0219 (gf = 0.02), A*0222 (gf = 0.15), HLA-B* 3520 (gf = 0.01), B*3521 (gf = 0.03), B*3912 (gf = 0.03) and B*4803 (gf = 0.16). Five of the six novel alleles differ from their putative progenitors by amino acid replacements in residues that contribute to the pockets of the peptide-binding site. Many of the variants defined by molecular methods were not identified correctly by serological typing. We calculated heterozygosity values (H) for HLA-A, -B, -C, DRB1, DQB1 and DPB . The highest values were observed at the HLA-B locus, followed by HLA-A, -DRB1 and DQB1. Residue positions 9, 24, 45, 62, 67, 95, 114, 116, 156, and 163 of HLA class I showed heterozygosity values greater than 0.50. Nine of them contribute to the peptide-binding specificity pockets and one to the T cell receptor binding site. If HLA antigens are useful for defense against pathogenic agents, heterozygosity would offer an advantage by allowing binding of a larger repertoire of peptides to the class I molecules. Individuals that are heterozygous at these positions would probably have a wider repertoire of peptide presentation to T cells. The observed results including the presence of novel alleles in the class I HLA loci suggest a functionally significant, more rapid evolution of class I compared to class II loci in this South American isolated population.

Relevant residues of DRbeta1 third hypervariable region contributing to the expression and to severity of rheumatoid arthritis (RA) in Mexicans.

Rheumatoid arthritis (RA) is a chronic autoimmune disease leading to destruction of the joints. Residues at positions 67-74 of the DRB1 third hypervariable region are involved in susceptibility (S) and resistance (P) to RA. DNA from 83 patients and 175 controls, all of them Mexican Mestizos were oligotyped using PCR-SSOP and PCR-SSP. The (S) alleles are DRB*0404 (p = 0.000004), *0401 (p = 0.007) and *1001 (p = 0.008). Those associated with P are DRB1*0701 (p = 0.0001); *1101 (p = 0.01); *1503 (p = 0.02); *0801 (p = 0.04); *1401 (p = 0.04). Susceptibility/protection are recessive traits; SS genotypes are increased in the patients (p = 0.0003) while PP genotypes are decreased in them (p = 0.00004). The motif at 67-74 and the valine or glycine at position 86 are relevant in the development and severity of RA in Mexicans. The associations suggest that residues 67, 70, 71 are central for susceptibility. The P alleles have D-70 or carry V-86 in the absence of D-70. Thus, susceptibility/protection depends on the combination of basic residues at these positions and a non-polar aa at 86 contributes to resistance. Severity is also HLA influenced. DQA1*03011-DQB1*0302 are associated to severe lesions in the presence of any DR4 subtype. Analyzing different ethnic groups is essential to elucidate the etiopathogenesis of RA.

Dissimilar evolution of B-locus versus A-locus and class II loci of the HLA region in South American Indian tribes.

Native American populations have a limited HLA polymorphism compared with other ethnic groups. In spite of this, many novel HLA-B locus alleles, not observed in other populations, have been identified in South American tribes, and rapid evolution of this locus has been suggested. We have studied unrelated subjects of the Toba (TOB n = 116), Wichi (WIC n = 46) and Pilaga (PIL n = 14) tribes from northeastern Argentina to investigate the extent of the HLA polymorphism and obtain clues of selective forces that may have acted in these populations. In these tribes the number of HLA alleles is small at all loci except HLA-B, which presents 22 alleles. Seven novel alleles were characterized including 5 of HLA-B (B*35092, B*3518, B*3519, B*4009, B*4803) 1 at HLA-A (A*0219) and 1 at DRB1 (DRB1*0417). All these variants may have arisen by gene conversion events. Some of the novel variants represent the most frequent alleles of these populations (B*4803 in TOB and PIL; B*3519 in WIC) or are the most frequent subtypes in their lineages. HLA-A, B, DRB1,DQA1 and DQB1, but not DPB1, display relatively similar gene frequencies. This results in high heterozygosity in all the tribes for all the loci studied except HLA-DPB1. The larger polymorphism and the generation and maintenance of novel alleles at the HLA-B locus suggests a more specialized response of this locus to evolutionary forces. These effects may be related to the nature of the polymorphism, to the number of founder alleles and to the functional characteristics of the individual alleles.

An epitope in the third hypervariable region of the DRB1 gene is involved in the susceptibility to endemic pemphigus foliaceus (fogo selvagem) in three different Brazilian populations.

Endemic pemphigus foliaceus or fogo selvagem (FS) in an organ-specific autoimmune skin disease characterized by epidermal vesicles and mediated by autoantibodies. Family cases are frequent and not everyone living in endemic region develops FS suggesting that host factors play a role in determining whether exposed individuals will be affected. Because our previous works with Brazilian Mestizos and with Xavante Indians have shown that particular HLA alleles confer increased risk for the disease, we decided to extend these studies to another homogeneous population, the Terena Indians. 19 out of 20 Terena patients were either positive for DRB1*0404, 1402 or 1406 (p < 0.005, RR = 14). These findings were in agreement with the data obtained from the Xavante study. In Mestizos the association was with DRB1*01. All these alleles involved in predisposition to the disease in different populations shared the same amino acid sequence at position 67-74 on the third hypervariable region of the DRB1 gene: LLEQRRAA, suggesting that inheritance of this sequence is involved in the susceptibility to FS. When patients and controls data from different studies were pooled and analyzed disregarding the ethnic background and the HLA alleles involved, the results obtained clearly supported the hypothesis that matching for this epitope is highly significant and predictive of FS predisposition (p < 0.00001, RR = 6.4).

Allelic heterogeneity of HLA-B35 subtypes in different populations as assessed by DNA typing.

HLA-B35, a class I antigen differentially associated to several diseases in different ethnic groups, comprises at least eight alleles which differ among them by one to six amino acids. In the present work a rapid DNA typing procedure was used to investigate the distribution of the various HLA-B35 alleles in different populations. The approach is based on a group-specific PCR amplification of a set of closely related HLA-B alleles sharing a Thr in position 45 of the alpha-1 domain. The amplified DNA was then hybridized to a panel of sequence-specific oligonucleotide (SSO) probes designed to recognize the polymorphic residues in previously reported HLA-B35 subtypes. This methodology was successfully tested in 100 individuals of four different populations, previously typed by serology as HLA-B35, and in six reference panel cells of the 10th International Histocompatibility Workshop. HLA-B*3501 was the predominant subtype in all populations. B*3502, B*3503 and, to a lesser extent B*3508, were also found. Among Mexican Mestizos, thirteen individuals had patterns of SSO hybridization suggestive of new B35 alleles. The evolutionary considerations on the different B35 alleles and their extended B35,Cw4 haplotypes are discussed.

Endemic pemphigus foliaceus (fogo selvagem) in native Americans from Brazil.

BACKGROUND: Fogo selvagem (FS) is an autoimmune disease that is endemic in certain regions of Brazil and appears to be precipitated by an environmental factor. OBJECTIVE: Our purpose was to confirm the occurrence and prevalence of FS in a population of Xavante Indians living in an endemic region of central Brazil. METHODS: Clinical, anthropologic, and immunologic studies were carried out in patients and in normal inhabitants of the Pimentel Barbosa Indian Reservation, Mato Grosso, Brazil. RESULTS: FS was identified and confirmed in 10 patients from a patient pool of 295 with various skin diseases. The Xavante settlement has a total population of 746. Anti-desmoglein 1 autoantibodies were detected in all patients with FS and were absent from more than 300 serum samples collected from randomly selected unaffected persons. CONCLUSION: FS is strongly linked to outdoor activities and is largely restricted to immunogenetically predisposed persons. FS appears to have been endemic in certain regions of South America for several centuries.

Differences in HLA class II alleles of isolated South American Indian populations from Brazil and Argentina.

We have studied the HLA class II alleles in 277 South American Indians, which included Argentinian tribes from the Gran Chaco: Toba (n = 135), Toba-Pilaga (n = 19), Mataco-Wichi (n = 49), and Xavantes, a tribe from Central Brazil (n = 74). In the Brazilian tribe, only four DR groups were found: DRB1*1602 (gf = 0.303), DRB1*04 including DRB1*0404 (gf = 0.070) and DRB1*0407 (gf = 0.077), DRB1*0802 (gf = 0.265), and DRB1*1402 (gf = 0.303). The HLA class II allele frequencies were similar among the different Argentinian tribes, and 90% of DRB1 alleles belonged to three families: DRB1*04 (including DRB1*0403, DRB1*0404, DRB1*0407, DRB1*0411, and DRB1*0417), DRB1*0802, and DRB1*14 (including DRB1*1402 and DRB1*1406). At the DPB1 locus, we found only seven alleles, the most frequent being DPB1*0402. Comparison of HLA class II alleles with those of North American Indians that we have previously studied shows that the frequency of some HLA class II alleles in Brazilian Xavantes resembles that of North American Indians more than that of the Argentinian Indian tribes. The allele DRB1*0417 was found exclusively in this population.

HLA class II DNA typing in two Brazilian populations.

Brazil constitutes a melting pot of populations arising from three major groups, including Amerindians, Africans, and Europeans predominantly from Portugal who were later supplemented by migrations from other European countries. Although every possible combination of racial mixture exists in Brazil, we have selected for this study two groups of subjects residing in Rio de Janeiro. A predominant White population, among whom some Amerindian admixture may exist, and a predominantly African population having little admixture from the other races. We have used the polymerase chain reaction (PCR) and hybridization with oligonucleotide probes to perform a complete typing of the HLA class II alleles. We report the allele frequencies for HLA-DRB1, DQA1, DQB1 and DPB1. We also report on the postulated DR-DQ haplotypes based on family studies and observations in homozygous B-cell lines. These results may serve as background for various types of clinical studies in Brazilian populations.

HLA antigens and risk for development of pemphigus foliaceus (fogo selvagem) in endemic areas of Brazil.

Endemic pemphigus foliaceus (EPF), is an autoimmune disease associated with production of IgG antibodies against epidermal antigens. We have tested 38 patients and 50 control subjects living in endemic areas to investigate whether HLA genes are associated with host factors that determine whether or not exposed individuals will develop this disease. A variant of HLA-DR1, an antigen common in Blacks (DRB1*0102), was found to be the main susceptibility factor (relative risk = 7.3, P less than 0.0002). Two amino acids, in positions 85 and 86 of DRB1, distinguish DRB1*0102 from DRB1*0101. These residues appear to be involved in the formation of a functional epitope that causes T cell recognition and determines disease susceptibility. Moreover, subjects having DQw2 did not develop the disease, while the frequency of DQw2 in controls was 22% (RR = 0.04, P less than 0.006). Thus HLA genes appear to play a crucial role in the response to an environmental factor which in this setting frequently leads to the development of autoimmune disease. An HLA-DQ allele, DQw2, appears to be associated with factors that prevent the development of the disease in exposed individuals.