David S. Dunn

IL28B, HLA-C, and KIR variants additively predict response to therapy in chronic hepatitis C virus infection in a European Cohort: a cross-sectional study.

Vijayaprakash Suppiah and colleagues show that genotyping hepatitis C patients for the IL28B, HLA-C, and KIR genes improves the ability to predict whether or not patients will respond to antiviral treatment.

Genome sequence of the pathogenic intestinal spirochete brachyspira hyodysenteriae reveals adaptations to its lifestyle in the porcine large intestine.

Brachyspira hyodysenteriae is an anaerobic intestinal spirochete that colonizes the large intestine of pigs and causes swine dysentery, a disease of significant economic importance. The genome sequence of B. hyodysenteriae strain WA1 was determined, making it the first representative of the genus Brachyspira to be sequenced, and the seventeenth spirochete genome to be reported. The genome consisted of a circular 3,000,694 base pair (bp) chromosome, and a 35,940 bp circular plasmid that has not previously been described. The spirochete had 2,122 protein-coding sequences. Of the predicted proteins, more had similarities to proteins of the enteric Escherichia coli and Clostridium species than they did to proteins of other spirochetes. Many of these genes were associated with transport and metabolism, and they may have been gradually acquired through horizontal gene transfer in the environment of the large intestine. A reconstruction of central metabolic pathways identified a complete set of coding sequences for glycolysis, gluconeogenesis, a non-oxidative pentose phosphate pathway, nucleotide metabolism, lipooligosaccharide biosynthesis, and a respiratory electron transport chain. A notable finding was the presence on the plasmid of the genes involved in rhamnose biosynthesis. Potential virulence genes included those for 15 proteases and six hemolysins. Other adaptations to an enteric lifestyle included the presence of large numbers of genes associated with chemotaxis and motility. B. hyodysenteriae has diverged from other spirochetes in the process of accommodating to its habitat in the porcine large intestine.

Genomic and phylogenetic analysis of the S100A7 (Psoriasin) gene duplications within the region of the S100 gene cluster on human chromosome 1q21

The human S100 gene family encodes the EF-hand superfamily of calcium-binding proteins, with at least 14 family members clustered relatively closely together on chromosome 1q21. We have analyzed the most recently available genomic sequence of the human S100 gene cluster for evidence of tandem gene duplications during primate evolutionary history. The sequences obtained from both GenBank and GoldenPath were analyzed in detail using various comparative sequence analysis tools. We found that of the S100A genes clustered relatively closely together within a genomic region of 260 kb, only the S100A7 (psoriasin) gene region showed evidence of recent duplications. The S100A7 gene duplicated region is composed of three distinct genomic regions, 33, 11, and 31 kb, respectively, that together harbor at least five identifiable S100A7-like genes. Regions 1 and 3 are in opposite orientation to each other, but each region carries two S100A7-like genes separated by an 11-kb intergenic region (region 2) that has only one S100A7-like gene, providing limited sequence resemblance to regions 1 and 3. The duplicated genomic regions 1 and 3 share a number of different retroelements including five Alu subfamily members that serve as molecular clocks. The shared (paralogous) Alu S insertions suggest that regions 1 and 3 were probably duplicated during or after the phase of AluS amplification some 30–40 mya. We used PCR to amplify an indel within intron 1 of the S100A7a and S100A7c genes that gave the same two expected product sizes using 40 human DNA samples and 1 chimpanzee sample, therefore confirming the presence of the region 1 and 3 duplication in these species. Comparative genomic analysis of the other S100 gene members shows no similarity between intergenic regions, suggesting that they diverged long before the emergence of the primates. This view was supported by the phylogenetic analysis of different human S100 proteins including the human S100A7 protein members. The S100A7 protein, also known as psoriasin, has important functions as a mediator and regulator in skin differentiation and disease (psoriasis), in breast cancer, and as a chemotactic factor for inflammatory cells. This is the first report of five copies of the S100A7 gene in the human genome, which may impact on our understanding of the possible dose effects of these genes in inflammation and normal skin development and pathogenesis.

Associations between HLA-DRB1*0102, HLA-B*5801, and hepatotoxicity during initiation of nevirapine-containing regimens in South Africa.

A three-drug regimen comprising nevirapine plus two nucleoside reverse transcriptase inhibitors is frequently prescribed for HIV-1 infection worldwide. Unfortunately, drug hypersensitivity which can include severe hepatotoxicity during the initial weeks of therapy with nevirapine, particularly when initiated in women with >250 CD4+ T cells/mm3, have prompted changes in prescribing guidelines. Although isolated non-severe skin rash associated with nevirapine may be managed conservatively, the major treatment-limiting side effect of nevirapine is a drug hypersensitivity syndrome comprising fever, skin rash and/or hepatitis. This affects as many as 5% of those starting the drug. Various HLA class I and II alleles have been associated with nevirapine rash and/or hepatitis across different populations [1–6]. A population based study from Western Australia first identified an association between HLA-DRB1*0101 and nevirapine-associated hepatitis among patients with ≥25% CD4+ T cells [1]. This work was complemented by ex vivo studies suggesting a CD4 T cell-dependent process. Associations were subsequently identified between various HLA class I alleles such as HLA-B*1402 and HLA-Cw8 and nevirapine hypersensitivity in Sardinian [2] and Japanese [3] populations. A case control study in a Thai population identified an association between HLA-B*3505 and nevirapine-associated rash or hypersensitivity [4]. Further analyses from Western Australia confirmed the original association between HLA-DRB1*0101 and ≥25% CD4 T cells, and identified an association between HLA-B*3501 and nevirapine-associated rash, suggesting that phenotype was particularly important for defining specific HLA associations [5]. Only in the presence of hepatitis was HLA-DRB1*0101 associated with rash, while HLA-B*3501 was associated with rash regardless of hepatitis [5]. A retrospective analysis involving 76 sites in 11 countries (none in Africa) enrolled 276 cases who had experienced severe rash and/or hepatitis within 8 weeks of nevirapine initiation, and 587 nevirapine-tolerant controls [6]. Based on two digit HLA typing, that study associated HLA-DRB1*01 with hepatitis in whites, both HLA-B*35 and HLA-Cw*04 with rash in Asians, and HLA-Cw*04 alone with rash in Whites and Blacks. Furthermore, CYP2B6 slow metabolizer genotype was associated with rash but not hepatotoxicity. The above studies suggest that diverse genetic, immunological and drug metabolism pathways contribute to nevirapine hypersensitivity syndromes, and that associations may differ by specific toxicity phenotype and population. Data from Africa, where HIV prevalence and nevirapine use is high have been scant. Herein we used data and specimens previously collected during a clinical trial in South Africa to characterize relationships between HLA I and II alleles and nevirapine hepatotoxicity.

Identification of genes associated with prophage-like gene transfer agents in the pathogenic intestinal spirochaetes Brachyspira hyodysenteriae, Brachyspira pilosicoli and Brachyspira intermedia

VSH-1 is an unusual prophage-like gene transfer agent (GTA) that has been described in the intestinal spirochaete Brachyspira hyodysenteriae. The GTA does not self-propagate, but it assembles into a virus-like particle and transfers random 7.5kb fragments of host DNA to other B. hyodysenteriae cells. To date the GTA VSH-1 has only been analysed in B. hyodysenteriae strain B204, in which 11 late function genes encoding prophage capsid, tail and lysis elements have been described. The aim of the current study was to look for these 11 genes in the near-complete genomes of B. hyodysenteriae WA1, B. pilosicoli 95/1000 and B. intermedia HB60. All 11 genes were found in the three new strains. The GTA genes in WA1 and 95/1000 were contiguous, whilst some of those in HB60 were not-although in all three strains some gene rearrangements were present. A new predicted open reading frame with potential functional importance was found in a consistent position associated with all four GTAs, located between the genes for head protein Hvp24 and tail protein Hvp53, overlapping with the hvp24 sequence. Differences in the nucleotide and predicted amino acid sequences of the GTA genes in the spirochaete strains were consistent with the overall genetic distances between the strains. Hence the GTAs in the two B. hyodysenteriae strains were considered to be strain specific variants, and were designated GTA/Bh-B204 and GTA/Bh-WA1 respectively. The GTAs in the strains of B. intermedia and B. pilosicoli were designated GTA/Bint-HB60 and GTA/Bp-95/1000 respectively. Further work is required to determine the extent to which these GTAs can transfer host genes between different Brachyspira species and strains.

HLA class I restricted CD8+ and class II restricted CD4+ t cells are implicated in the pathogenesis of nevirapine hypersensitivity

Objectives:This study sought to examine nevirapine hypersensitivity (NVP HSR) phenotypes and their relationship with differing major histocompatibility complex (MHC) Class I and Class II alleles and the associated CD4+ and CD8+ T-cell NVP-specific responses and their durability over time. Methods:A retrospective cohort study compared HIV-positive patients with NVP HSR, defined by fever and hepatitis and/or rash, with those tolerant of NVP for more than 3 months. Covariates included class I (HLA-A, B, C) and class II (HLA-DR) alleles. Cellular studies examined NVP-specific CD4+ and CD8+ T-cell responses by interferon-gamma (IFN&ggr;) ELISpot assay and intracellular cytokine staining (ICS). Results:NVP HSR occurred in 19 out of 451 (4%) NVP-exposed individuals between March 1993 and December 2011. HLA associations were phenotype dependent with HLA-DRB1*01 : 01 associated with hepatitis (P = 0.02); HLA-B*35 : 01 and HLA-Cw4 associated with cutaneous NVP HSR (P = 0.001, P = 0.01), and HLA-Cw*08 was associated with NVP HSR with eosinophilia (P = 0.04) and multisystemic NVP HSR (P = 0.02). NVP-specific INF&ggr; responses waned significantly more than 3 months from the original reaction and were diminished or completely abrogated when either CD4+ or CD8+ T cells were depleted from the peripheral blood mononuclear cells culture. Conclusion:The association of specific class I and II allele pairings with specific phenotypes of NVP HSR, and cellular studies showing both CD4+ and CD8+ T-cell NVP-specific responses suggest that specific combinations of NVP reactive class I restricted CD8+ and class II restricted CD4+ T cells contribute to the immunopathogenesis of NVP HSR.

Polymorphic Alu insertions within the Major Histocompatibility Complex class I genomic region: a brief review.

Most polymorphic Alu insertions (POALINs) belong to a subgroup of the Alu multicopy retrotransposon family of short interspersed nucleotide elements (SINEs) that are categorized as AluYb8 and AluYa5. The number of AluYb8/AluYa5 members (∼4,492 copies) is significantly less than the ∼one million fixed Alu copies per human genome. We have studied the presence of POALINs within the Major Histocompatibility Complex (MHC) class I region on the short arm of chromosome 6 (6p21.3) because this region has a high gene density, many genes with immune system functions, large sequence variations and diversity, duplications and redundancy, and a strong association with more than 100 different diseases. Since little is known about POALINs within the MHC genomic region, we undertook to identify some of the members of the AluYb8/AluYa5 subfamily and to study their frequency of distribution and genetic characteristics in different populations. As a result of our comparative genomic analyses, we identified the insertion sites for five POALINs distributed within the MHC class I region. This brief review outlines the locations of the insertions and sequence features of the five MHC POALINs, their single site and haplotype frequencies in different geographic populations, and their association with different HLA class I genes and disease. We show that the MHC POALINs have a potential value as lineage and linkage markers for the study of human population genetics, disease associations, genomic diversity and evolution.

The Association Between HLA-A Alleles and Young Alu Dimorphisms Near the HLA-J, -H, and -F Genes in Workshop Cell Lines and Japanese and Australian Populations

At least two polymorphic Alu insertions have been previously identified and characterized within the class I region of the major histocompatibility complex (MHC). We have identified another two new polymorphic Alu insertions, AluyHJ and AluyHF, located near HLA-J and HLA-F, respectively, within the a block of the MHC. Here we report on (1) the haplotypic relationships between the Alu dimorphisms and the HLA-A locus within a panel of 51 IHW homozygous cell lines representing at least 36 HLA class I haplotypes, (2) the Alu genotype, allele, and haplotype frequencies present in the Australian Caucasians and Japanese populations, and (3) the frequency of association between the different Alu dimorphisms and the HLA-A alleles in 109 Australian Caucasians and 99 Japanese. PCR was used to detect the presence or absence of insertion for AluyHJ, AluyHG, and AluyHF within the DNA samples prepared from the cell lines and the two population groups that had been previously typed for HLA-A. In the homozygous cell lines, all three Alu insertions were found in only one HLA class I haplotype (HLA-A1, -B57, -Cw6), no Alu insertions were detected in six HLA class I haplotypes and one or more of the Alu insertions were found in 29 HLA class I haplotypes. At least one of the Alu insertions was found in about 86% of the Japanese and Australian individuals, with the AluyHJ generally related inversely to AluyHG and/or AluyHF. The gene frequency of the AluyHJ and AluyHF insertions was significantly different (p <0.05) BETWEEN JAPANESE AND AUSTRALIANS, WHEREAS THERE WAS NO DIFFERENCE (P > 0.05) between the frequencies of AluyHG in the two populations. The Alu haplotype frequencies were also significantly different between the Japanese and the Australians. In the cell lines and the population groups, the AluyHJ insertion was most frequently found associated with HLA-A1 or A24, AluyHG with HLA-A2, and AluyHF with HLA-A2, -A10, or -A26. This study suggests that the three polymorphic Alu elements have been inserted into the a block of the MHC in different progenitor groups and therefore will be useful lineage and linkage markers in human population studies and for elucidating the evolution of HLA class I haplotypes.

Alu polymorphism within the MICB gene and association with HLA-B alleles

We describe the finding of an Alu repeat dimorphism within the first intron of the MICB gene. The frequencies of the two AluyMICB alleles, AluyMICB*0 (absence of insertion) and AluyMICB*1 (presence of insertion), and their associations with the highly polymorphic HLA-B locus were determined for 51 human cell lines and for 109 and 200 Caucasians and northeastern Thais, respectively. Analysis of the AluyMICB and HLA-B allelic relationships revealed that AluyMICB*1 occurred at relatively low gene frequency (0.113–0.118) but was strongly associated with HLA-B17 (HLA-B57,HLA-B58) and HLA-B13. The AluyMICB locus provides a useful dimorphic marker for investigations on the level of linkage disequilibrium between MICB, MICA, and HLA-B loci.

Polymorphic Alu insertions and their associations with MHC class I alleles and haplotypes in the northeastern Thais.

Polymorphic Alu insertions (POALINs) are known to contribute to the strong polymorphic nature of the Major Histocompatibility Complex (MHC). Previous population studies on MHC POALINs were limited to only Australian Caucasians and Japanese. Here, we report on the individual insertion frequency of the five POALINs within the MHC class I region, their HLA‐A and ‐B associations, and the three and four locus alpha block POALIN haplotype frequencies in the Northeastern (NE) Thai population. Of the five POALINs, the lowest frequency was 0.018 for AluyHF and the highest frequency was 0.292 for AluyHJ and AluyHG. The strongest positive associations between the POALINs and HLA class I alleles was between AluyMICB and HLA‐B*57, AluyHJ and HLA‐A*24 and HLA‐A*01, and AluyHG and HLA‐A*02, supporting previous findings in Caucasians and Japanese. Single POALIN haplotypes were found more frequently than multiple POALIN haplotypes. However, of the seven different POALIN haplotypes within the MHC alpha block, there were only two significant differences between the NE Thais, Caucasians and Japanese. This study confirms that the MHC POALINs are in linkage disequilibrium with HLA‐A and –B alleles and that there are significant frequency differences for some of the POALINs when compared between NE Thai, Caucasians and Japanese.