Very little is currently known about the major histocompatibility complex (MHC) region of cynomolgus macaques (Macaca fascicularis; Mafa) from Chinese breeding centers. We performed comprehensive MHC class I haplotype analysis of 100 cynomolgus macaques from two different centers, with animals from different reported original geographic origins (Vietnamese, Cambodian, and Cambodian/Indonesian mixed-origin). Many of the samples were of known relation to each other (sire, dam, and progeny sets), making it possible to characterize lineage-level haplotypes in these animals. We identified 52 Mafa-A and 74 Mafa-B haplotypes in this cohort, many of which were restricted to specific sample origins. We also characterized full-length MHC class I transcripts using Pacific Biosciences (PacBio) RS II single-molecule real-time (SMRT) sequencing. This technology allows for complete read-through of unfragmented MHC class I transcripts (~1100 bp in length), so no assembly is required to unambiguously resolve novel full-length sequences. Overall, we identified 311 total full-length transcripts in a subset of 72 cynomolgus macaques from these Chinese breeding facilities; 130 of these sequences were novel and an additional 115 extended existing short database sequences to span the complete open reading frame. This significantly expands the number of Mafa-A, Mafa-B, and Mafa-I full-length alleles in the official cynomolgus macaque MHC class I database. The PacBio technique described here represents a general method for full-length allele discovery and genotyping that can be extended to other complex immune loci such as MHC class II, killer immunoglobulin-like receptors, and Fc gamma receptors.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Price includes VAT (USA)
Tax calculation will be finalised during checkout.
Aarnink A, Apoil PA, Takahashi I, Osada N, Blancher A (2011) Characterization of MHC class I transcripts of a Malaysian cynomolgus macaque by high-throughput pyrosequencing and EST libraries. Immunogenetics 63(11):703–713
Bontrop RE (2006) Comparative genetics of MHC polymorphisms in different primate species: duplications and deletions. Hum Immunol 67(6):388–397
Budde ML, Wiseman RW, Karl JA, Hanczaruk B, Simen BB, O’Connor DH (2010) Characterization of Mauritian cynomolgus macaque major histocompatibility complex class I haplotypes by high-resolution pyrosequencing. Immunogenetics 62(11–12):773–780
Campbell KJ, Detmer AM, Karl JA, Wiseman RW, Blasky AJ, Hughes AL, Bimber BN, O’Connor SL, O’Connor DH (2008) Characterization of 47 MHC class I sequences in Filipino cynomolgus macaques. Immunogenetics 61(3):177–187
Daza-Vamenta R, Glusman G, Rowen L, Guthrie B, Geraghty DE (2004) Genetic divergence of the rhesus macaque major histocompatibility complex. Genome Res 14(8):1501–1515
de Groot N, Doxiadis GG, Otting N, de Vos-Rouweler AJ, Bontrop RE (2014) Differential recombination dynamics within the MHC of macaque species. Immunogenetics 66(9–10):535–544
de Groot NG, Otting N, Robinson J, Blancher A, Lafont BA, Marsh SG, O’Connor DH, Shiina T, Walter L, Watkins DI, Bontrop RE (2012) Nomenclature report on the major histocompatibility complex genes and alleles of great ape, old and new world monkey species. Immunogenetics 64(8):615–631
Heimbruch KE, Karl JA, Wiseman RW, Dudley DM, Johnson Z, Kaur A, O’Connor DH (2015) Novel MHC class I full-length allele and haplotype characterization in sooty mangabeys. Immunogenetics 67(8):437–445
Karl JA, Bohn PS, Wiseman RW, Nimityongskul FA, Lank SM, Starrett GJ, O’Connor DH (2013) Major histocompatibility complex class I haplotype diversity in Chinese rhesus macaques. G3 (Bethesda) 3(7):1195–1201
Karl JA, Heimbruch KE, Vriezen CE, Mironczuk CJ, Dudley DM, Wiseman RW, O’Connor DH (2014) Survey of major histocompatibility complex class II diversity in pig-tailed macaques. Immunogenetics 66(11):613–623
Kita YF, Hosomichi K, Kohara S, Itoh Y, Ogasawara K, Tsuchiya H, Torii R, Inoko H, Blancher A, Kulski JK, Shiina T (2009) MHC class I a loci polymorphism and diversity in three southeast Asian populations of cynomolgus macaque. Immunogenetics 61(9):635–648
Krebs KC, Jin Z, Rudersdorf R, Hughes AL, O’Connor DH (2005) Unusually high frequency MHC class I alleles in Mauritian origin cynomolgus macaques. J Immunol 175(8):5230–5239
Lawrence J, Orysiuk D, Prashar T, Pilon R, Fournier J, Rud E, Sandstrom P, Plummer FA, Luo M (2012) Identification of 23 novel MHC class I alleles in cynomolgus macaques of Philippine and Philippine/Mauritius origins. Tissue Antigens 79(4):306–307
Ling F, Zhuo M, Ni C, Zhang GQ, Wang T, Li W, Wei LQ, Du HL, Wang JF, Wang XN (2012) Comprehensive identification of high-frequency and co-occurring mafa-B, mafa-DQB1, and mafa-DRB alleles in cynomolgus macaques of Vietnamese origin. Hum Immunol 73(5):547–553
Otting N, de Groot N, de Vos-Rouweler AJ, Louwerse A, Doxiadis GG, Bontrop RE (2012) Multilocus definition of MHC haplotypes in pedigreed cynomolgus macaques (Macaca fascicularis). Immunogenetics 64(10):755–765
Otting N, de Vos-Rouweler AJ, Heijmans CM, de Groot NG, Doxiadis GG, Bontrop RE (2007) MHC class I a region diversity and polymorphism in macaque species. Immunogenetics 59(5):367–375
Otting N, Doxiadis GG, Bontrop RE (2009) Definition of mafa-a and -B haplotypes in pedigreed cynomolgus macaques (Macaca fascicularis). Immunogenetics 61(11–12):745–753
Otting N, Heijmans CM, Noort RC, de Groot NG, Doxiadis GG, van Rood JJ, Watkins DI, Bontrop RE (2005) Unparalleled complexity of the MHC class I region in rhesus macaques. Proc Natl Acad Sci U S A 102(5):1626–1631
Pendley CJ, Becker EA, Karl JA, Blasky AJ, Wiseman RW, Hughes AL, O’Connor SL, O’Connor DH (2008) MHC class I characterization of Indonesian cynomolgus macaques. Immunogenetics 60(7):339–351
Robinson J, Halliwell JA, McWilliam H, Lopez R, Parham P, Marsh SG (2013) The IMGT/HLA database. Nucleic Acids Res 41(Database issue):D1222–D1227
Saito Y, Naruse TK, Akari H, Matano T, Kimura A (2012) Diversity of MHC class I haplotypes in cynomolgus macaques. Immunogenetics 64(2):131–141
Shiina T, Yamada Y, Aarnink A, Suzuki S, Masuya A, Ito S, Ido D, Yamanaka H, Iwatani C, Tsuchiya H, Ishigaki H, Itoh Y, Ogasawara K, Kulski JK, Blancher A (2015) Discovery of novel MHC-class I alleles and haplotypes in Filipino cynomolgus macaques (Macaca fascicularis) by pyrosequencing and sanger sequencing : mafa-class I polymorphism. Immunogenetics 67(10):563–578
Uda A, Tanabayashi K, Fujita O, Hotta A, Terao K, Yamada A (2005) Identification of the MHC class I B locus in cynomolgus monkeys. Immunogenetics 57(3–4):189–197
Uda A, Tanabayashi K, Yamada YK, Akari H, Lee YJ, Mukai R, Terao K, Yamada A (2004) Detection of 14 alleles derived from the MHC class I a locus in cynomolgus monkeys. Immunogenetics 56(3):155–163
Wang HB, Ling F, Zhuo M, Wang JF, Wang XN (2011) Twenty-three novel major histocompatibility complex class I B alleles identified in cynomolgus macaques of Vietnamese origin. Tissue Antigens 77(4):346–348
Westbrook CJ, Karl JA, Wiseman RW, Mate S, Koroleva G, Garcia K, Sanchez-Lockhart M, O’Connor DH, Palacios G (2015) No assembly required: full-length MHC class I allele discovery by PacBio circular consensus sequencing. Hum Immunol 76(12):891–896
Wiseman RW, Karl JA, Bimber BN, O’Leary CE, Lank SM, Tuscher JJ, Detmer AM, Bouffard P, Levenkova N, Turcotte CL, Szekeres EJ, Wright C, Harkins T, O’Connor DH (2009) Major histocompatibility complex genotyping with massively parallel pyrosequencing. Nat Med 15(11):1322–1326
Wiseman RW, Karl JA, Bohn PS, Nimityongskul FA, Starrett GJ, O’Connor DH (2013) Haplessly hoping: macaque major histocompatibility complex made easy. ILAR J 54(2):196–210
Wiseman RW, Wojcechowskyj JA, Greene JM, Blasky AJ, Gopon T, Soma T, Friedrich TC, O’Connor SL, O’Connor DH (2007) Simian immunodeficiency virus SIVmac239 infection of major histocompatibility complex-identical cynomolgus macaques from Mauritius. J Virol 81(1):349–361
Wu J, Bassinger S, Montoya GD, Chavez L, Jones CE, Holder-Lockyer B, Masten B, Williams TM, Prilliman KR (2008) Allelic diversity within the high frequency Mamu-A2*05/mane-A2*05 (mane-a*06)/mafa-A2*05 family of macaque MHC-a loci. Tissue Antigens 72(1):29–38
Zhang GQ, Ni CH, Ling F, Qiu W, Wang HB, Xiao Y, Guo XJ, Huang JY, Du HL, Wang JF, Zhao SJ, Zhuo M, Wang XN (2012) Characterization of the major histocompatibility complex class I a alleles in cynomolgus macaques of Vietnamese origin. Tissue Antigens 80(6):494–501
Zhuo M, Wang HB, Ling F, Wang JF, Wang XN (2011) Eighteen novel MHC class I a alleles identified in Vietnamese-origin cynomolgus macaques. Tissue Antigens 78(2):139–142
The authors thank Catherine Westbrook, Suzanne Mate, and Galina Koroleva at the US Army Medical Research Institute of Infectious Diseases, and Katherine Munson and the staff of the University of Washington PacBio Sequencing Services core for performing PacBio RS II sequencing. We also thank Annemiek J.M. de Vos-Rouweler for RNA isolations and preparation of cDNAs used in this study. Finally, we thank Nel Otting, Natasja de Groot and the IMGT Non-human Primate Nomenclature Committee for obtaining accession numbers and providing official Mafa transcript nomenclature designations for the sequences described here. This research was supported by contracts HHSN272201600007C and HHSN272201100013C from the National Institute of Allergy and Infectious Diseases of the National Institutes of Health, and was conducted at a facility constructed with support from the Research Facilities Improvement Program (RR15459-01, RR020141-01).
Electronic supplementary material
(PDF 103 kb).
(PDF 397 kb).
(PDF 383 kb).
(PDF 1888 kb).
(PDF 888 kb).
(XLSX 217 kb).
(XLSX 159 kb).
(XLSX 70 kb).
(XLSX 83 kb).
About this article
Cite this article
Karl, J.A., Graham, M.E., Wiseman, R.W. et al. Major histocompatibility complex haplotyping and long-amplicon allele discovery in cynomolgus macaques from Chinese breeding facilities. Immunogenetics 69, 211–229 (2017). https://doi.org/10.1007/s00251-017-0969-7
- Macaca fascicularis
- MHC class I
- PacBio long-amplicon sequencing
- RNA transcript-based haplotypes