, Volume 58, Issue 2–3, pp 203–215 | Cite as

Paucity of class I MHC gene heterogeneity between individuals in the endangered Hawaiian monk seal population

  • Brian M. Aldridge
  • Lizabeth Bowen
  • Brett R. Smith
  • George A. Antonelis
  • Frances Gulland
  • Jeffrey L. Stott
Original Paper


The Hawaiian monk seal population has experienced precipitous declines in the last 50 years. In this study, we provide evidence that individuals from remaining endangered population exhibit alarming uniformity in class I major histocompatibility (MHC) genes. The peripheral blood leukocyte-derived mRNA of six captive animals rescued from a stranding incident on the French frigate shoals in the Hawaiian archipelago was used to characterize genes in the monk seal class I MHC gene family, from which techniques for genotyping the broader population were designed using degenerate primers designed for the three major established human MHC class I loci (HLA-A, HLA-B, and HLA-C), and by sequencing multiple clones, six unique full-length classical MHC class I gene transcripts were identified among the six animals, three of which were only found in single individuals. Since The low degree of sequence variation between these transcripts and the similarity of genotype between individuals provided preliminary evidence for low class I MHC variability in the population. The sequence information from the class I transcripts from these six animals was used to design several primer sets for examining the extent of MHC variability in the remaining population using a combination of polymerase chain reaction and denaturing gradient gel electrophoresis (DGGE). Several DGGE assays, each one amplifying subtly different class I MHC gene combinations, were designed to compare exons encoding the highly polymorphic domains of the putative peptide-binding region of MHC class I. In combination, these assays failed to show interindividual variability at any of the class I MHC gene loci examined in either the six captive seals or in 80 free-ranging animals (∼6.7% of the estimated population) representing all six major subpopulations of Hawaiian monk seal.


Major histocompatibility complex Hawaiian monk seal Endangered species Immunogenetics Immunology 



Major histocompatibility complex


Denaturing gradient gel electrophoresis


Polymerase chain reaction


Human leukocyte antigen

Mosc-Mhc I

Monachus schauinslandi MHC class I



These studies were made possible by a generous grant from the Morris Animal Foundation. The authors thank Dr. Les Dalton, Dr. Jim McBain, and the staff of Sea World of San Antonio, TX, and Dr. Robert Braun (contract veterinarian) and staff of the Pacific Islands Fisheries Science Center, National Oceanic and Atmospheric Administration for the collection of samples from captive and wild seals, respectively. The collection of samples for this research was authorized by Marine Mammal Permit 848-1335 and by permit issued by Hawaiian Islands National Wildlife Refuge, U.S. Fish and Wildlife Service, U.S. Department of Interior. Experiments described in this paper conform to the current laws in the United States regarding the care and use of vertebrate animals for research.


  1. Aguilar A (1999) Status of Mediterranean monk seal (Monachus monachus) populations. In: RAC-SPA, United Nations Environment Program (UNEP). Aloes Editions, Tunis, pp 1–60Google Scholar
  2. Aguilar A, Roemer G, Debenham S, Binns M, Garcelon D, Wayne RK (2004) High MHC diversity maintained by balancing selection in an otherwise genetically monomorphic mammal. Proc Natl Acad Sci U S A 101:3490–3494CrossRefPubMedADSGoogle Scholar
  3. Aldridge BM, McGuirk SM, Clark RJ, Knapp LA, Watkins D, Lunn DP (1998) Denaturing gradient gel electrophoresis: a rapid method for differentiating BoLA-DRB3 alleles. Anim Genet 29:389–394CrossRefPubMedGoogle Scholar
  4. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410CrossRefPubMedGoogle Scholar
  5. Arnason U, Bodin K, Gullberg A, Ledje C, Mouchaty S (1995) A molecular view of pinniped relationships with particular emphasis on the true seals. J Mol Evol 40:78–85CrossRefPubMedGoogle Scholar
  6. Banish LD, Gilmartin WG (1992) Pathological findings in the Hawaiian monk seal. J Wildl Dis 28:428–434PubMedGoogle Scholar
  7. Bodmer JG, Marsh SGE, Albert ED, Bodmer WF, Bontrop RE, Dupont B, Erlich HA, Hansen JA, Mach B, Mayr WR, Parham P, Petersdorf EW, Sasazuki T, Schreuder GMT, Strominger JL, Svejgaard A, Terasaki PI (1999) Nomenclature for factors of the HLA system, 1998. Tissue Antigens 53:407–446CrossRefPubMedGoogle Scholar
  8. Bowen L, Aldridge BM, Stott JL, Gulland F, Woo J, Van Bonn W, DeLong R, Lowenstine L, Johnson ML (2002) Molecular characterization of expressed DQA and DQB genes in the California sea lion (Zalophus californianus). Immunogenetics 54:332–347CrossRefPubMedGoogle Scholar
  9. Bowen L, Aldridge BM, Gulland F, Van Bonn W, DeLong R, Melin S, Lowenstine LJ, Stott JL, Johnson ML (2004) Class II multiformity generated by variable MHC-DRB region configurations in the California sea lions (Zalophus californianus). Immunogenetics 56:12–27CrossRefPubMedGoogle Scholar
  10. Bowen L, Aldridge BM, DeLong R, Gulland F, Lowenstine L, Stott J, Johnson M (2005) An immunogenetic basis for the urogenital cancer epidemic of California sea lions (Zalophus californianus). Immunogenetics 56(11):846–848Google Scholar
  11. Carretta JV, Barlow J, Forney KA, Muto MM, Baker J (2001) U.S. Pacific Marine Mammal Stock Assessments: 2001. U.S. Department of Commerce, NOAA Tech. Memo. NMFS-TM-NMFS-SWFSC-317, p 284Google Scholar
  12. Costas E, Lopez-Rodas V (1998) Paralytic phycotoxins in monk seal mass mortality. Vet Rec 142:643–644PubMedCrossRefGoogle Scholar
  13. De Swart RL, Ross PS, Vos JG, Osterhaus ADME (1996) Impaired immunity in harbour seals (Phoca vitulina) exposed to bioaccumulated environmental contaminants: review of a long-term feeding study. Environ Health Perspect 104:823–828PubMedGoogle Scholar
  14. Dyall R, Messaoudi I, Janetzki S, Nikolic-Zugic J (2000) MHC polymorphism can enrich the T cell repertoire of the species by shifts in intrathymic selection. J Immunol 164(4):1695–1698PubMedGoogle Scholar
  15. Harvell CD, Kim K, Burkholder JM, Colwell RR, Epstein PR, Grimes DJ, Hofmann EE, Lipp EK, Osterhaus ADME, Overstreet RM, Porter JW, Smith GW, Vasta GR (1999) Emerging marine diseases-climate links and anthropogenic factors. Science 285:1505–1510CrossRefPubMedGoogle Scholar
  16. Harwood J (1998) What killed the monk seals? Nature 393:17–18CrossRefPubMedADSGoogle Scholar
  17. Hedrick PW, Kim T (2000) Genetics of complex polymorphisms: parasites and maintenance of MHC variation. In: Singh R, Krimbas C (eds) Evolutionary genetics: from molecules to morphology. Cambridge University Press, Cambridge, UK, pp 204–234Google Scholar
  18. Hughes AL, Nei M (1988) Pattern of nucleotide substitution at major histocompatibility complex class I loci reveals overdominant selection. Nature 335(6186):167–170CrossRefPubMedADSGoogle Scholar
  19. Hughes AL, Yeager M (1998) Natural selection at major histocompatibility complex loci of vertebrates. Annu Rev Genet 32:415–435CrossRefPubMedGoogle Scholar
  20. IUCN (1996) Red list of threatened animals. IUCN, Gland, SwitzerlandGoogle Scholar
  21. Jimenez JA, Hughes KA, Alaks G, Graham L, Lacy RC (1994) An experimental study of inbreeding depression in a natural habitat. Science 266(5183):271–273PubMedADSGoogle Scholar
  22. Knapp LA, Lehmann E, Hennes L, Eberle ME, Watkins DI (1997) High-resolution HLA-DRB typing using denaturing gradient gel electrophoresis and direct sequencing. Tissue Antigens 50:170–177PubMedCrossRefGoogle Scholar
  23. Kretzmann MB, Gilmartin WG, Meyer A, Zegers GP, Fain SR, Taylor BF, Costa DP (1997) Low genetic variability in the Hawaiian monk seal. Conserv Biol 11:482–490CrossRefGoogle Scholar
  24. Messaoudi I, Guevara Patino JA, Dyall R, LeMaoult J, Nikolich-Zugich J (2002) Direct link between MHC polymorphism, T cell avidity, and diversity in immune defense. Science 298:1797–1800CrossRefPubMedADSGoogle Scholar
  25. Nei M, Gojobori T (1986) Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions. Mol Biol Evol 3:418–426PubMedGoogle Scholar
  26. Osterhaus ADJ, Groen H, Niesters H, van de Bildt M, Martina B, Vedder L, Vos J, van Egmond H, Sidi BA, Barham MEO (1997) Morbilivirus in monk seal mass mortality. Nature 388:838–839CrossRefPubMedADSGoogle Scholar
  27. Parham P (2005) Putting a face to MHC restriction. J Immunol 174:3–5PubMedGoogle Scholar
  28. Pastor T, Garza JC, Allen P, Amos W, Aguilar A (2004) Low genetic variability in the highly endangered Mediterranean monk seal. J Hered 95:291–300CrossRefPubMedGoogle Scholar
  29. Ragen TJ, Lavigne DM (1997) The Hawaiian monk seal: biology of an endangered species. In: Twiss JR Jr, Reeves RR (eds) Conservation and management of marine mammals. Smithsonian Institution Press, Washington, D.C., pp 224–245Google Scholar
  30. Robinson J, Waller M, Parham P, Bodmer JG, Marsh SGE (2001) IMGT/HLA Database—a sequence database for the human major histocompatibility complex. Nucleic Acids Res 29:210–213CrossRefPubMedGoogle Scholar
  31. Sheffield VC, Cox DR, Lerman LS, Myers RM (1989) Attachment of a 40-base-pair G + C-rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain reaction results in improved detection of single-base changes. Proc Natl Acad Sci U S A 86:232–236PubMedADSGoogle Scholar
  32. Spielman D, Brook BW, Frankham R (2004) Most species are not driven to extinction before genetic factors impact them. Proc Natl Acad Sci U S A 101(42):15261–15264CrossRefPubMedADSGoogle Scholar
  33. Stanley HF, Harwood J (1997) Genetic differentiation among subpopulations of the highly endangered Mediterranean monk seal. In: Tew TE, Usher MB, Crawford TJ, Stevens D, Warren J, Spencer J (eds) The role of genetics in conserving small populations. Joint Nature Conservation Committee, Peterborough, pp 97–101Google Scholar
  34. Takei GH, Leong GH (1981) Macro-analytical methods used to analyze tissues of the Hawaiian monk seal, Monachus schauinslandi, for organochlorine pesticides, polychlorobiphenyls, and pentachlorophenol. Bull Environ Contam Toxicol 27:489–498CrossRefPubMedGoogle Scholar
  35. van de Bildt MW, Martina BE, Vedder EJ, Androukaki E, Kotomatas S, Komnenou A, Sidi BA, Jiddoi AB, Barham ME, Niesters HG, Osterhaus AD (2000) Identification of morbilliviruses of probable cetacean origin in carcases of Mediterranean monk seals (Monachus monachus). Vet Rec 146:691–694PubMedCrossRefGoogle Scholar
  36. Wirtz WO II (1968) Reproduction, growth and development, and juvenile mortality in the Hawaiian monk seal. J Mammal 49:229–238PubMedGoogle Scholar
  37. Yuhki N, O’Brien SJ (1988) Molecular characterization and genetic mapping of class I and class II MHC genes of the domestic cat. Immunogenetics 27:414–425CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Brian M. Aldridge
    • 1
    • 2
  • Lizabeth Bowen
    • 2
  • Brett R. Smith
    • 2
  • George A. Antonelis
    • 3
  • Frances Gulland
    • 4
  • Jeffrey L. Stott
    • 2
  1. 1.Veterinary Clinical SciencesRoyal Veterinary CollegeNorth MymmsUK
  2. 2.Laboratory for Marine Mammal Immunology, Department of Pathology, Microbiology and Immunology, School of Veterinary MedicineUniversity of CaliforniaDavisUSA
  3. 3.The Marine Mammal Center (TMMC)GGNRASausalitoUSA
  4. 4.National Marine Fisheries ServiceHonoluluUSA

Personalised recommendations