Molecular Studies of Antigens in HAP Organisms

  • R. Y. C. Lo


The use of recombinant DNA technology to study the HAP organisms has greatly improved our ability to characterize the many potential virulence factors of these bacteria. Instead of relying on conventional identification and purification methods based on biochemical, biophysical or immunological techniques, the genes coding for the virulence factors could be isolated for more detailed studies. After isolation and characterization of the genes, the recombinant antigens could be expressed for detailed biochemical and immunological studies. Further, large scale production of the recombinant antigens is also possible by high level expression in a heterologous system. These recombinant antigens could be used in vaccine trial and challenge experiments to investigate their protective capabilities. As a result of these molecular approaches, we now have a better understanding of some of the virulence factors of the HAP organisms. Some of these characterized antigens are being used as potential candidates for the development of an efficacious vaccine against these opportunistic pathogens in animals. In addition to studying the virulence antigens of the bacteria, the accumulation of molecular data is beginning to give us a picture into the biology, genetics and evolution of these bacteria.


Neisseria Meningitidis Recombinant Antigen galE Gene Actinobacillus Actinomycetemcomitans galT Gene 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Abdullah, K.M., 1991. Biochemical and genetic characterization of a Pasteurella haemolytica Al glycoprotease. Ph.D. Thesis, University of Guelph.Google Scholar
  2. Abdullah, K.M., Lo, R.Y.C., and Mellors, A., 1991, Cloning. nucleotide sequence, and expression of the Pasteurella haemolyticn AI glycoprotease gene. J. Bacteriol. 173: 5597–5603.PubMedGoogle Scholar
  3. Abdullah. K.M, Udoh, E.A., Shewen, P.E., and Mellors, A., 1992. A neutral glycoprotease of Pasteurella haemolyticn AI specifically cleaves 0-sialoglycoproteins. Infect. Immun. 60: 56–62.Google Scholar
  4. Adhya, S., 1987. The galactose operon. in Escherichia coli and Salmonella typhimurium: Cellular andGoogle Scholar
  5. Molecular Biology. F.C. Neidhart et al.,. eds., American Society for Microbiology, Wachington, D.C. Braun, V., and Focareta, T., 1991. Pore-forming bacterial protein hemolysins (cytolysins). Crit. Rev. Microhiol. 18: 115–158.Google Scholar
  6. Burrows, L.L., 1993, Molecular characterization of the RTX cytolysin determinants from Gram-negativeGoogle Scholar
  7. pathogens of veterinary significance. Ph.D. Thesis, University of Guelph, Guelph. Ontario, Canada. Burrows, L.L., and Lo, R.Y.C., 1992, Molecular characterization of an RTX toxin determinant from Actinobacillus suis. Infect. immun. 60: 2166–2173.Google Scholar
  8. Burrows, L.L., Olah-Winfield. E., and Lo, R.Y.C., 1993, Molecular analysis of the leukotoxin determinants from Pasteurella haemolytica serotypes 1 to 16. infect. immun. 61: 5001–5007.Google Scholar
  9. Carle, G.F., Frank, M. and Olson, M.V., 1986, Electrophoretic separation of large molecules by periodic inversion of the electric field. Science 232: 65–68.PubMedCrossRefGoogle Scholar
  10. Cavalieri. S.J., Bohach, G.A., and Snyder, i.S., 1984, Escherichia coli à-haemolysin: characteristics and probable role in pathogenicity. Microhiol. Rev. 48: 326–343.Google Scholar
  11. Chang, Y.-F., Young. R., Moulds. T.L., and Struck, D.K., 1989, Secretion of the Pasteurella leukotoxin by Escherichia coli. FEMS Microhiol. Lett. 60: 169–174.Google Scholar
  12. Chen, L., and Coleman. Jr., W.G., 1993, Cloning and characterization of the Escherichia coli K-12 (rfaC) gene, a gene required for lipopolysaccharide inner core synthesis. J. Bacteriol. 175: 2534–2540.PubMedGoogle Scholar
  13. Cho, H.J. and Jericho, K.W.F., 1986, Induction of immunity against pneumonic pasteurellosis following experimental infection in calves. Can. J. Vet. Res. 50: 27–31.Google Scholar
  14. Chu, G., Vollrath. D., and Davis. R.W., 1986. Separation of large DNA molecules by contour-clamped homogeneous electric fields. Science 234: 1582–1585.Google Scholar
  15. Confer, A.W., and Simons, K.R., 1986, Effects of Pasteurella haemo/vtica lipopolysaccharide on selected functions of bovine leukocytes. Am. J. Vet. Res. 47: 154–157.Google Scholar
  16. Conlon. J.A., Shewen, P.E., and Lo. R.Y.C., 1991. Efficacy of a recombinant leukotoxin in protection against pneumoniae challenge with Pasteurella haemolyticn Al. infect. Immun. 59: 587–591.Google Scholar
  17. Cooney. B.J., and Lo. R.Y.C., 1993, Three contiguous lipoprotein genes in Pasteurella haernolvtica which are homologous to a lipoprotein gene in Haemophilia influen:ae type B. infect. immun. 61: 4682–4688.Google Scholar
  18. Coote, J.G., 1992, Structural and functional relationships among the RTX toxin determinants of Gram-negative bacteria. FEMS Microbiol. Rev. 88: 137–162.Google Scholar
  19. Cope, L.D., Yogev, R., Mertsola. J., Latimer, J.L., Hanson, M.S., McCracken. Jr., G.H., and Hansen. E.J., 1991, Molecular cloning of a gene involved in lipopogosaccharide biosynthesis and virulence expression by Haentophiltts influenzae type B. Molec. Microhiol. 5: 1113–1124.Google Scholar
  20. Cornelissen, C.N., Biswas, G.D., Tsai, J., Paruchuri, D.K., Thompson, S.A., and Spading, P.F., 1992. Gonococcal transferrin-binding protein 1 is required for transferrin utilization and is homologous to TonB-dependent outer membrane receptors. J. Bacteriol. 174: 5788–5797.PubMedGoogle Scholar
  21. Frey, J., 1994, RTX-toxins in Actinobacillus pleuropn eu noniae and their potential role in virulence. in “Molecular Mechanisms of Bacterial Virulence”, C. I. Kado and J.H. Crosa, eds., Kluwer Academic Publishers.Google Scholar
  22. Frey, J., Beck. M., and Nicolet, J., 1994. RTX-toxins of Actinobacilhts pleta’opneunonine. In “Bacterial Protein Toxins. Freer et al. eds., Gustav Fisher.Google Scholar
  23. Gauthier, A., Turmel, M., and Lemieux, C., 1991, A group I intron in the chloroplast large subunit rRNA gene of Chlamydarnonas eugametos encodes a double-strand endonuclease that cleaves the homing site of this intros. Cuff. Genet. 19: 43–47.Google Scholar
  24. Gonzalez, G.C., Caamano, D,L., and Schryvers, A.B., 1990, Identification and characterization of a porcinespecific transferrin receptor in Actinobacillus pleuropnetumoniae. Molec. Microbiol. 4: 1173–1179.Google Scholar
  25. Gonzaleze-Rayos, C., Lo, R.Y.C., Shewen, P.S., and Beveridge, T.J., 1986, Cloning of a serotype-specific antigen from Pasteurella haemolytica Al. infect. Immun. 53: 505–510.Google Scholar
  26. Herrington, D.A., and Spading, P.F., 1985, Haemophilus influenzae can use human transferrin as a sole source for required iron. Infect. Immun. 48: 248–251.Google Scholar
  27. High, N.J., Deadman, M•E., and E.R. Moxon, 1993, The role of a repetitive DNA motif (5’-CAAT-3’) in the variable expression of the Haemophilus influenzae lipopolysaccharide epitope 1 Gal(1–4) Gal. Molec. Microbiol. 9: 1275–1282.Google Scholar
  28. Hone,D., Morona, R., Attridge, S., and Hackett, J., 1987, Construction of defined galE mutants of Salmonella for use as vaccines. J. infect. Dis. 156: 167–174.Google Scholar
  29. Houng, H.S,H., Kopecko, D.J., and Baron, L.S., 1990. Molecular cloning and physical and functional characterization of the Salmonella typhimurium and Salmonella typhi galactose utilization opérons. J. Bacteriol. 172: 4392–4398.Google Scholar
  30. Jennings, M.P., van der Ley, P., Wilks, K.E., Maskell, D.J., Poolman, J.T., and Moxon, E.R., 1993, Cloning and molecular analysis of the galE gene of Neisseria meningitidis and its role in lipopolysaccharide biosynthesis. Molec. Microbiol. 10: 361–369.Google Scholar
  31. Kadner, R.J., 1990, Vitamin B 12 transport in Escherichia coli: energy coupling between membranes. Moles. Microbiol. 4: 2027–2033.Google Scholar
  32. Kciss, R.E., Will, D.H., and Collier. J.R., 1964, Skin toxicity and hemodynamic properties of endotoxin derived from Pasteurella haemolytica. Am. J. Vet. Res. 25: 935–942.Google Scholar
  33. Kolodrubetz, D., Dailey. T., Ebersole. J., and Kraig, E., 1989, Cloning and expression of the leukotoxin gene from Actinobacillus actinomycetemcomitcm.s. infect. immun. 57: 1465–1469.Google Scholar
  34. Koronakis, V., Cross, M., Senior, B., Koronakis, E., and Hughes, C., 1987, The secreted hemolysins of Proteus mirabilis, Proteus vul garis, and Morgane/la morganii are genetically related to each other and to the alpha-hemolysin of Escherichia coli. J. Bacteriol. 169: 1509–1515.PubMedGoogle Scholar
  35. Koronakis, V., Stanley, P., Koronakis, E., and Hughes, C., 1992, The HiyB/HiyD-dependent secretion of toxins by Gram-negative bacteria. FEMS Microbiol. Immunol. 105: 45–54.Google Scholar
  36. LaCroix, R.P., Duncan, J.R., Jenkins, R.P., Leitch. R.A., Petry, J.A., and Richards. J.C., 1993. Structural and serological specificities of Pasteurella haemolytica lipolysaccharides. infect. Immun. 61: 170–181.Google Scholar
  37. Legrain, M., Mazarin. V., Irwin, S.W., Bouchon. B., Quentin-Millet. M., Jacobs, E., and Schryvers, A.B., 1993, Cloning and characterization of Neisseria meningitidis genes encoding the transferrin-binding proteins Tbp 1 and Tbp2. Gene, 130: 73–80.Google Scholar
  38. Lemaire, H.-G., and Muller-Hill. B., 1986, Nucleotide sequences of the galE gene and the galT gene of E. coli. Nucl. Acids Res. 14: 7705–7711.Google Scholar
  39. Lo, R.Y.C., 1990, Molecular characterization of cytotoxins produced by Haemophilus, Actinobacillus, Pasteurella. Can. J. Vet. Res. 54: S33 - S35.PubMedGoogle Scholar
  40. Lo, R.Y.C., 1992, An analysis of the codon usage of Pasteurella haemolytica Al. FEMS Microbiol. Lett. 100: 125–132.Google Scholar
  41. Lo, R.Y.C., and Cameron, L.A., 1986. A simple immunological detection method for the direct screening of genes form clone Banks. Can. J. Biochem. Cell Biol. 64: 73–76.Google Scholar
  42. Lo. R.Y.C., Shewen, P.E., Strathdee, C.A., and Greer, C.N., 1985, Cloning and expression of the leukotoxin gene of Pasteurella haemolytica AI in Escherichia coli K-12. Infect. Immun. 50: 667–671.Google Scholar
  43. Lo, R.Y.C., Strathdee, C.A., and Shewen. P.E., 1987. Nucleotide sequence of the leukotoxin genes of Pasteurella haemolytica A 1. Infect. immun. 55: 1987–1996.PubMedGoogle Scholar
  44. Lo, R.Y.C., Strathdee, C.A., Shewen, P.E., and Cooney, B.J., 1991, Molecular studies of Ssal, a serotypespecific antigen of Pasteurella haemolytica Al. infect. Immun. 59: 3398–3406.Google Scholar
  45. Lo, R.Y.C., Watt, M.-A., Gyorffy, S., and Mellors, A., 1994, Preparation of recombinant glycoprotease of Pasteurella haemolytica Al utilizing the Escherichia coli ahemolysin secretion system. FEMS Microbiol. Lett. 116: 225–230.Google Scholar
  46. Mannheim, W. 1984, In Bergey’s Manual of Systematic Bacteriology, Vol.l. (Kraig, N.R., Ed.). pp. 550–552, Williams and Wilkins Press, Baltimore/London.Google Scholar
  47. Marshall. P., and Lemieux, C., 1991, Cleavage pattern of the homing endonuclease encoded by the fifth intros in the chloroplast large subunit rRNA-encoding gene of Chlamydomonas eugametos. Gene 104: 241–245.CrossRefGoogle Scholar
  48. Marsh, J.C.W., Sutherland, D.R., Davidson, J., Mellors, A., and Keating, A., 1992, Retention of progenitor cell fraction in CD34’ cells purified using a nova) 0-sialoglycoprotease. Leukemia 6: 926–934.PubMedGoogle Scholar
  49. Maskell, D.J., Szabo, M.J., Dcadman, M.E., and Moxon, E.R., 1992, The gal locus from Haemophilus influenzae: cloning, sequencing and the use of gal mutants to study lipopolysaccharide. Molec. Microbiol. 6: 3051–3063.Google Scholar
  50. Mickelsen, P.A., and Sparling, P.F., 1981, Ability of Neisseria gonorrhoeae, Neisseria meningitidis and commensal Neisseria species to obtain iron form transferrin and iron compounds. Infect. Immun. 33: 555–564.Google Scholar
  51. Neilands, J.B., 1981, Microbial iron compounds. Annu. Rev. Microbiol. 50: 715–731.Google Scholar
  52. Ogunnariwo, J.A., and Schryvers. A.B., 1990, iron acquisition in Pasteurella haemolytica: expression and identification of bovine-specific transferrin receptor. Infect. Immun. 58: 2091–2097.Google Scholar
  53. Otto. B.R., Verweij-van Vught, A.M.J.J., and MacLaren. D.M., 1992, Transferrins and heme-compounds as iron sources for pathogenic bacteria. Crit. Rev. Microbiol. 13: 217–233.Google Scholar
  54. Paulsen. D.B., Mosier, D.A., Clinkenbeard, K.D., and Confer, A.W., 1989, Direct effects of Pasteurella haemolytica lipopolysaccharide on bovine pulmonary endothelial cells in vitro. Am. J. Vet. Res. 50: 1633–1637.Google Scholar
  55. Paulsen. D.B., Confer, A.W., Clinkenbeard, K.D., and Mosier, D.A., 1990, Pasteurella haemolytica Google Scholar
  56. lipopolysaccharide-induced arachidonic acid release from and neutrophil adherence to bovine pulmonary artery endothelial cells. Am J. Vet. Res. 51: 1635–1639.Google Scholar
  57. Postle, K., 1990, TonB and the Gram-negative dilemma. Molec. Microbiol. 4: 2019–2025.Google Scholar
  58. Potter, M.D. and Lo, R.Y.C., 1993, Genetic characterization of Pasteurella haemolytica Al LPS biosynthetic genes. 93rd Ann. Meet. Am. Soc. Microbiol., 1993. Atlanta, Georgia, Abstr. B354.Google Scholar
  59. Raetz, C.R.H., 1990, Biochemistry of endotoxins. Annu. Rev. Biochem. 59: 129–170.Google Scholar
  60. Rimsay, R.L., Coyle-Dennis. J.E., Lauerman. L.H., and Squire, P.G., 1981. Purification and biologicalGoogle Scholar
  61. characterization of endotoxin fractions from Pasteurella haemolytica. Am. J. Vet. Res. 42:2134–2138.Google Scholar
  62. Robertson. B.D., Frosch, M., and van Pulten. J.P.M., 1993. The role of galE in the biosynthesis and function of gonococcal lipopolysaccharide. Molec. Microbiol. 8: 891–901.Google Scholar
  63. Schnaitman. C.A., and Klena, J.D., 1993, Genetics of lipopolysaccharide biosynthesis in enteric bacteria. Microbiol. Rev. 57: 655–682.Google Scholar
  64. Schryvers, A.B., and Morris, L.J., 1988, Identification and characterization of the transferrin receptor from Neisseria meningitidis. Molec. Microbiol. 2: 281–288.Google Scholar
  65. Severn, W.B., and Richards, J.C., 1993, Characterization of the O-polysaccharide of Pasteurella haemolytica serotype Al. Carbohydr. Res. 240: 277–285.Google Scholar
  66. Shewen. P.E., and Wilkie, B.N., 1988. Vaccination of calves with leukotoxic culture supernatant from Pasteurella haemolytica. Can. J. Vet. Res. 52: 30–36.Google Scholar
  67. Shewen, P.E., Sharpe, A., and Wilkie, B.N., 1988. Efficacy testing of a Pasteurella haemolytica extract vaccine. Vet. Med. 10: 1078–1083.Google Scholar
  68. Sirisena, D.M., Brozek, K.A., MacLachlan, P.R., Sanderson, K.E., and Raetz, C.R.H., 1992, The rfaC gene of Salmonella typhimurium. J. Biol. Chem. 267: 18874–18884.Google Scholar
  69. Slocombe, R.F., Mulks, M., Killingsworth, C.R., Derksen, F.J., and Robinson. N.E., 1990. Effect of Pasteurella haemolytica-derived endotoxin on pulmonary structure and function in calves. Am. J. Vet. Res. 51: 433–438.Google Scholar
  70. Smits, M.A., Briaire, J., Jansen, R., Smith. H.E., Kamp, E.M., and Gielkens, A.L.J., 1991, Cytolysins of Actinobacillus pleuropnetunoniae serotype 9. Infect. Immun. 59: 4497–4504.Google Scholar
  71. Sneath, P.H.A. and Stevens, M., 1990, Actinobacillus rossi sp. nov., Actinobacillus seminis sp. nov., nom. rev., Pasteurella bettii sp. nov. Pasteurella lymphangitidis sp. nov., Pasteurella main’ sp. nov., and Pasteurella trehalosi sp. Nov. Int. J. Syst. Bact. 40: 148–153.Google Scholar
  72. Strathdee, C.A., 1989, Molecular characterization of the Pasteurella haemolytica leukotoxin determinant. Ph.D. Thesis, University of Guelph, Guelph. Ontario, Canada.Google Scholar
  73. Strathdee, C.A., and Lo, R.Y.C., 1987, Extensive homology between the leukotoxin of Pasteurella haemolytica Al and the alpha-hemolysin of Escherichia coli. Infect. Immun. 55: 3233–3236.Google Scholar
  74. Strathdee. C.A., and Lo. R.Y.C., 1989. Cloning, nucleotide sequence, and characteriztion of genes encoding the secretion function of the Pasteurella haemolytica leukotoxin determinant. J. Bacteriol. 171: 916–928.Google Scholar
  75. Sutherland, A.D., and Donachie, W., 1986, Cytotoxic effect of serotypes of Pasteurella haemolytica on sheep bronchoalveolar macrophages. Vet. Microbiol. 11: 331–336.Google Scholar
  76. Welch, R.A., 1991, Pore-forming cytolysins of Gram-negative bacteria. Molec. Microbiol. 5: 521–528.Google Scholar
  77. Welch, R.A., Forestier, C., Lobo, A., Pellett, S., Thomas, Jr., W., and Rowe, G., 1992, The synthesis and function of the Escherichia coli hemolysin and related RTX exotoxins. FEMS Microbiol. Immunol. 105: 29–36.Google Scholar
  78. Williams, P., and Griffiths, E., 1992, Bacterial transferrin receptors - structure, function and contribution to virulence. Med. Microbial. Immunol. 181: 301–322.Google Scholar
  79. Whiteley, L.O., Maheswaran, S.K., Weiss, D.J., and Ames, T.R., 1990, Immunohistochemical localization of Pasteurella haemolytica A 1-derived endotoxin, leukotoxin, and capsular polysaccharide in experimental bovine pasteurella pneumoniae. Vet Pathol. 27: 150–161.PubMedCrossRefGoogle Scholar
  80. Woo, T.K.W., and Lo, R.Y.C., 1993, The transferrin binding protein of Pasteurella haemolytica is homologous to the gonococcal transferrin binding protein. 93rd Ann. Meet. American Society for Microbiol., Atlanta Georgia, Abstr. B294.Google Scholar

Copyright information

© Springer Science+Business Media New York 1995

Authors and Affiliations

  • R. Y. C. Lo
    • 1
  1. 1.Department of MicrobiologyUniversity of GuelphGuelphCanada

Personalised recommendations