1887

Abstract

The brucellae are facultative intracellular pathogens of mammals that are transmitted by contact with infected animals or contaminated materials. Several major lipidic components of the brucella cell envelope are imperfectly recognized by innate immunity, thus contributing to virulence. These components carry large proportions of acyl chains of lactobacillic acid, a long chain cyclopropane fatty acid (CFA). CFAs result from addition of a methylene group to unsaturated acyl chains and contribute to resistance to acidity, dryness and high osmolarity in many bacteria and to virulence in mycobacteria. We examined the role of lactobacillic acid in virulence by creating a mutant in ORF BAB1_0476, the putative CFA synthase gene. The mutant did not incorporate [C]methyl groups into lipids, lacked CFAs and synthesized the unsaturated precursors, proving that BAB1_0476 actually encodes a CFA synthase. BAB1_0476 promoter–AB fusion studies showed that CFA synthase expression was promoted by acid pH and high osmolarity. The mutant was not attenuated in macrophages or mice, strongly suggesting that CFAs are not essential for intracellular life. However, when the mutant was tested under high osmolarity on agar and acid pH, two conditions likely to occur on contaminated materials and fomites, they showed reduced ability to grow or survive. Since CFA synthesis entails high ATP expenses and brucellae produce large proportions of lactobacillic acyl chains, we speculate that the CFA synthase has been conserved because it is useful for survival extracellularly, thus facilitating persistence in contaminated materials and transmission to new hosts.

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2012-04-01
2020-01-26
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References

  1. Alton G. G., Jones L. M., Angus R. D., Verger J. M.. ( 1988;). Techniques for the Brucellosis Laboratory Paris: INRA;
    [Google Scholar]
  2. Arellano-Reynoso B., Lapaque N., Salcedo S., Briones G., Ciocchini A. E., Ugalde R., Moreno E., Moriyón I., Gorvel J. P.. ( 2005;). Cyclic β-1,2-glucan is a Brucella virulence factor required for intracellular survival. Nat Immunol6:618–625 [CrossRef][PubMed]
    [Google Scholar]
  3. Barkan D., Rao V., Sukenick G. D., Glickman M. S.. ( 2010;). Redundant function of cmaA2 and mmaA2 in Mycobacterium tuberculosis cis cyclopropanation of oxygenated mycolates. J Bacteriol192:3661–3668 [CrossRef][PubMed]
    [Google Scholar]
  4. Barquero-Calvo E., Chaves-Olarte E., Weiss D. S., Guzmán-Verri C., Chacón-Díaz C., Rucavado A., Moriyón I., Moreno E.. ( 2007;). Brucella abortus uses a stealthy strategy to avoid activation of the innate immune system during the onset of infection. PLoS ONE2:e631 [CrossRef][PubMed]
    [Google Scholar]
  5. Bligh E. G., Dyer W. J.. ( 1959;). A rapid method of total lipid extraction and purification. Can J Biochem Physiol37:911–917 [CrossRef][PubMed]
    [Google Scholar]
  6. Chang Y. Y., Eichel J., Cronan J. E. Jr. ( 2000;). Metabolic instability of Escherichia coli cyclopropane fatty acid synthase is due to RpoH-dependent proteolysis. J Bacteriol182:4288–4294 [CrossRef][PubMed]
    [Google Scholar]
  7. Coloe P. J., Sinclair A. J., Slattery J. F., Burke D.. ( 1984;). Differentiation of Brucella ovis from Brucella abortus by gas-liquid chromatographic analysis of cellular fatty acids. J Clin Microbiol19:896–898[PubMed]
    [Google Scholar]
  8. Conde-Álvarez R., Grilló M. J., Salcedo S. P., de Miguel M. J., Fugier E., Gorvel J. P., Moriyón I., Iriarte M.. ( 2006;). Synthesis of phosphatidylcholine, a typical eukaryotic phospholipid, is necessary for full virulence of the intracellular bacterial parasite Brucella abortus . Cell Microbiol8:1322–1335 [CrossRef][PubMed]
    [Google Scholar]
  9. Corbel M. J.. ( 2006;). Brucellosis in Humans and Animals Geneva: WHO Press;
    [Google Scholar]
  10. Courtois F., Guérard C., Thomas X., Ploux O.. ( 2004;). Escherichia coli cyclopropane fatty acid synthase. Eur J Biochem271:4769–4778 [CrossRef][PubMed]
    [Google Scholar]
  11. Cronan J. E. Jr. ( 2002;). Phospholipid modifications in bacteria. Curr Opin Microbiol5:202–205 [CrossRef][PubMed]
    [Google Scholar]
  12. Dao D. N., Sweeney K., Hsu T., Gurcha S. S., Nascimento I. P., Roshevsky D., Besra G. S., Chan J., Porcelli S. A., Jacobs W. R.. ( 2008;). Mycolic acid modification by the mmaA4 gene of M. tuberculosis modulates IL-12 production. PLoS Pathog4:e1000081 [CrossRef][PubMed]
    [Google Scholar]
  13. De B. K., Stauffer L., Koylass M. S., Sharp S. E., Gee J. E., Helsel L. O., Steigerwalt A. G., Vega R., Clark T. A.. & other authors ( 2008;). Novel Brucella strain (BO1) associated with a prosthetic breast implant infection. Clin Microbiol46:43–49 [CrossRef]
    [Google Scholar]
  14. Dees S. B., Hollis D. G., Weaver R. E., Moss C. W.. ( 1981;). Cellular fatty acids of Brucella canis and Brucella suis . J Clin Microbiol14:111–112[PubMed]
    [Google Scholar]
  15. Dricot A., Rual J. F., Lamesch P., Bertin N., Dupuy D., Hao T., Lambert C., Hallez R., Delroisse J. M.. & other authors ( 2004;). Generation of the Brucella melitensis ORFeome version 1.1. Genome Res14:2201–2206 [CrossRef][PubMed]
    [Google Scholar]
  16. Freer E., Moreno E., Moriyón I., Pizarro-Cerdá J., Weintraub A., Gorvel J. P.. ( 1996;). BrucellaSalmonella lipopolysaccharide chimeras are less permeable to hydrophobic probes and more sensitive to cationic peptides and EDTA than are their native Brucella sp. counterparts. J Bacteriol178:5867–5876[PubMed]
    [Google Scholar]
  17. Gamazo C., Moriyón I.. ( 1987;). Release of outer membrane fragments by exponentially growing Brucella melitensis cells. Infect Immun55:609–615[PubMed]
    [Google Scholar]
  18. Gerhardt P.. ( 1958;). The nutrition of brucellae. Bacteriol Rev22:81–98[PubMed]
    [Google Scholar]
  19. Glickman M. S., Cox J. S., Jacobs W. R. Jr. ( 2000;). A novel mycolic acid cyclopropane synthetase is required for cording, persistence, and virulence of Mycobacterium tuberculosis . Mol Cell5:717–727 [CrossRef][PubMed]
    [Google Scholar]
  20. Gómez-Miguel M. J., Moriyón I.. ( 1986;). Demonstration of a peptidoglycan-linked lipoprotein and characterization of its trypsin fragment in the outer membrane of Brucella spp. Infect Immun53:678–684[PubMed]
    [Google Scholar]
  21. Gorvel J. P., Moreno E., Moriyón I.. ( 2009;). Is Brucella an enteric pathogen?. Nat Rev Microbiol7:250–, author reply 250 [CrossRef][PubMed]
    [Google Scholar]
  22. Grilló M. J., Manterola L., de Miguel M. J., Muñoz P. M., Blasco J. M., Moriyón I., López-Goñi I.. ( 2006;). Increases of efficacy as vaccine against Brucella abortus infection in mice by simultaneous inoculation with avirulent smooth bvrS/bvrR and rough wbkA mutants. Vaccine24:2910–2916 [CrossRef][PubMed]
    [Google Scholar]
  23. Grogan D. W., Cronan J. E. Jr. ( 1997;). Cyclopropane ring formation in membrane lipids of bacteria. Microbiol Mol Biol Rev61:429–441[PubMed]
    [Google Scholar]
  24. Hacker S., Sohlenkamp C., Aktas M., Geiger O., Narberhaus F.. ( 2008;). Multiple phospholipid N-methyltransferases with distinct substrate specificities are encoded in Bradyrhizobium japonicum . J Bacteriol190:571–580 [CrossRef][PubMed]
    [Google Scholar]
  25. Hallez R., Letesson J. J., Vandenhaute J., De Bolle X.. ( 2007;). Gateway-based destination vectors for functional analyses of bacterial ORFeomes: application to the Min system in Brucella abortus . Appl Environ Microbiol73:1375–1379 [CrossRef][PubMed]
    [Google Scholar]
  26. Huddleson I. F.. ( 1943;). Brucellosis in Men and Animals, Revised Edn.. New York: The Commonwealth Fund;
    [Google Scholar]
  27. Ingrosso D., Fowler A. V., Bleibaum J., Clarke S.. ( 1989;). Sequence of the d-aspartyl/l-isoaspartyl protein methyltransferase from human erythrocytes. Common sequence motifs for protein, DNA, RNA, and small molecule S-adenosylmethionine-dependent methyltransferases. J Biol Chem264:20131–20139[PubMed]
    [Google Scholar]
  28. Iñón de Iannino N., Briones G. C., Iannino F., Ugalde R. A.. ( 2000;). Osmotic regulation of cyclic 1,2-β-glucan synthesis. Microbiology146:1735–1742[PubMed]
    [Google Scholar]
  29. Iriarte M., González D., Delrue R. M., Monreal D., Conde-Álvarez R., López-Goñi I., Letesson J. J., Moriyón I.. ( 2004;). Brucella lipopolysacharide: structure, biosynthesis and genetics. Brucella: Molecular and Cellular Biology159–192 López-Goñi I., Moriyón I.. Wymondham, UK: Horizon Bioscience;
    [Google Scholar]
  30. Jubier-Maurin V., Loisel S., Liautard J. P., Köhler S.. ( 2004;). The intramacrophagic environment of Brucella spp. and their replicative niche. Brucella: Molecular and Cellular Biology313–340 López-Goñi I., Moriyón I.. Wymondham, UK: Horizon Bioscience;
    [Google Scholar]
  31. Kaneko T., Nakamura Y., Sato S., Asamizu E., Kato T., Sasamoto S., Watanabe A., Idesawa K., Ishikawa A.. & other authors ( 2000;). Complete genome structure of the nitrogen-fixing symbiotic bacterium Mesorhizobium loti . DNA Res7:331–338 [CrossRef][PubMed]
    [Google Scholar]
  32. Lapaque N., Takeuchi O., Corrales F., Akira S., Moriyón I., Howard J. C., Gorvel J. P.. ( 2006;). Differential inductions of TNF-α and IGTP, IIGP by structurally diverse classic and non-classic lipopolysaccharides. Cell Microbiol8:401–413 [CrossRef][PubMed]
    [Google Scholar]
  33. Martínez de Tejada G., Moriyón I.. ( 1993;). The outer membranes of Brucella spp. are not barriers to hydrophobic permeants. J Bacteriol175:5273–5275[PubMed]
    [Google Scholar]
  34. Martínez de Tejada G., Pizarro-Cerdá J., Moreno E., Moriyón I.. ( 1995;). The outer membranes of Brucella spp. are resistant to bactericidal cationic peptides. Infect Immun63:3054–3061[PubMed]
    [Google Scholar]
  35. Moriyón I., Berman D. T.. ( 1982;). Effects of nonionic, ionic, and dipolar ionic detergents and EDTA on the Brucella cell envelope. J Bacteriol152:822–828[PubMed]
    [Google Scholar]
  36. Palacios-Chaves L., Conde-Álvarez R., Gil-Ramírez Y., Zúñiga-Ripa A., Barquero-Calvo E., Chacón-Díaz C., Chaves-Olarte E., Arce-Gorvel V., Gorvel J. P.. & other authors ( 2011;). Brucella abortus ornithine lipids are dispensable outer membrane components devoid of a marked pathogen-associated molecular pattern. PLoS ONE6:e16030 [CrossRef][PubMed]
    [Google Scholar]
  37. Rao V., Fujiwara N., Porcelli S. A., Glickman M. S.. ( 2005;). Mycobacterium tuberculosis controls host innate immune activation through cyclopropane modification of a glycolipid effector molecule. J Exp Med201:535–543 [CrossRef][PubMed]
    [Google Scholar]
  38. Rao V., Gao F., Chen B., Jacobs W. R. Jr, Glickman M. S.. ( 2006;). Trans-cyclopropanation of mycolic acids on trehalose dimycolate suppresses Mycobacterium tuberculosis-induced inflammation and virulence. J Clin Invest116:1660–1667 [CrossRef][PubMed]
    [Google Scholar]
  39. Roop R. M. II, Gee J. M., Robertson G. T., Richardson J. M., Ng W. L., Winkler M. E.. ( 2003;). Brucella stationary-phase gene expression and virulence. Annu Rev Microbiol57:57–76[PubMed][CrossRef]
    [Google Scholar]
  40. Roset M. S., Ciocchini A. E., Ugalde R. A., Iñón de Iannino N.. ( 2006;). The Brucella abortus cyclic beta-1,2-glucan virulence factor with O-ester-linked succinyl residues. J Bacteriol188:5003–5013[CrossRef]
    [Google Scholar]
  41. Saborido Basconcillo L., Zaheer R., Finan T. M., McCarry B. E.. ( 2009;). Cyclopropane fatty acyl synthase in Sinorhizobium meliloti . Microbiology155:373–385 [CrossRef][PubMed]
    [Google Scholar]
  42. Scupham A. J., Triplett E. W.. ( 1997;). Isolation and characterization of the UDP-glucose 4′-epimerase-encoding gene, galE, from Brucella abortus 2308. Gene202:53–59 [CrossRef][PubMed]
    [Google Scholar]
  43. Sieira R., Comerci D. J., Sánchez D. O., Ugalde R. A.. ( 2000;). A homologue of an operon required for DNA transfer in Agrobacterium is required in Brucella abortus for virulence and intracellular multiplication. J Bacteriol182:4849–4855 [CrossRef][PubMed]
    [Google Scholar]
  44. Theys T. E., Geeraerd A. H., Verhulst A., Poot K., Van Bree I., Devlieghere F., Moldenaers P., Wilson D., Brocklehurst T., Van Impe J. F.. ( 2008;). Effect of pH, water activity and gel micro-structure, including oxygen profiles and rheological characterization, on the growth kinetics of Salmonella Typhimurium. Int J Food Microbiol128:67–77 [CrossRef][PubMed]
    [Google Scholar]
  45. Thiele O. W., Schwinn G.. ( 1973;). The free lipids of Brucella melitensis and Bordetella pertussis . Eur J Biochem34:333–344 [CrossRef][PubMed]
    [Google Scholar]
  46. Thiele O. W., Schwinn G.. ( 1974;). Bacterial ornithine lipids. Z Allg Mikrobiol14:435–443 [CrossRef][PubMed]
    [Google Scholar]
  47. Thiele O. W., Lacave C., Asselineau J.. ( 1969;). On the fatty acids of Brucella abortus and Brucella melitensis . Eur J Biochem7:393–396 [CrossRef][PubMed]
    [Google Scholar]
  48. Vasiurenko Z. P., Siniak K. M., Korotich A. S., Antonova L. A.. ( 1977;). Fatty acid makeup of various Brucella species and its relationship to the culture medium. Zh Mikrobiol Epidemiol Immunobiol8:53–59[PubMed]
    [Google Scholar]
  49. Velasco J., Bengoechea J. A., Brandenburg K., Lindner B., Seydel U., González D., Zähringer U., Moreno E., Moriyón I.. ( 2000;). Brucella abortus and its closest phylogenetic relative, Ochrobactrum spp., differ in outer membrane permeability and cationic peptide resistance. Infect Immun68:3210–3218 [CrossRef][PubMed]
    [Google Scholar]
  50. Viadas C., Rodríguez M. C., Sangari F. J., Gorvel J. P., García-Lobo J. M., López-Goñi I.. ( 2010;). Transcriptome analysis of the Brucella abortus BvrR/BvrS two-component regulatory system. PLoS ONE5:e10216 [CrossRef][PubMed]
    [Google Scholar]
  51. Wang A. Y., Cronan J. E. Jr. ( 1994;). The growth phase-dependent synthesis of cyclopropane fatty acids in Escherichia coli is the result of an RpoS(KatF)-dependent promoter plus enzyme instability. Mol Microbiol11:1009–1017 [CrossRef][PubMed]
    [Google Scholar]
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