Characters of homogentisate oxygenase gene mutation and high clonality of the natural pigment-producing Vibrio cholerae strains

BMC Microbiology

Biao Kan²  Baowei Diao²  Junjie Yue¹  Yuelan Wang¹  Haijian Zhou²  Hengliang Wang¹  Ruibai Wang² 

[1] Beijing Institute of Biotechnology, State Key Laboratory of Pathogen and Biosecurity, Beijing 100071, PR China;State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, PR China

关键词: clonality;    homogentisate oxygenase;    pigment;    Vibrio cholerae;   

Others  :  1223696
DOI  :  10.1186/1471-2180-11-109

 received in 2010-12-21, accepted in 2011-05-18,  发布年份 2011

【 摘 要 】

Background

Some microorganisms can produce pigments such as melanin, which has been associated with virulence in the host and with a survival advantage in the environment. In Vibrio cholerae, studies have shown that pigment-producing mutants are more virulent than the parental strain in terms of increased UV resistance, production of major virulence factors, and colonization. To date, almost all of the pigmented V. cholerae strains investigated have been induced by chemicals, culture stress, or transposon mutagenesis. However, during our cholera surveillance, some nontoxigenic serogroup O139 strains and one toxigenic O1 strain, which can produce pigment steadily under the commonly used experimental growth conditions, were obtained in different years and from different areas. The genes VC1344 to VC1347, which correspond to the El Tor strain N16961 genome and which comprise an operon in the tyrosine catabolic pathway, have been confirmed to be associated with a pigmented phenotype. In the present study, we investigated the mechanism of pigment production in these strains.

Results

Sequencing of the VC1344, VC1345, VC1346, and VC1347 genes in these pigmented strains suggested that a deletion mutation in the homogentisate oxygenase gene (VC1345) may be associated with the pigmented phenotype, and gene complementation confirmed the role of this gene in pigment production. An identical 15-bp deletion was found in the VC1345 gene of all six O139 pigment-producing strains examined, and a 10-bp deletion was found in the VC1345 gene of the O1 strain. Strict sequence conservation in the VC1344 gene but higher variance in the other three genes of this operon were observed, indicating the different stress response functions of these genes in environmental adaption and selection. On the basis of pulsed-field gel electrophoresis typing, the pigment-producing O139 strains showed high clonality, even though they were isolated in different years and from different regions. Additionally all these O139 strains belong to the rb4 ribotype, which contains the O139 strains isolated from diarrheal patients, although these strains are cholera toxin negative.

Conclusion

Dysfunction of homogentisate oxygenase (VC1345) causes homogentisate accumulation and pigment formation in naturally pigmented strains of V. cholerae. The high clonality of these strains may correlate to an environmental survival advantage in the V. cholerae community due to their pigment production, and may imply a potential protective function of melanin in environmental survival of such strains.

【 授权许可】

   
2011 Wang et al; licensee BioMed Central Ltd.

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【 参考文献 】

• [1]Kaper JB, Morris JG Jr, Levine MM: Cholera. Clin Microbiol Rev 1995, 8(1):48-86.
• [2]Reidl J, Klose KE: Vibrio cholerae and cholera: out of the water and into the host. FEMS Microbiol Rev 2002, 26(2):125-139.
• [3]Karaolis DK, Johnson JA, Bailey CC, Boedeker EC, Kaper JB, Reeves PR: A Vibrio cholerae pathogenicity island associated with epidemic and pandemic strains. Proc Natl Acad Sci USA 1998, 95(6):3134-3139.
• [4]Coelho A, Andrade JR, Vicente AC, Dirita VJ: Cytotoxic cell vacuolating activity from Vibrio cholerae hemolysin. Infect Immun 2000, 68(3):1700-1705.
• [5]Lin W, Fullner KJ, Clayton R, Sexton JA, Rogers MB, Calia KE, Calderwood SB, Fraser C, Mekalanos JJ: Identification of a vibrio cholerae RTX toxin gene cluster that is tightly linked to the cholera toxin prophage. Proc Natl Acad Sci USA 1999, 96(3):1071-1076.
• [6]von Kruger WM, Humphreys S, Ketley JM: A role for the PhoBR regulatory system homologue in the Vibrio cholerae phosphate-limitation response and intestinal colonization. Microbiology 1999, 145(Pt 9):2463-2475.
• [7]Paz S: Impact of temperature variability on cholera incidence in southeastern Africa, 1971-2006. Ecohealth 2009, 6(3):340-345.
• [8]Sedas VT: Influence of environmental factors on the presence of Vibrio cholerae in the marine environment: a climate link. J Infect Dev Ctries 2007, 1(3):224-241.
• [9]Constantin de Magny G, Colwell RR: Cholera and climate: a demonstrated relationship. Trans Am Clin Climatol Assoc 2009, 120:119-128.
• [10]Nosanchuk JD, Casadevall A: The contribution of melanin to microbial pathogenesis. Cell Microbiol 2003, 5(4):203-223.
• [11]Nosanchuk JD, Casadevall A: Impact of melanin on microbial virulence and clinical resistance to antimicrobial compounds. Antimicrob Agents Chemother 2006, 50(11):3519-3528.
• [12]Steenbergen JN, Casadevall A: The origin and maintenance of virulence for the human pathogenic fungus Cryptococcus neoformans. Microbes Infect 2003, 5(7):667-675.
• [13]Brownlee JM, Johnson-Winters K, Harrison DH, Moran GR: Structure of the ferrous form of (4-hydroxyphenyl)pyruvate dioxygenase from Streptomyces avermitilis in complex with the therapeutic herbicide, NTBC. Biochemistry 2004, 43(21):6370-6377.
• [14]Kavana M, Moran GR: Interaction of (4-hydroxyphenyl)pyruvate dioxygenase with the specific inhibitor 2-[2-nitro-4-(trifluoromethyl)benzoyl]-1,3-cyclohexanedione. Biochemistry 2003, 42(34):10238-10245.
• [15]Sanchez-Amat A, Ruzafa C, Solano F: Comparative tyrosine degradation in Vibrio cholerae strains. The strain ATCC 14035 as a prokaryotic melanogenic model of homogentisate-releasing cell. Comp Biochem Physiol B Biochem Mol Biol 1998, 119(3):557-562.
• [16]Lerner AB, Fitzpatrick TB: Biochemistry of melanin formation. Physiol Rev 1950, 30(1):91-126.
• [17]Wheeler MH, Bell AA: Melanins and their importance in pathogenic fungi. Curr Top Med Mycol 1988, 2:338-387.
• [18]Ivins BE, Holmes RK: Isolation and characterization of melanin-producing (mel) mutants of Vibrio cholerae. Infect Immun 1980, 27(3):721-729.
• [19]Ivins BE, Holmes RK: Factors affecting phaeomelanin production by a melanin-producing (mel) mutant of Vibrio cholerae. Infect Immun 1981, 34(3):895-899.
• [20]Coyne VE, al-Harthi L: Induction of melanin biosynthesis in Vibrio cholerae. Appl Environ Microbiol 1992, 58(9):2861-2865.
• [21]Kotob SI, Coon SL, Quintero EJ, Weiner RM: Homogentisic acid is the primary precursor of melanin synthesis in Vibrio cholerae, a Hyphomonas strain, and Shewanella colwelliana. Appl Environ Microbiol 1995, 61(4):1620-1622.
• [22]Ruzafa C, Sanchez-Amat A, Solano F: Characterization of the melanogenic system in Vibrio cholerae, ATCC 14035. Pigment Cell Res 1995, 8(3):147-152.
• [23]Valeru SP, Rompikuntal PK, Ishikawa T, Vaitkevicius K, Sjoling A, Dolganov N, Zhu J, Schoolnik G, Wai SN: Role of melanin pigment in expression of Vibrio cholerae virulence factors. Infect Immun 2009, 77(3):935-942.
• [24]Wang RB, Gao SY, Kan B: Application of transposon to screening of pigment-production genes of Vibrio cholerae. Bull Acad Millit Med Sci 2005, 29(4):4.
• [25]Cooper KL, Luey CK, Bird M, Terajima J, Nair GB, Kam KM, Arakawa E, Safa A, Cheung DT, Law CP, et al.: Development and validation of a PulseNet standardized pulsed-field gel electrophoresis protocol for subtyping of Vibrio cholerae. Foodborne Pathog Dis 2006, 3(1):51-58.
• [26]Heidelberg JF, Eisen JA, Nelson WC, Clayton RA, Gwinn ML, Dodson RJ, Haft DH, Hickey EK, Peterson JD, Umayam L, et al.: DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae. Nature 2000, 406(6795):477-483.
• [27]Qu M, Xu J, Ding Y, Wang R, Liu P, Kan B, Qi G, Liu Y, Gao S: Molecular epidemiology of Vibrio cholerae O139 in China: polymorphism of ribotypes and CTX elements. J Clin Microbiol 2003, 41(6):2306-2310.
• [28]Titus GP, Mueller HA, Burgner J, Rodriguez De Cordoba S, Penalva MA, Timm DE: Crystal structure of human homogentisate dioxygenase. Nat Struct Biol 2000, 7(7):542-546.