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Gallibacterium elongation factor-Tu possesses amyloid-like protein characteristics, participates in cell adhesion, and is present in biofilms

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Abstract

Gallibacterium, which is a bacterial pathogen in chickens, can form biofilms. Amyloid proteins present in biofilms bind Congo red dye. The aim of this study was to characterize the cell-surface amyloid-like protein expressed in biofilms formed by Gallibacterium strains and determine the relationship between this protein and curli, which is an amyloid protein that is commonly expressed by members of the Enterobacteriaceae family. The presence of amyloid-like proteins in outer membrane protein samples from three strains of G. anatis and one strain of Gallibacterium genomospecies 2 was evaluated. A protein identified as elongation factor-Tu (EF-Tu) by mass spectrometric analysis and in silico analysis was obtained from the G. anatis strain F149T. This protein bound Congo red dye, cross-reacted with anti-curli polyclonal serum, exhibited polymerizing properties and was present in biofilms. This protein also reacted with pooled serum from chickens that were experimentally infected with G. anatis, indicating the in vivo immunogenicity of this protein. The recombinant EF-Tu purified protein, which was prepared from G. anatis 12656-12, polymerizes under in vitro conditions, forms filaments and interacts with fibronectin and fibrinogen, all of which suggest that this protein functions as an adhesin. In summary, EF-Tu from G. anatis presents amyloid characteristics, is present in biofilms and could be relevant for the pathogenesis of G. anatis.

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References

  • Ashman, E.E. and Chapman, R.R. 2008. Polymerizing the fibre between bacteria and host cells: the biogenesis of functional amyloid fibres. Cell Microbiol. 10, 1413–1420.

    Article  Google Scholar 

  • Bager, R.J., Nesta, B., Pors, S.E., Soriani, M., Serino, L., Boyce, J.D., Adler, B., and Bojesen, A.M. 2013. The fimbrial protein FlfA from Gallibacterium anatis is a virulence factor and vaccine candidate. Infect. Immun. 81, 1964–1973.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Balasubramanian, S., Kannan, T.R., and Baseman, J.B. 2008. The surface-exposed carboxyl region of Mycoplasma pneumoniae elongation factor Tu interacts with fibronectin. Infect. Immun. 76, 3116–3123.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Beck, B.D., Arscott, P.G., and Jacobson, A. 1978. Novel properties of bacterial elongation factor Tu. Proc. Natl. Acad. Sci. USA 75, 1250–1254.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bisgaard, M. 1977. Incidence of Pasteurella haemolytica in the respiratory tract of apparently healthy chickens and chickens with infectious bronchitis. Characterization of 213 strains. Avian Pathol. 6, 285–292.

    Article  CAS  Google Scholar 

  • Costerton, J.W., Stewart, P.S., and Greenberg, E.P. 1999. Bacterial biofilms: a common cause of persistent infections. Science 284, 1318–1322.

    Article  CAS  PubMed  Google Scholar 

  • Dallo, S.F., Zhang, B., Denno, J., Hong, S., Tsai, A., Haskins, W., Ye, J.Y., and Weitao, T. 2012. Association of Acinetobacter baumannii EF-Tu with cell surface, outer membrane vesicles, and fibronectin. Scientific World Journal 128705. doi: 10.1100/2012/128705.

    Google Scholar 

  • Foulston, L., Elsholz, A.K., DeFrancesco, A.S., and Losick, R. 2014. The extracellular matrix of Staphylococcus aureus biofilms comprises cytoplasmic proteins that associate with the cell surface in response to decreasing pH. MBio 5, e01667–14.

    Article  Google Scholar 

  • Fowler, D.M., Koulov, A.V., Balch, W.E., and Kelly, J.W. 2007. Functional amyloid–from bacteria to humans. Trends Biochem. Sci. 32, 217–224.

    Article  CAS  PubMed  Google Scholar 

  • Furano, A.V. 1975. Content of elongation factor Tu in Escherichia coli. Proc. Natl. Acad. Sci. USA 72, 4780–4784.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Granato, D., Bergonzelli, G.E., Pridmore, R.D., Marvin, L., Rouvet, M., and Corthésy-Theulaz, I.E. 2004. Cell surface-associated elongation factor Tu mediates the attachment of Lactobacillus johnsonii NCC533 (La1) to human intestinal cells and mucins. Infect. Immun. 72, 2160–2169.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • He, Y., Wang, H., and Chen, L. 2015. Comparative secretomics reveals novel virulence-associated factors of Vibrio parahaemolyticus. Front. Microbiol. 6, 707.

    PubMed  PubMed Central  Google Scholar 

  • Helms, M.K., Marriott, G., Sawyer, W.H., and Jameson, D.M. 1996. Dynamics and morphology of the in vitro polymeric form of elongation factor Tu from Escherichia coli. Biochim. Biophys. Acta. 1291, 122–130.

    Article  PubMed  Google Scholar 

  • Hughes, D. 1990. Both genes for EF-Tu in Salmonella typhimurium are individually dispensable for growth. J. Mol. Biol. 215, 41–51.

    Article  CAS  PubMed  Google Scholar 

  • Johnson, T.J., Danzeisen, J.L., Trampel, D., Nolan, L.K., Seemann, T., Bager, R.J., and Bojesen, A.M. 2013. Genome analysis and phylogenetic relatedness of Gallibacterium anatis strains from poultry. PLoS One 8, e54844.

    Article  Google Scholar 

  • Kimizuka, R., Kato, T., Hashimoto, S., Yamanaka-Okada, A., Okuda, K., and Ishihara, K. 2009. Congo red binding protein in roughphenotype Aggregatibacter actinomycetemcomitans is amyloidlike fiber. Bull. Tokyo Dent. Coll. 50, 23–29.

    Article  CAS  PubMed  Google Scholar 

  • Kittigul, L., Suthachana, S., Kittigul, C., and Pengruangrojanachai, V. 1989. Immunoglobulin M-capture biotin-streptavidin enzymelinked immunosorbent assay for detection of antibodies to dengue viruses. Am. J. Trop. Med. Hyg. 59, 352–356.

    Article  Google Scholar 

  • Larsen, P., Nielsen, J.L., Dueholm, M.S., Wetzel, R., Otzen, D., and Nielsen, P.H. 2007. Amyloid adhesins are abundant in natural biofilms. Environ. Microbiol. 9, 3077–3090.

    Article  CAS  PubMed  Google Scholar 

  • Maurer-Stroh, S., Debulpaep, M., Kuemmerer, N., Lopez de la Paz, M., Martins, I.C., Reumers, J., Morris, K.L., Copland, A., Serpell, L., Serrano, L., et al. 2010. Exploring the sequence determinants of amyloid structure using position-specific scoring matrices. Nat. Methods 7, 237–242.

    Article  CAS  PubMed  Google Scholar 

  • Mena-Rojas, E., Vázquez Cruz, C., Vaca, P.S., García, G.O., Pérez Márquez, V., Pérez Méndez, A., Ibarra-Caballero, J., de la Garza, M., Zenteno, E., and Negrete-Abascal, E. 2004. Antigenic secreted proteins from Haemophilus paragallinarum. A 110 kDa putative RTX-protein. FEMS Microbiol. Lett. 232, 83–87.

    Article  CAS  PubMed  Google Scholar 

  • Meneses, N., Mendoza-Hernandez, G., and Encarnacion, S. 2010. The extracellular proteome of Rhizobium etli CE3 in exponential and stationary growth phase. Proteome Sci. 8, 51.

    Article  PubMed  PubMed Central  Google Scholar 

  • Montes-García, J.F., Vaca, S., Vazquez-Cruz, C., Soriano-Vargas, E., Aguilar-Romero, F., Blackall, P.J., and Negrete Abascal, E. 2016. Identification of a hemagglutinin from Gallibacterium anatis. Curr. Microbiol. 72, 450–456.

    Article  PubMed  Google Scholar 

  • Muñoz-Pruvencio, D., Pérez-Martínez, G., and Monedero, A. 2011. Identification of surface proteins from Lactobacillus casei BL23 able to bind fibronecting and collagen. Probiotics Antimicrob. Proteins 3, 15–20.

    Google Scholar 

  • Neubauer, C., De Souza-Pilz, M., Bojesen, A.M., Bisgaard, M., and Hess, M. 2009. Tissue distribution of haemolytic Gallibacterium anatis isolates in laying birds with reproductive disorders. Avian Pathol. 38, 1–7.

    Article  CAS  PubMed  Google Scholar 

  • Nieves, W., Heang, J., Asakrah, S., Höner zu Bentrup, K., Roy, C.J., and Morici, L.A. 2010. Immunospecific responses to bacterial elongation factor Tu during Burkholderia infection and immunization. PLoS One 5, e14361.

    Article  Google Scholar 

  • Nishiyama, K., Ochiai, A., Tsubokawa, D., Ishihara, K., Yamamoto, Y., and Mukai, T. 2013. Identification and characterization of sulfated carbohydrate-binding protein from Lactobacillus reuteri. PLoS One 8, e83703.

    Article  Google Scholar 

  • Papa, R., Artini, M., Cellini, A., Tilotta, M., Galano, E., Pucci, P., Amoresano, A., and Selan, L. 2013. New anti-infective strategy to reduce the spreading of antibiotic resistance by the action on adhesion-mediated virulence factors in Staphylococcus aureus. Microb. Pathog. 63, 44–53.

    Article  CAS  PubMed  Google Scholar 

  • Persson, G. and Bojesen, A.M. 2015. Bacterial determinants of importance in the virulence of Gallibacterium anatis in poultry. Vet. Res. 46, 57.

    Article  PubMed  PubMed Central  Google Scholar 

  • Reichhardt, C., McCrate, O.A., Zhou, X., Lee, J., Thongsomboon, W., and Cegelski, L. 2016. Influence of the amyloid dye Congo red on curli, cellulose, and the extracellular matrix in E.coli during growth and matrix purification. Anal Bioanal. Chem. 408, 7709–7717.

    Article  CAS  PubMed  Google Scholar 

  • Salgado-Lucio, M.L., Vaca, S., Vázquez, C., Zenteno, E., Pérez-Márquez, V.M., and Negrete Abascal, E. 2012. Adhesion of Gallibacterium anatis to chicken oral-pharyngeal epithelial cells and identification of putative fimbriae. Adv. Microbiol. 2, 505–510.

    Article  Google Scholar 

  • Sambrook, J., Fritsch, E.F., and Maniatis, F. 1989 Molecular Cloning. A laboratory manual. Second Edition. Cold Spring Harbor Laboratory.

    Google Scholar 

  • Sharma, J., Mishra, B.B., Li, Q., and Teale, J.M. 2011. TLR4-dependent activation of inflammatory cytokine response in macrophages by Francisella elongation factor Tu. Cell Immunol. 269, 69–73.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shinoy, M., Dennehy, R., Coleman, L., Carberry, S., Schaffer, K., Callaghan, M., Doyle, S., and McClean, S. 2013. Immunoproteomic analysis of proteins expressed by two related pathogens, Burkholderia multivorans and Burkholderia cenocepacia, during human infection. PLoS One 8, e80796.

    Article  Google Scholar 

  • Sitaras, C., Naghavi, M., Herrington, M.B. 2011. Sodium dodecyl sulfate–agarose gel electrophoresis for the detection and isolation of amyloid curli fibers. Anal. Biochem. 408, 328–331.

    Article  CAS  PubMed  Google Scholar 

  • Szabó, E., Skedsmo, A., Sonnevend, A., Al-Dhaheri, K., Emody, L., Usmani, A., and Pál, T. 2005. Curli expression of enterotoxigenic Escherichia coli. Folia Microbiol. (Praha) 50, 40–46.

    Article  Google Scholar 

  • Uhlich, G.A., Cooke, P.H., and Solomon, E.B. 2006. Analyses of the red-dry-rough phenotype of an Escherichia coli O157:H7 strain and its role in biofilm formation and resistance to antibacterial agents. Appl. Environ. Microbiol. 72, 2564–2572.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vaca, S., Monroy, E., Rojas, L., Vázquez, C., Sánchez, P., Soriano Vargas, E., Bojesen, A.M., and Negrete Abascal, E. 2011. Adherence of Gallibacterium anatis to inert surfaces. J. Anim. Vet. Adv. 10, 1688–1693.

    Article  CAS  Google Scholar 

  • Vanden Bergh, P., Heller, M., Braga-Lagache, S., and Joachim, F. 2013. The Aeromonas salmonicida subsp. salmonicida exoproteome: global analysis, moonlighting proteins and putative antigens for vaccination against furunculosis. Proteome Sci. 11, 44.

    Article  PubMed  PubMed Central  Google Scholar 

  • Wolff, D.G., Castiblanco-Valencia, M.M., Abe, C.M., Monaris, D., Souza, G.O., Vasconcellos, S.A., Isaac, L., Abreu, P.A., and Barbosa, A.S. 2013. Interaction of Leptospira elongation factor Tu with plasminogen and complement factor H: A metabolic leptospiral protein with moonlighting activities. PLoS One 8, e81818.

    Article  Google Scholar 

  • Xicohtencatl-Cortes, J., Saldaña, Z., Deng, W., Castañeda, E., Freer, E., Tarr, P.I., Finlay, B.B., Puente, J.L., and Girón, J.A. 2010. Bacterial macroscopic rope-like fibres with cytopathic and adhesive properties. J. Biol. Chem. 285, 32336–32342.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zepeda, A., Ramírez, S., Vega, V., Morales, V., Talavera, M., Salgado-Miranda, C., Simón-Martínez, J., Bojesen, A.M., and Soriano-Vargas, E. 2009. Hemagglutinating activity of Gallibacterium strains. Avian Dis. 53, 115–118.

    Article  CAS  PubMed  Google Scholar 

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Correspondence to Erasmo Negrete-Abascal.

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López-Ochoa, J., Montes-García, J.F., Vázquez, C. et al. Gallibacterium elongation factor-Tu possesses amyloid-like protein characteristics, participates in cell adhesion, and is present in biofilms. J Microbiol. 55, 745–752 (2017). https://doi.org/10.1007/s12275-017-7077-0

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  • DOI: https://doi.org/10.1007/s12275-017-7077-0

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