Contribution of the Outer Membrane Protein OmpQ to biofilm formation by Bordetella bronchiseptica

CATTELÁN NATALIA; SERRA, DIEGO; ARNAL LAURA; YANTORNO OSVALDO

Mar del Plata

Congreso; VIII Congreso Argentino de Microbiología General; 2012

Sociedad Argentina de Microbiología General

Bordetella species are small aerobic, Gram-negative bacteria that colonize the respiratory tract of humansand animals. Bordetella pertussis, a strictly obligate human pathogen, is the etiologic agent of whoopingcough, while B. bronchiseptica mainly infects animals causing a variety of respiratory diseases (1). It hasbeen demonstrated by us and other groups that these bacteria are capable of living as sessilecommunities, known as biofilms, on abiotic surfaces and in the respiratory tract of mice (2-4). OmpQ is ageneral outer membrane porin of Bordetella, the expression of which is regulated by the two-componentsystem BvgAS (6). We had previously found, using proteomic approaches, a higher expression level of thegeneral porins OmpP and OmpQ in B. pertussis biofilm cells than in planktonic cells, either for referenceand clinical strains. (5). The purpose of this work was to study the contribution of OmpQ in the biofilmformation by B. bronchiseptica. To this aim an in-frame, non-polar mutation was first obtained throughallelic exchange on B. bronchiseptica RB50 strain. Then, we analyzed in a comparative manner theplanktonic growth and the biofilm formation ability of the wild-type and the mutant strain using themicrotiter dish assay and Confocal Laser Scanner Microscopy (CLSM). We found that the mutation had noeffect on the planktonic growth. Regarding biofilm development, the ability of the mutant strain to adhereto cover glasses, evaluated by epifluorescense microscopy, showed no significant differences between thetwo strains at the attachment stage. However, the quantification of the biomass adhered to the wells ofmicrotiter plates at 48 and 72 h of biofilm culture showed significant differences between wild-type andOmpQ defective strains. Confocal images, acquired from bacterial cultures grown as biofilm expressing thegreen fluorescent protein, were analyzed with the COMSTAT2 software. Biofilms developed by the wildtypestrain exhibited an average thickness of 33.021 ± 3,853 m after 72h of growth. In contrast, underthe same experimental conditions the ompQ mutant strain formed a biofilm with an average thickness of12.957 ± 3,604 m. Overall, these data showed that the presence of OmpQ is not required for theattachment of B. bronchiseptica to abiotic surfaces but appears to have an important role for biofilmdevelopment. Our results contribute to the identification of new key factors regulated by the BvgAS systemfor Bordetella biofilm formation, leading to a better understanding of the mechanisms involved in thisprocess. 1- Mattoo, S. and Cherry, J.D. Clin Microbiol Rev, 2005. 18(2): p. 326-82. 2- Mishra, M., et al. JBacteriol, 2005. 187(4): p. 1474-84. 3- Sloan, G.P., et al. J Bacteriol, 2007. 189(22): p. 8270-6. 4- Serra,D.O., et al. PLoS ONE, 2012. 6(12): e28811. doi:10.1371/journal.pone.0028811 5- Serra, D.O., et al.Proteomics, 2008. 8(23-24): p. 4995-5010. 6- Finn, T.M., et al. J Bacteriol, 1995. 177(3): p. 805-9.