Cellular Microbiology (2006) 8(12), 1877–1887 doi:10.1111/j.1462-5822.2006.00754.x First published online 4 July 2006 © 2006 The Authors Journal compilation © 2006 Blackwell Publishing Ltd Blackwell Publishing LtdOxford, UKCMICellular Microbiology1462-5814© 2006 The Authors; Journal compilation © 2006 Blackwell Publishing Ltd 200681218771887Original ArticleBrucella SP41 adhesinE. I. Castañeda-Roldán et al. Received 25 April, 2006; revised 17 May, 2006; accepted 19 May, 2006. *For correspondence. E-mail jagiron@email.arizona.edu; Tel. (+1) 520 626 0104; Fax (+1) 520 626 2100. Characterization of SP41, a surface protein of Brucella associated with adherence and invasion of host epithelial cells Elsa I. Castañeda-Roldán, 1 Safia Ouahrani-Bettache, 2 Zeus Saldaña, 3 Fabiola Avelino, 1 María A. Rendón, 3 Jacques Dornand 2 and Jorge A. Girón 3 * 1 Centro de Investigaciones en Ciencias Microbiológicas, Benemérita Universidad Autónoma de Puebla, Ciudad Universitaria, Puebla, 72570, México. 2 INSERM U431 Université de Montpellier II, 34075 Montpellier, France. 3 Department of Immunobiology, University of Arizona, Tucson, AZ 85724, USA. Summary Brucella is an invasive organism that multiplies and survives within eukaryotic cells. The brucellae are able to adhere to the surface of cultured epithelial cells, a mechanism that may facilitate penetration and dissemination to other host tissues. However, no adhesins that allow the bacteria to interact with the surface of epithelial cells before migration within polymorphonuclear leukocytes, monocytes and mac- rophages have been described. Here, we show that Brucella surface proteins (SPs) with apparent molec- ular masses of 14, 18 and 41 kDa bound selectively to HeLa cells. However, only antibodies directed against the 41 kDa surface protein (SP41) inhibited in dose– response manner, bacterial adherence and invasion of HeLa cells. HeLa cells treated with neuraminidase did not bind SP41, suggesting the involvement of cel- lular sialic acid residues in this interaction. Biochem- ical analysis of SP41 revealed that this protein is the predicted product of the ugpB locus, which showed significant homology to the glycerol-3-phosphate- binding ATP-binding cassette (ABC) transporter pro- tein found in several bacterial species. SP41 appears to be exposed on the bacterial surface as determined by immunofluorescence and immunogold labelling with anti-SP41 antibody. An isogenic DugpB mutant showed a significant inhibitory effect on Brucella adherence and invasion of human cultured epithelial cells and this effect could be reversed by restoration of the ugpB on a plasmid. Lastly, we also show that most of the sera from individuals with acute brucello- sis, but not sera obtained from healthy donors or patients with chronic brucellosis, mount antibody reactivity against SP41, suggesting that this protein is produced in vivo and that it elicits an antibody immune response. These data are novel findings that offer new insights into understanding the interplay between this bacterium and host target cells, and identify a new target for vaccine development and prevention of brucellosis. Introduction Adherence of pathogenic bacteria to host tissues is an essential and complex stage for bacterial colonization and consequently, for the establishment of bacterial infectious disease. Bacterial adherence is then considered an important virulence trait, because it enables bacterial pathogens to deliver toxins efficiently to host tissues, to interact closely with the cell membrane favouring intracel- lular penetration, to overcome peristaltic clearance and to establish microbial communities in biological niches. Bac- teria are able to selectively adhere and colonize host cells through the presence of surface adhesins that specifically recognize host cell receptors. Adhesins in the form of polymeric filamentous structures called pili or as outer membrane proteins may mediate direct binding of the bacteria to host cells or favour colonization by mediating bacteria–bacteria interactions. Brucella is a Gram-negative organism responsible for causing brucellosis and is classified as a select agent that could potentially be used as a bioweapon. The disease brucellosis is a highly common bacterial zoonosis world- wide, and an important cause of human disease and economic loss (Letesson et al., 2002). Brucellosis repre- sents a threat to farm animals and humans in which the bacteria are known to cause systemic infection due to the ability of the microorganism to traverse epithelial barriers, invade phagocytic and non-phagocytic cells, and localize to the reticulo endothelial system to cause infection (Boschiroli and O’Callaghan, 2001; Letesson et al., 2002; Kohler et al., 2003). In animals, brucellosis causes abor-