Purification and partial characterization of the OmpA family of proteins of Pasteurella haemolytica

This study was conducted to partially characterize and identify the purity of two major outer membrane proteins (OMPs) (with molecular weights of 32,000 and 35,000 [32K and 35K, respectively]) of Pasteurella haemolytica. The 35K and 32K major OMPs, designated Pasteurella outer membrane proteins A and B (PomA and PomB, respectively), were extracted from P. haemolytica by solubilization in N-octyl polyoxyl ethylene. The P. haemolytica strain used was a mutant serotype A1 from which the genes expressing the 30-kDa lipoproteins had been deleted. PomA and PomB were separated and partially purified by anion-exchange chromatography. PomA but not PomB was heat modifiable. The N-terminal amino acid sequences of the two proteins were determined and compared with reported sequences of other known proteins. PomA had significant N-terminal sequence homology with the OmpA protein of Escherichia coli and related proteins from other gram-negative bacteria. Moreover, polyclonal antiserum raised against the E. coli OmpA protein reacted with this protein. PomA was surface exposed, was conserved among P. haemolytica biotype A serotypes, and had porin activity in planar bilayers. No homology between the N-terminal amino acid sequence of PomB and those of other known bacterial proteins was found. Cattle vaccinated with live P. haemolytica developed a significant increase in serum antibodies to partially purified PomA, as shown by enzyme-linked immunosorbent assays, and to purified PomA and PomB, as detected on Western blots and by densitometry.Peer reviewedAnatomy, Pathology and PharmacologyInfectious Disease and Physiolog

A Novel Secretion Pathway of Salmonella enterica Acts as an Antivirulence Modulator during Salmonellosis

Salmonella spp. are Gram-negative enteropathogenic bacteria that infect a variety of vertebrate hosts. Like any other living organism, protein secretion is a fundamental process essential for various aspects of Salmonella biology. Herein we report the identification and characterization of a horizontally acquired, autonomous and previously unreported secretion pathway. In Salmonella enterica serovar Typhimurium, this novel secretion pathway is encoded by STM1669 and STM1668, designated zirT and zirS, respectively. We show that ZirT is localized to the bacterial outer membrane, expected to adopt a compact β-barrel conformation, and functions as a translocator for ZirS. ZirS is an exoprotein, which is secreted into the extracellular environment in a ZirT-dependent manner. The ZirTS secretion pathway was found to share several important features with two-partner secretion (TPS) systems and members of the intimin/invasin family of adhesions. We show that zirTS expression is affected by zinc; and that in vivo, induction of zirT occurs distinctively in Salmonella colonizing the small intestine, but not in systemic sites. Additionally, strong expression of zirT takes place in Salmonella shed in fecal pellets during acute and persistent infections of mice. Inactivation of ZirTS results in a hypervirulence phenotype of Salmonella during oral infection of mice. Cumulatively, these results indicate that the ZirTS pathway plays a unique role as an antivirulence modulator during systemic disease and is involved in fine-tuning a host–pathogen balance during salmonellosis