U.S. Department of Energy

Pacific Northwest National Laboratory

Discovery of novel secreted virulence factors from Salmonella enterica serovar Typhimurium by proteomic analysis of culture supernatants.

TitleDiscovery of novel secreted virulence factors from Salmonella enterica serovar Typhimurium by proteomic analysis of culture supernatants.
Publication TypeJournal Article
Year of Publication2011
AuthorsNiemann GS, Brown RN, Gustin JK, Stufkens A, Shaikh-Kidwai AS, Li J, McDermott JE, Brewer HM, Schepmoes A, Smith RD, Adkins JN, Heffron F
JournalInfect Immun
KeywordsAlgorithms, Animals, Artificial Intelligence, Bacterial Proteins, Gene Expression Regulation, Bacterial, Mice, Mice, 129 Strain, Mutation, Salmonella Infections, Animal, Salmonella typhimurium, Virulence Factors
Abstract

Salmonella enterica serovar Typhimurium is a leading cause of acute gastroenteritis throughout the world. This pathogen has two type III secretion systems (TTSS) encoded in Salmonella pathogenicity islands 1 and 2 (SPI-1 and SPI-2) that deliver virulence factors (effectors) to the host cell cytoplasm and are required for virulence. While many effectors have been identified and at least partially characterized, the full repertoire of effectors has not been catalogued. In this proteomic study, we identified effector proteins secreted into defined minimal medium designed to induce expression of the SPI-2 TTSS and its effectors. We compared the secretomes of the parent strain to those of strains missing essential (ssaK::cat) or regulatory (ΔssaL) components of the SPI-2 TTSS. We identified 20 known SPI-2 effectors. Excluding the translocon components SseBCD, all SPI-2 effectors were biased for identification in the ΔssaL mutant, substantiating the regulatory role of SsaL in TTS. To identify novel effector proteins, we coupled our secretome data with a machine learning algorithm (SIEVE, SVM-based identification and evaluation of virulence effectors) and selected 12 candidate proteins for further characterization. Using CyaA' reporter fusions, we identified six novel type III effectors and two additional proteins that were secreted into J774 macrophages independently of a TTSS. To assess their roles in virulence, we constructed nonpolar deletions and performed a competitive index analysis from intraperitoneally infected 129/SvJ mice. Six mutants were significantly attenuated for spleen colonization. Our results also suggest that non-type III secretion mechanisms are required for full Salmonella virulence.

DOI10.1128/IAI.00771-10
Alternate JournalInfect. Immun.
PubMed ID20974834
PubMed Central IDPMC3019877
Grant ListA1022933-22A1 / / PHS HHS / United States
R01 AI022933 / AI / NIAID NIH HHS / United States
Y1-A1-8401-01 / / PHS HHS / United States
| Pacific Northwest National Laboratory