title: Use of in vivo-induced antigen technology (IVIAT) to identify genes uniquely expressed during human infection with Vibrio cholerae.
abstract: In vivo-induced antigen technology is a method to identify proteins expressed by pathogenic bacteria during human infection. Sera from 10 patients convalescing from cholera infection in Bangladesh were pooled, adsorbed against in vitro-grown El Tor Vibrio cholerae O1, and used to probe a genomic expression library in Escherichia coli constructed from El Tor V. cholerae O1 strain N16961. We identified 38 positive clones in the screen, encoding pili (PilA and TcpA), cell membrane proteins (PilQ, MshO, MshP, and CapK), methyl-accepting chemotaxis proteins, chemotaxis and motility proteins (CheA and CheR), a quorum-sensing protein (LuxP), and four hypothetical proteins. Analysis of immune responses to purified PilA and TcpA in individual patients demonstrated that the majority seroconverted to these proteins, confirming results with pooled sera. These results suggest that PilA and its outer membrane secretin, PilQ, are expressed during human infection and may be involved in colonization of the gastrointestinal tract. These results also demonstrate substantial immune responses to TcpA in patients infected with El Tor V. cholerae O1. In vivo-induced antigen technology provides a simple method for identifying microbial proteins expressed during human infection, but not during in vitro growth.
authors.affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA.
authors.forename: Long
authors.initials: L
authors.lastname: Hang
authors.forename: Manohar
authors.initials: M
authors.lastname: John
authors.forename: Muhammad
authors.initials: M
authors.lastname: Asaduzzaman
authors.forename: Emily Anna
authors.initials: EA
authors.lastname: Bridges
authors.forename: Cecily
authors.initials: C
authors.lastname: Vanderspurt
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Kirn
authors.forename: Ronald K
authors.initials: RK
authors.lastname: Taylor
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Hillman
authors.forename: Ann
authors.initials: A
authors.lastname: Progulske-Fox
authors.forename: Martin
authors.initials: M
authors.lastname: Handfield
authors.forename: Edward T
authors.initials: ET
authors.lastname: Ryan
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Calderwood
mesh_terms: D000907:Antibodies, Bacterial / Q000096:biosynthesis / Q000276:immunology; D000942:Antigens, Bacterial / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D002771:Cholera / Q000235:genetics / Q000276:immunology / Q000382:microbiology*; D003289:Convalescence; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D036781:Fimbriae Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D015723:Gene Library; D006801:Humans; D007163:Immunosorbent Techniques; D014734:Vibrio cholerae / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C085807:TcpA protein, Vibrio cholerae; C084295:pilQ protein, bacteria; D036781:Fimbriae Proteins