title: In Vivo-Expressed Proteins of Virulent Leptospira interrogans Serovar Autumnalis N2 Elicit Strong IgM Responses of Value in Conclusive Diagnosis.
abstract: Leptospirosis is a serious zoonosis that is underdiagnosed because of limited access to laboratory facilities in Southeast Asia, Central and South America, and Oceania. Timely diagnosis of locally distributed serovars of high virulence is crucial for successful care and outbreak management. Using pooled patient sera, an expression gene library of a virulent Leptospira interrogans serovar Autumnalis strain N2 isolated in South India was screened. The identified genes were characterized, and the purified recombinant proteins were used as antigens in IgM enzyme-linked immunosorbent assay (ELISA) either singly or in combination. Sera (n = 118) from cases of acute leptospirosis along with sera (n = 58) from healthy subjects were tested for reactivity with the identified proteins in an ELISA designed to detect specific IgM responses. We have identified nine immunoreactive proteins, ArgC, RecA, GlpF, FliD, TrmD, RplS, RnhB, Lp28.6, and Lrr44.9, which were found to be highly conserved among pathogenic leptospires. Apparently, the proteins ArgC, RecA, GlpF, FliD, TrmD, and Lrr44.9 are expressed during natural infection of the host and undetectable in in vitro cultures. Among all the recombinant proteins used as antigens in IgM ELISA, ArgC had the highest sensitivity and specificity, 89.8% and 95.5%, respectively, for the conclusive diagnosis of leptospirosis. The use of ArgC and RecA in combination for IgM ELISA increased the sensitivity and specificity to 95.7% and 94.9%, respectively. ArgC and RecA thus elicited specific IgM responses and were therefore effective in laboratory confirmation of Leptospira infection.
authors.affiliation: Medical Microbiology Laboratory, Department of Microbiology, Centre of Excellence in Life Sciences, Bharathidasan University, Tiruchirappalli, India.
authors.forename: Veerapandian
authors.initials: V
authors.lastname: Raja
authors.affiliation: Medical Microbiology Laboratory, Department of Microbiology, Centre of Excellence in Life Sciences, Bharathidasan University, Tiruchirappalli, India.
authors.forename: Santhanam
authors.initials: S
authors.lastname: Shanmughapriya
authors.affiliation: Medical Microbiology Laboratory, Department of Microbiology, Centre of Excellence in Life Sciences, Bharathidasan University, Tiruchirappalli, India.
authors.forename: Murugesan
authors.initials: M
authors.lastname: Kanagavel
authors.affiliation: Gluck Equine Research Center, University of Kentucky, Lexington, Kentucky, USA.
authors.forename: Sergey C
authors.initials: SC
authors.lastname: Artiushin
authors.affiliation: Gluck Equine Research Center, University of Kentucky, Lexington, Kentucky, USA.
authors.forename: Sridhar
authors.initials: S
authors.lastname: Velineni
authors.affiliation: Gluck Equine Research Center, University of Kentucky, Lexington, Kentucky, USA.
authors.forename: John F
authors.initials: JF
authors.lastname: Timoney
authors.affiliation: Medical Microbiology Laboratory, Department of Microbiology, Centre of Excellence in Life Sciences, Bharathidasan University, Tiruchirappalli, India natarajaseenivasan@rediffmail.com.
authors.forename: Kalimuthusamy
authors.initials: K
authors.lastname: Natarajaseenivasan
mesh_terms: D000445:Aldehyde Oxidoreductases / Q000276:immunology; D000907:Antibodies, Bacterial / Q000097:blood; D000942:Antigens, Bacterial / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000145:classification / Q000276:immunology* / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D015723:Gene Library; D016680:Genome, Bacterial; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood*; D007194:India; D041301:Leptospira interrogans serovar autumnalis / Q000737:chemistry* / Q000235:genetics / Q000276:immunology* / Q000472:pathogenicity; D007922:Leptospirosis / Q000175:diagnosis* / Q000276:immunology* / Q000382:microbiology; D008297:Male; D011938:Rec A Recombinases / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D007075:Immunoglobulin M; D011994:Recombinant Proteins; D000445:Aldehyde Oxidoreductases; C089846:argC protein, Bacteria; D011938:Rec A Recombinases