title: Evaluation of an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of Chlamydia trachomatis infections.
abstract: BACKGROUND
  The OmcB protein is one of the most immunogenic proteins in C. trachomatis and C. pneumoniae infections. This protein is highly conserved leading to serum cross reactivity between the various chlamydial species. Since previous studies based on recombinant proteins failed to identify a species specific immune response against the OmcB protein, this study evaluated an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of C. trachomatis infections.
  
  
  RESULTS
  Using the ClustalW and Antigenic programs, we have selected two predicted specific and immunogenic regions in the OmcB protein: the N-terminal (Nt) region containing three epitopes and the C-terminal (Ct) region containing two epitopes with high scores. These regions were cloned into the PinPoint Xa-1 and pGEX-6P-1 expression vectors, incorporating a biotin purification tag and a glutathione-S-transferase tag, respectively. These regions were then expressed in E. coli. Only the pGEX-6P-1 has been found suitable for serological studies as its tag showed less cross reactivity with human sera and was retained for the evaluation of the selected antigens. Only the Ct region of the protein has been found to be well expressed in E. coli and was evaluated for its ability to be recognized by human sera. 384 sera were tested for the presence of IgG antibodies to C. trachomatis by our in house microimmunofluorescence (MIF) and the developed ELISA test. Using the MIF as the reference method, the developed OmcB Ct ELISA has a high specificity (94.3%) but a low sensitivity (23.9). Our results indicate that the use of the sequence alignment tool might be useful for identifying specific regions in an immunodominant antigen. However, the two epitopes, located in the selected Ct region, of the 24 predicted in the full length OmcB protein account for approximately 25% of the serological response detected by MIF, which limits the use of the developed ELISA test when screening C. trachomatis infections.
  
  
  CONCLUSIONS
  The developed ELISA test might be used as a confirmatory test to assess the specificity of serological results found by MIF.
authors.affiliation: Department of Microbiology and research laboratory Microorganismes et Pathologie Humaine, Habib Bourguiba hospital of Sfax, Tunisia. olfafrikhagargouri@ymail.com
authors.forename: Olfa
authors.initials: O
authors.lastname: Frikha-Gargouri
authors.forename: Radhouane
authors.initials: R
authors.lastname: Gdoura
authors.forename: Abir
authors.initials: A
authors.lastname: Znazen
authors.forename: Boutheina
authors.initials: B
authors.lastname: Gargouri
authors.forename: Jalel
authors.initials: J
authors.lastname: Gargouri
authors.forename: Ahmed
authors.initials: A
authors.lastname: Rebai
authors.forename: Adnene
authors.initials: A
authors.lastname: Hammami
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000097:blood; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000276:immunology*; D015153:Blotting, Western; D002648:Child; D002675:Child, Preschool; D002690:Chlamydia Infections / Q000175:diagnosis*; D002692:Chlamydia trachomatis / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D019295:Computational Biology; D004797:Enzyme-Linked Immunosorbent Assay / Q000592:standards; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D012698:Serologic Tests / Q000379:methods*
chemical_list: D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; C436968:OmcB protein, bacteria; D011994:Recombinant Proteins