Development of a rapid immunodiagnostic test for Haemophilus ducreyi.
Haemophilus ducreyi is the etiologic agent of chancroid, a sexually transmitted disease that increases the rate of transmission of human immunodeficiency virus. Chancroid ulcerations are difficult to distinguish from those produced by syphilis and herpes. Diagnosis based solely on clinical grounds is inaccurate, and culture is insensitive. Highly sensitive PCR has largely superseded culture as the preferred method of laboratory diagnosis; however, neither culture nor PCR is feasible where chancroid is endemic. We developed a rapid (15-min) diagnostic test based on monoclonal antibodies (MAbs) to the hemoglobin receptor of H. ducreyi, HgbA. This outer membrane protein is conserved in all strains of H. ducreyi tested and is required for the establishment of experimental human infection. MAbs to HgbA were generated and tested for cross-reactivity against a panel of geographically diverse strains. Three MAbs were found to be unique and noncompetitive and bound to all strains of H. ducreyi tested. Using an immunochromatography format, we evaluated the sensitivity and specificity of the test using geographically diverse strains of H. ducreyi, other Haemophilus strains, and other bacteria known to superinfect genital ulcers. All H. ducreyi strains were positive, and all other bacteria were negative, resulting in a specificity of 100%. The minimum number of CFU of H. ducreyi detected was 2 x 10(6) CFU, and the minimum amount of purified HgbA protein detected was 8.5 ng. Although this level of sensitivity may not be sufficient to detect H. ducreyi in all clinical specimens, further work to increase the sensitivity could potentially make this a valuable bedside tool in areas where chancroid is endemic.
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000032:analysis* / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins; D002352:Carrier Proteins / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D002602:Chancroid / Q000382:microbiology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006191:Haemophilus ducreyi / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D006801:Humans; D007120:Immunochemistry; D007159:Immunologic Tests / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; C017206:hemoglobin-binding protein, bacteria