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Title: SEQUENCING OF THE CHLAMYDOPHILA PSITTACI OMPA GENE REVEALS A NEW GENOTYPE, E/B, AND THE NEED FOR A RAPID DISCRIMINATORY GENOTYPING METHOD

Author
item GEENS, TOM - GHENT UNIVERSITY
item DESPLANQUES, ANN - GHENT UNIVERSITY
item VAN LOOCK, MARNIX - UNIVERSITEIT LEUVEN
item BONNER, B - JUSTUS-LIEBIG-UNIV
item KALETA, E - JUSTUS-LIEBIG-UNIV
item MAGNINO, S - NATL REF LAB ANIM CHLAMYD
item Andersen, Arthur
item EVERETT, KARIN - INST ENVIRONMENTAL HLTH
item VANROMPAY, D - GHENT UNIVERSITY

Submitted to: Journal of Clinical Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/20/2004
Publication Date: 5/20/2005
Citation: Geens, T., Desplanques, A., Van Loock, M., Bonner, B.M., Kaleta, E.F., Magnino, S., Andersen, A.A., Everett, K.D.E., Vanrompay, D. 2005. Sequencing of the Chlamydophila psittaci ompA gene reveals a new genotype, E/B, and the need for a rapid discriminatory genotyping method. Journal of Clinical Microbiology. 43(5):2456-2461.

Interpretive Summary: The currently used method of identifying chlamydial isolates, serotyping with monoclonal antibodies, was compared with two methods of genotyping isolates, PCR-RFLP and sequence analysis. These methods were used to identify 21 European chlamydial isolates from birds. The serotyping used a panel of 6 monoclonal antibodies to the known avian strains. The genotyping methods characterized the isolates by analysis of the genome of the major outer membrane protein. The serotyping method identified the strains of 19 of the 21 isolates, while both of the genotyping methods, PCR-RFLP and sequence analysis, differentiated all of the isolates. In addition, the genotyping was able to detect mixed infections missed by the serotyping, and the sequencing analysis indicated the existence of a new genotype that reacted with 2 of the monoclonal antibodies. The results show that genotyping methods have potential for improving epidemiological studies and stress the need to develop more rapid genotyping methods.

Technical Abstract: Avian Chlamydophila psittaci field isolates from 4 different European countries (Belgium, Germany, Italy and the Netherlands) were characterized using OmpA restriction fragment length and sequence analysis as well as serovar-specific monoclonal antibodies in an indirect immunofluorescence test. The serotyping panel was first characterized by isotyping. Western blotting and a radio-immunoprecipitation assay. All serovar-specific monoclonal antibodies recognized the major outer membrane protein of Chlamydophila psittaci. Only 19 on 21 isolates could be serotyped while all isolates were successfully genotyped using either OmpA RFLP or sequence analysis. In addition, genotyping revealed the presence of mixed infections in 5 on 21 isolates while serotyping could only detect one of these mixed infections. Genotypes A and B were preferentially associated with psittacines and pigeons, respectively. Interestingly, OmpA sequencing indicated the existence of a new genotype designated E/B, reacting with both serovar B- and E-specific monoclonal antibodies, having a genotype E restriction pattern. Results of the present study enabled the comparison between current available serotyping and genotyping methods for future epidemiological studies. OmpA RFLP analysis is preferred above the serotyping method as it is more sensitive. However, the genotype E/B can only be identified using OmpA sequence analysis, stressing the need for a more rapid discriminatory genotyping method.