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| Wolf-J„ckel, G.A.; J„ckel, C.; Museux, K.; Hoelzle, K.; Tasker, S.; Lutz, H.; Hofmann-Lehmann, R. | |
| Identification, characterization, and application of a recombinant antigen for the serological investigation of feline hemotropic Mycoplasma infections | |
| 2010 Clinical and Vaccine Immunology (17): 1917-1925 | |
In felids, three hemotropic mycoplasma species (hemoplasmas) have been described: _Mycoplasma haemofelis_, "_Candidatus _Mycoplasma haemominutum," and "_Candidatus _Mycoplasma turicensis." In particular, _M. haemofelis _may cause severe, potentially life-threatening hemolytic anemia. No routine serological assays for feline hemoplasma infections are available. Thus, the goal of our project was to identify and characterize an _M. haemofelis _antigen (DnaK) that subsequently could be applied as a recombinant antigen in a serological assay. The gene sequence of this protein was determined using consensus primers and blood samples from two naturally _M. haemofelis-_infected Swiss pet cats, an experimentally _M. haemofelis-_infected specific-pathogen-free cat, and a naturally _M. haemofelis-_infected Iberian lynx (_Lynx pardinus_). The _M. haemofelis _DnaK gene sequence showed the highest identity to an analogous protein of a porcine hemoplasma (72%). _M. haemofelis _DnaK was expressed recombinantly in an _Escherichia coli _DnaK knockout strain and purified using Ni affinity, sizeexclusion, and anion-exchange chromatography. It then was biochemically and functionally characterized and showed characteristics typical for DnaKs (secondary structure profile, thermal denaturation, ATPase activity, and DnaK complementation). Moreover, its immunogenicity was assessed using serum samples from experimentally hemoplasma-infected cats. In Western blotting or enzyme-linked immunosorbent assays, it was recognized by sera from cats infected with _M. haemofelis_, "_Ca. _Mycoplasma haemominutum," and "_Ca. _Mycoplasma turicensis," respectively, but not from uninfected cats. This is the first description of a full-length purified recombinant feline hemoplasma antigen that can readily be applied in future pathogenesis studies and may have potential for application in a diagnostic serological test. |
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