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Afipia Felis

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Claus Koch – 1st expert on this subject based on the ideXlab platform

  • Immunopurified extracellular Bartonella henselae antigen for detecting specific antibodies by enzyme immunoassay.
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    Protein antigens of Bartonella henselae bacterial sonicate supernatant and concentrated cell-free culture filtrate were examined by SDS-PAGE. The sonicate supernatant gave 38 bands and the culture filtrate at least 21, of which 18 were of bacterial origin. Immunoblotting against 13 monoclonal antibodies obtained from mice infected with live B. henselae showed that 10 of these antibodies reacted with a narrow 225 kDa band and varying smears of bands ranging from 36 to 240 kDa in the sonicate, but only with a single 200 kDa band in the culture filtrate. Testing of pre- and post-infection rabbit sera in immunoblotting against culture filtrate demonstrated that the 200 kDa component gave the most prominent specific reaction with post-infection sera. The 200 kDa antigen was isolated by immunoaffinity chromatography of concentrated culture filtrate, and its molecular size determined by size-exclusion chromatography as >1000 kDa. The immunopurified antigen was compared with bacterial sonicate as coating antigen in EIA for determining humoral immune responses in rabbits inoculated with live B. henselae. The two antigens gave almost identical results for IgM and IgG responses. The specificity of the immunopurified antigen was tested in EIA against hyperimmune rabbit sera and sera of rabbits inoculated with live B. henselae, B. quintana and Afipia Felis. Only the hyperimmune serum against B. henselae and the sera of the rabbits inoculated with live B. henselae reacted with the immunopurified antigen, whereas the B. henselae sonicate cross-reacted with hyperimmune and post-infection sera of rabbits inoculated with B. quintana and A. Felis.

  • production and characterization of mouse monoclonal antibodies against Afipia Felis
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    : A series of 10 monoclonal antibodies reacting with Afipia Felis antigens were selected from mice immunized with live organisms of the reference strain ATCC 53690. Immunoblotting against SDS-PAGE-separated A Felis sonicate allowed the antibodies to be classified into three groups: 1) 168-4, -6, -7 and -10 reacted with a 53 kDa antigen, 2) 168-1, -3 and -9 reacted with both 53 kDa and 60 kDa antigens, and 3) 168-2, -5 and -9 reacted with other antigens. Antibodies of group 1 did not cross-react with other Afipia species or 36 unrelated bacteria, whereas those of groups 2 and 3 reacted with other Afipia species and some unrelated bacteria. Immunoblots of crossed immunoelectrophoretic patterns of A. Felis sonicate against rabbit antiserum showed that antibodies of groups 1 and 2 bound to the same precipitin arcs. Antibodies of group 1 reacted with a species-specific epitope on the 53 kDa antigen, while those of group 2 reacted with other epitopes shared by the 53 kDa and 60 kDa antigens. The binding of antibodies of group 1 to A. Felis sonicate was inhibited by post-infection rabbit serum, whereas no inhibition was observed for antibodies of group 2. The species-specific epitope of the 53 kDa antigen and the early appearance of antibodies against this epitope after infection suggest that this antigen can be used in a serodiagnostic test for A. Felis infection.

  • identification of Afipia Felis antigens in culture medium reaction with human sera
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    : Fourteen protein antigens were identified on SDS-PAGE of Afipia Felis culture supernatant. Immunoblotting against 10 monoclonal antibodies obtained from mice infected with live A. Felis showed that 4 antibodies reacted with a 56 kDa band and 3 with both 56 kDa and 62 kDa bands. Compared with A. Felis sonicate, the reacting proteins in culture supernatant showed an increase in molecular mass of 2-3 kDa, suggesting that they were more glycosylated. Purified antigen obtained by affinity chromatography of culture supernatant on the seven immobilized antibodies was tested against antibodies reacting with the 56 kDa and 62 kDa bands. All eluates contained both components, suggesting that the antibodies were directed against different epitopes of a double antigen held together during the affinity chromatography but cleaved by reduction and SDS-PAGE. The molecular size of the uncleaved protein in culture supernatant was determined by size-exclusion chromatography as > 1000 kDa. Testing of pre- and post-infection rabbit sera in immunoblotting against culture supernatant demonstrated that the 56 kDa and 62 kDa components gave the most prominent specific reactions with post-infection sera. One of fifty human sera submitted for testing for cat-scratch disease and 1 of 50 sera from healthy blood donors reacted with several bands in A. Felis culture supernatant, including the 56 kDa and 62 kDa bands.

Didier Raoult – 2nd expert on this subject based on the ideXlab platform

  • description of Afipia birgiae sp nov and Afipia massiliensis sp nov and recognition of Afipia Felis genospecies a
    International Journal of Systematic and Evolutionary Microbiology, 2002
    Co-Authors: Bernard La Scola, Marienoelle Mallet, Patrick A D Grimont, Didier Raoult

    Abstract:

    On the basis of phenotypic characterization and DNA relatedness, two novel species are proposed, Afipia birgiae sp. nov. (type strain 34632T = CIP 106344T = CCUG 43108T) and Afipia massiliensis sp. nov. (type strain 34633T = CIP 107022T = CCUG 45153T). A new genospecies is described, named Afipia Felis genospecies A, closely related to Afipia Felis. The complexity encountered in the taxonomy of the Bradyrhizobiaceae group within the alpha-2 subgroup of the Proteobacteria is discussed and the description of these novel species highlights the need for new tools for phylogenetic analysis in the group. The novel species herein described are fastidious bacteria isolated from a hospital water supply in co-culture with amoebae. It is hypothesized that this group of bacteria are a potential cause of nosocomial infections.

  • Legionella-like and other amoebal pathogens as agents of community-acquired pneumonia.
    Emerging Infectious Diseases, 2001
    Co-Authors: Thomas J. Marrie, Didier Raoult, Bernard La Scola, Richard J. Birtles, Elena De Carolis

    Abstract:

    We tested serum specimens from three groups of patients with pneumonia by indirect immunofluorescence against Legionella-like amoebal pathogens (LLAPs) 1–7, 9, 10, 12, 13; Parachlamydia acanthamoeba strains BN 9 and Hall’s coccus; and Afipia Felis. We found that LLAPs play a role (albeit an infrequent one) in community-acquired pneumonia, usually as a co-pathogen but sometimes as the sole identified pathogen. A number of bacteria that grow only within amoebae and are closely related phylogenetically to Legionella species, Legionella -like amoebal pathogens (LLAPs), have been identified and characterized (1). The role of these bacteria as human pathogens is still largely unknown. Other microorganisms, e.g., Parachlamydia acanthamoeba strains BN 9 (2) and Hall’s coccus (3), also grow within amoebae. Afipia Felis (once thought to be the etiologic agent of cat-scratch disease), a gram-negative rod, is difficult to grow on artificial medium but grows well in human monocytes and HeLa cells (4); this organism was recently reported to be an environmental bacterium probably associated with free-living amoebae and living in water (5). We tested serum specimens from three groups of patients with pneumonia to determine if any of these microorganisms cause disease. The Study We used 511 specimens from a 1985 study of a random sample of the Nova Scotia population (6); 121 acute- and convalescent-phase serum specimens from a study (Nova Scotia, 1991-1994) of 149 ambulatory patients with communityacquired pneumonia (7); and specimens from a prospective study of community-acquired pneumonia requiring hospitalization conducted at 15 teaching hospitals in eight Canadian provinces (1996-1997). All serum specimens from both groups of patients with pneumonia were tested for antibodies to Mycoplasma pneumoniae; influenza viruses A and B; parainfluenza viruses 1,2,3; adenovirus; and Respiratory syncytial virus (RSV) by a standard complement fixation technique in microtiter plates. Serum specimens from 60% of the patients (randomly selected from the group of patients with community-acquired pneumonia requiring hospitalization) were tested by the microimmunofluorescence test (8-10) for immunoglobulin (Ig) G and IgM antibodies to Chlamydia pneumoniae (AR 39 strain); C. psittaci (avian strain 6BC, feline pneumonitis strain FP, turkey strain TT 3, and pigeon strain CP 3); C. pecorum (ovine polyarthritis strain); and C. trachomatis

  • isolation of new fastidious α proteobacteria and Afipia Felis from hospital water supplies by direct plating and amoebal co culture procedures
    FEMS Microbiology Ecology, 2000
    Co-Authors: Bernard La Scola, Lina Barrassi, Didier Raoult

    Abstract:

    As water is a source of nosocomial infections in hospitals, the presence of fastidious Gram-negative bacteria in water samples taken in a university hospital was investigated. Water samples were inoculated onto agar plates and into amoebal microplates for co-culture. Sixty-eight α proteobacteria isolates were obtained and characterized using phenotypic methods and 16S rRNA gene sequence comparison. The latter approach divided the strains into seven clusters. Of these, one corresponded to previously recognized Afipia Felis and it is likely that six were closely related new species. As these bacteria are fastidious and can not be cultivated on standard microbiological media, their possible role in hospital-acquired human infections should be investigated.

Kraesten Engbaek – 3rd expert on this subject based on the ideXlab platform

  • Immunopurified extracellular Bartonella henselae antigen for detecting specific antibodies by enzyme immunoassay.
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    Protein antigens of Bartonella henselae bacterial sonicate supernatant and concentrated cell-free culture filtrate were examined by SDS-PAGE. The sonicate supernatant gave 38 bands and the culture filtrate at least 21, of which 18 were of bacterial origin. Immunoblotting against 13 monoclonal antibodies obtained from mice infected with live B. henselae showed that 10 of these antibodies reacted with a narrow 225 kDa band and varying smears of bands ranging from 36 to 240 kDa in the sonicate, but only with a single 200 kDa band in the culture filtrate. Testing of pre- and post-infection rabbit sera in immunoblotting against culture filtrate demonstrated that the 200 kDa component gave the most prominent specific reaction with post-infection sera. The 200 kDa antigen was isolated by immunoaffinity chromatography of concentrated culture filtrate, and its molecular size determined by size-exclusion chromatography as >1000 kDa. The immunopurified antigen was compared with bacterial sonicate as coating antigen in EIA for determining humoral immune responses in rabbits inoculated with live B. henselae. The two antigens gave almost identical results for IgM and IgG responses. The specificity of the immunopurified antigen was tested in EIA against hyperimmune rabbit sera and sera of rabbits inoculated with live B. henselae, B. quintana and Afipia Felis. Only the hyperimmune serum against B. henselae and the sera of the rabbits inoculated with live B. henselae reacted with the immunopurified antigen, whereas the B. henselae sonicate cross-reacted with hyperimmune and post-infection sera of rabbits inoculated with B. quintana and A. Felis.

  • production and characterization of mouse monoclonal antibodies against Afipia Felis
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    : A series of 10 monoclonal antibodies reacting with Afipia Felis antigens were selected from mice immunized with live organisms of the reference strain ATCC 53690. Immunoblotting against SDS-PAGE-separated A Felis sonicate allowed the antibodies to be classified into three groups: 1) 168-4, -6, -7 and -10 reacted with a 53 kDa antigen, 2) 168-1, -3 and -9 reacted with both 53 kDa and 60 kDa antigens, and 3) 168-2, -5 and -9 reacted with other antigens. Antibodies of group 1 did not cross-react with other Afipia species or 36 unrelated bacteria, whereas those of groups 2 and 3 reacted with other Afipia species and some unrelated bacteria. Immunoblots of crossed immunoelectrophoretic patterns of A. Felis sonicate against rabbit antiserum showed that antibodies of groups 1 and 2 bound to the same precipitin arcs. Antibodies of group 1 reacted with a species-specific epitope on the 53 kDa antigen, while those of group 2 reacted with other epitopes shared by the 53 kDa and 60 kDa antigens. The binding of antibodies of group 1 to A. Felis sonicate was inhibited by post-infection rabbit serum, whereas no inhibition was observed for antibodies of group 2. The species-specific epitope of the 53 kDa antigen and the early appearance of antibodies against this epitope after infection suggest that this antigen can be used in a serodiagnostic test for A. Felis infection.

  • identification of Afipia Felis antigens in culture medium reaction with human sera
    Apmis, 1997
    Co-Authors: Kraesten Engbaek, Lars Otto Uttenthal, Claus Koch

    Abstract:

    : Fourteen protein antigens were identified on SDS-PAGE of Afipia Felis culture supernatant. Immunoblotting against 10 monoclonal antibodies obtained from mice infected with live A. Felis showed that 4 antibodies reacted with a 56 kDa band and 3 with both 56 kDa and 62 kDa bands. Compared with A. Felis sonicate, the reacting proteins in culture supernatant showed an increase in molecular mass of 2-3 kDa, suggesting that they were more glycosylated. Purified antigen obtained by affinity chromatography of culture supernatant on the seven immobilized antibodies was tested against antibodies reacting with the 56 kDa and 62 kDa bands. All eluates contained both components, suggesting that the antibodies were directed against different epitopes of a double antigen held together during the affinity chromatography but cleaved by reduction and SDS-PAGE. The molecular size of the uncleaved protein in culture supernatant was determined by size-exclusion chromatography as > 1000 kDa. Testing of pre- and post-infection rabbit sera in immunoblotting against culture supernatant demonstrated that the 56 kDa and 62 kDa components gave the most prominent specific reactions with post-infection sera. One of fifty human sera submitted for testing for cat-scratch disease and 1 of 50 sera from healthy blood donors reacted with several bands in A. Felis culture supernatant, including the 56 kDa and 62 kDa bands.