Graduate Thesis Or Dissertation
 

Serological identification of common and strain-specific antigens in Agrobacterium

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  • Two new methods, one of broad specificity and the other of narrow specificity, were developed for the identification of agrobacteria. The former method was based on ribosomal antigens that were common to members of the closely related Agrobacterium and Rhizobium genera. Antisera to unwashed 50 S ribosomal subunits from five Agrobacterium strains were tested in gel immunodiffusion against ammonium sulfate-washed ribosomes from 34 Agrobacterium, one Rhizobium meliloti and three Rhizobium leguminosarum strains, and nine other genera outside the Rhizobiaceae. Only the agrobacteria and rhizobia reacted to form a single band of precipitation common to each of the strains tested. However, when the same antisera were reacted against unwashed ribosome preparations from the same strains, heterogeneous precipitation patterns with multiple bands were produced. The heterogeneous patterns were due to heat-stable cell surface antigens contaminating the unwashed ribosomes. We hypothesized that the heterogeneous patterns represented antigenic differences among strains. Subsequently, antisera to unwashed ribosomes from six different Agrobacterium strains were tested against water-phenol extracts from 39 Agrobacterium strains; a precipitin band formed only with the homologous Agrobacterium strain. The strain-specific serological reaction was due to lipopolysaccharide antigens present as contaminants in the ribosomal preparation. The serological uniqueness of each strain was confirmed by EcoRI digestion profiles of total DNA. The profiles were different for each of the Agrobacterium strains. The utility of the broadly specific method was demonstrated when antisera to 50 S ribosomal subunits were used successfully to identify putative agrobacteria isolated from a natural habitat. In contrast, the strain specific method allowed the identification of Agrobacterium radiobacter K84 strains recovered six months after they were introduced into the environment.
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