Graduate Thesis Or Dissertation
 

Host range, serological and electron microscope studies of phages of Agrobacterium tumefaciens (Smith and Townsend) Conn

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  • Phages of crown gall bacteria were among the first to be isolated and certain properties of these phages have been studied by a number of investigators. However, several important characteristics have not been studied in detail. The present investigation was made in order to determine the major sources of phages of A. tumefaciens and to compare the host range pattern, morphology and serological relationships among phages from different sources. Thirty-one phage isolates were obtained from sewage, soil and crown gall tissues. Phages were prevalent in sewage, but scarce in soil and gall tissue. Prevalence of these phages in sewage, where A. tumefaciens is not expected to occur, poses some interesting questions regarding host range of the phages and possible relationships between crown gall bacteria and bacteria present in sewage. Plaques of the phages were always round and clear with a sharp margin, but the size varied. There appeared to be four different strains of phage based on plaque size. The most common plaque type was 1.0-2.5 mm in diameter. One isolate produced small plaques, 0.25-0.50 mm, and six isolates produced large ones, 3.5-5.0 mm. One isolate produced plaques which were 2.5-3.5 mm in diameter. The size of the plaques did not correlate with source of isolation. Forty-one strains of A. tumefaciens and a number of other bacteria associated with plants were tested for host specificity of the phage isolates. The phages were very specific as they lysed only certain strains of A. tumefaciens. Eight strains of A. tumefaciens and all the other bacteria tested including representatives of three other species of Agrobacterium were not lysed. Among the 31 phage isolates, there were 26 different strains based on host range patterns within A. tumefaciens. In some instances, the phages were closely related in host range, but most were entirely different. The source of isolation and plaque size of the phages did not correlate with the host range. All 31 phage isolates were used as antigens to stimulate production of antibodies in rabbits. Each of the antisera produced was tested against all of the phage isolates by the Ouchterlony agar double-diffusion method. The phages could be divided into five serological groups based on the reactions which occurred. The phages in each group contained at least two antigens, and all five groups were more or less interrelated. The host bacterium on which the phages were reproduced appeared to be correlated with serological properties of phage. Host specificity seemed to be related to serological properties in some cases but not in the others. Source of isolation and plaque size were not correlated with serological properties. Seven phage isolates with representatives from each source of isolation were observed with the electron microscope. All of the phage particles were similar in shape but were somewhat varied in size. The heads were polyhydral, 42 mμ in width and 80-92 mμ in length. The tails were 8 mμ in width and 90 -125 mμ in length. Phage morphology did not appear to be related to source of isolation, plaque size, host range or serological properties.
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