Pathogenic leptospires of serovar hardjo isolated from North American cattle were compared genetically and antigenically to reference strain hardjoprajitno of the Sejroe serogroup. Guanine-plus-cytosine (G+C) contents were determined for the genomes, and microscopic agglutination, Western blotting (immunoblotting), and immunoprecipitation were used to characterize antigenic relatedness. Major differences were demonstrated between the isolates and the reference strain. The G+C content of the reference strain was calculated to be approximately 34 1%, and those of the isolates were calculated at 39 + 1%. Antigenic differences between the isolates and the reference strain were identified by using rabbit immune serum raised against a hardjo isolate exhaustively adsorbed with hardjoprajitno whole and sonicated cells. Western blot analysis and immunoprecipitation using this adsorbed serum revealed antigens apparently unique for the hardjo isolates. Microscopic agglutination with the adsorbed rabbit serum did not agglutinate hardjoprajitno when diluted 1:2 but agglutinated bovine isolates to a 1:32 dilution. Bovine antiserum raised against the isolates was also used to identify antigens by immunoprecipitation.
Bacteriophages were isolated from 22 of 38 strains of Vibrio fetus by an enrichment process, utilizing the donor and host strains growing together in fluid thioglycollate medium. One phage, V-45, isolated by the conventional lawn-spot method, was characterized by stability in broth, growth kinetics, and morphology. It was sensitive to rapid thermal inactivation, chloroform, and pH values above 6.5. Calcium was required for phage replication and stability in broth. Magnesium provided the best protection against thermal inactivation at 50 C in the pH range of 6.5 to 7.5. The minimum latent period was 135 min, rise time was 75 min, and average burst size was 35 plaque-forming units per infected cell. Phage V-45 resembled Bradley's morphological group B, having a long tail without contractile sheath. Dimensions were: head, about 50 nm; tail, about 7 by 240 nm; and tail lumen, 2 to 3 nm.
We have developed a rapid method for the isolation of leptospiral chromosomal DNA which yields DNA of a purity suitable for restriction endonuclease analysis. A small volume (15 to 20 ml) of an exponentially growing culture of leptospires yielded 2 to 4 ,ug of chromosomal DNA. In a 1-day protocol, the DNA was isolated, restricted with endonucleases, and fractionated on an agarose gel. Chromosomal DNA from dinger zones (visible subsurface zones of leptospiral growth) of first semisolid subcultures of field isolates was also isolated and characterized, thus greatly speeding up the diagnostic process.
SUMMARY. Auto-agglutinated and non-agglutinated cells of Campylobacter jejuni and C. coli were examined by transmission electronmicroscopy in phosphotungstate negative stain. Agglutination was induced by three factors (I) extracellular DNA, (2) an aggregated protein, probably a bacteriophage precursor, and (3) free phage-tail sheaths. Auto-agglutinated cells were often "leaky," with a mantle of adhering DNA. About 80% of the auto-agglutinated cells could be resuspended after treatment with DNAase. Flagella were loosely embedded in protein aggregates, especially in phage-infected cultures. They were clumped in a side-by-side arrangement by free phage-tail sheaths. These findings suggest that auto-agglutination could be minimised in suspensions of organisms intended for use in agglutination tests by harvesting early logarithmic-phase cells containing no more than a low phage population. The most common C. jejuni phage had a contractile tail, a head diameter of 60-70 nm, and an overall length of 180-2 10 nm. A phage isolated from C. jejuni strain 1590 was morphologically identical with C. coli phage.
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