ISOLATION OF AUXOTROPHS OF BACILLUS CEREUS.
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Biomedical subjects
Publications and source records attributed to C B THORNE.
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Gwinn, Darrel D. (Oregon State University, Corvallis), and Curtis B. Thorne. Transformation of Bacillus licheniformis. J. Bacteriol. 87:519-526. 1964.-When a series of 28 auxotrophic mutants of Bacillus licheniformis were screened for transformation, only three of them, M28 (glycine(-)), M30 (uncharacterized), and M33 (purine(-)), produced a detectable number of transformants. The screening method consisted of spreading auxotrophic cells and deoxyribonucleic acid (DNA) from the prototrophic strain 9945A on minimal agar plates and observing the plates for development of prototrophic colonies. M28 transformed at a higher frequency than did the two other mutants, and it was studied in greater detail. Although up to 20% of the recipient cells spread on the plates in the presence of DNA gave rise to prototrophic colonies over a period of 72 hr, only about 10(-3)% of the cells produced transformants when they were incubated with DNA in liquid suspension for 1 hr. The most competent cultures of many tested were those grown on a shaker for 22 hr in a medium composed of nutrient broth, salts, and glycerol. When mutations resulting in requirements for histidine, leucine, serine, and trytophan were introduced singly into the glycine mutant, transformants for the leucine, serine, and histidine markers could be obtained at will, but transformants for the tryptophan marker were not detected even though all four of the double mutants could be transformed to glycine independence.
Taylor, Martha J. (U.S. Army Biological Laboratories, Fort Detrick, Frederick, Md.) and Curtis B. Thorne. Transduction of Bacillus licheniformis and Bacillus subtilis by each of two phages. J. Bacteriol. 86:452-461. 1963.-A second transducing bacteriophage, designated SP-15, was isolated from the same soil-sample culture filtrate that supplied the Bacillus subtilis transducing phage, SP-10, reported earlier from this laboratory. SP-10 and SP-15 differ serologically and in several other respects, but share the ability to propagate on B. subtilis W-23-S(r) (streptomycin-resistant) and B. licheniformis ATCC 9945a, and to mediate general transduction in either species when propagated homologously. Attempts to transduce between the species have failed. SP-10 forms plaques readily on both W-23-S(r) and 9945a; SP-15 forms minute plaques on W-23-S(r) and has shown no evidence of any lytic activity on 9945a. Maximal recoveries of prototrophic colonies from mixtures of SP-10 with auxotrophs of either W-23-S(r) or 9945a were obtained only when excess phage was neutralized by post-transduction treatment with specific phage antiserum. Such treatment was not necessary for maximal recovery of transductants effected by SP-15. Unlike SP-10, SP-15 propagated on W-23-S(r) did not transduce B. subtilis 168 (indole(-)). SP-15 transduced B. licheniformis more efficiently than did SP-10. Neither phage was able to transduce B. licheniformis as efficiently as it transduced B. subtilis. The differing influences of multiplicity of infection were compared for the two phages in both species.
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Thorne, Curtis B. (Fort Detrick, Frederick, Md.). Transduction in Bacillus subtilis. J. Bacteriol. 83:106-111. 1962.-A bacteriophage, SP-10, isolated from soil carries out general transduction in Bacillus subtilis. Phage propagated on a streptomycin-resistant mutant of the wild-type strain W-23 was capable of transducing to prototrophy strain 168 (indole(-)), as well as all of the auxotrophic mutants of W-23-S(r) tested, which included mutants requiring arginine, histidine, adenine, guanine, thiamine, leucine, or methionine. Although strain 168 was transduced by phage SP-10, lytic activity on this strain could not be detected and attempts to propagate the phage on it failed. Transductions occurred at frequencies in the range of 10(-6) to 10(-5) per plaque-forming unit. Homologous phage was ineffective, deoxyribonuclease had no effect on the frequency of transduction, and transduction was prevented by the addition of phage antiserum. Phage SP-10 was capable of lysogenizing strain W-23-S(r), and this condition was maintained through repeated growth and sporulation cycles in potato-extract medium. Although heating at 65 C for 60 min inactivated free phage particles, spores retained their lysogenic condition after such heat treatment. When heat-treated spores of the lysogenic cultures were used as inocula for growth in a nutrient broth-yeast extract-glucose medium, filtrates contained 10(9), or more, phage particles per ml.
Beall, Francis A. (U.S. Army Chemical Corps, Frederick, Md.), Martha J. Taylor, and Curtis B. Thorne. Rapid lethal effect in rats of a third component found upon fractionating the toxin of Bacillus anthracis. J. Bacteriol. 83:1274-1280. 1962.-Rats were found to be more susceptible to the lethal effect of toxin produced by Bacillus anthracis in vitro than were several species considered less resistant to anthrax. Rats were killed much faster by less toxin per gram of body weight than were mice. Guinea pigs survived doses of toxin that killed rats. Intravenous injection of Fischer 344 rats is a rapid test for lethal activity, which facilitates the demonstration of two components, different from protective antigen, in toxin. One of these, a lethal factor, was separated from the other component, which causes cutaneous edema in the guinea pig. The latter component was not necessary for lethal effect. Neither of these factors was active unless combined with protective antigen. Although the guinea pig skin reaction has been used routinely to assay the toxicity of samples, the present results show that this test does not assay the lethal component.
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