The structural gene for group A streptococcal pyrogenic exotoxin (SPE) type A was cloned into E. coli. DNA fragments used for cloning the toxin gene were isolated from bacteriophage T12. Toxin, present in cell lysates of E. coli clones, immunoprecipitated with antisera raised against purified SPE type A and formed a line of identity with streptococcal-derived A toxin. The cloned toxin shared the following biological activities with streptococcal A toxin: Pyrogenicity; enhancement of host susceptibility to lethal endotoxin shock; nonspecific lymphocyte mitogenicity; and alteration of immunoglobulin production. The physical location of the toxin gene on the phage T12 genome was determined.

A series of general preliminary studies was made on the physiology of delayed germination in Avena fatua, the results of which may be summarized as follows: (1) Great variations were found in the after-ripening periods of a number of A. fatua selections. (2) Evidence was obtained which strongly indicated that delayed germination is determined by a condition of the seed coat which develops after fertilization. (3) Results from tests of entire panicles indicated a correlation between germinability and the position of the seed in the panicle. (4) The after-ripening period of secondary grains was shown to be much longer than that of primary grains. (5) The placing of incompletely after-ripened grains under germinative conditions induced secondary dormancy. (6) Exposure to light appeared slightly to stimulate germination in seeds which were in the early stages of after-ripening, but appeared to have a harmful effect upon seeds which were more or less completely after-ripened. (7) Low dry-storage temperatures retarded the after-ripening process. Storage in a frozen condition at freezing temperatures resulted in increased germination. Seeds moistened and subjected to outdoor conditions failed to germinate. (8) Dormancy was more or less completely overcome by breaking the seed coat over the embryo, or by soaking seeds in potassium nitrate solutions. The exposure of seeds under germinative conditions to an atmosphere having an increased oxygen concentration definitely stimulated germination. Treatments with pure oxygen, ether, and sodium thiocyanate had more or less indifferent effects upon germination, while ethylene chlorhydrin and dichlorethylene were definitely injurious.It is inferred from the combined results that delayed germination is due to post-fertilization changes, related either to tissue absorption or development, which occur in the seed coats of A. fatua but not in readily germinable species, and which result in a restriction of the oxygen supply to the embryo. It is believed that the after-ripening process may consist, essentially, of a series of changes in the tissues of the seed coat which result in an increased permeability to oxygen.

Use of herbal therapies is not uncommon among diabetic patients. Many of the herbs reported have potential efficacy in treating diabetes or may result in adverse effects or interactions. In practical use, however, the herbs reported in this study are unlikely to have a significant effect on clinical outcomes in diabetes, either positively or negatively.

The nucleotide sequence of the gene encoding group A streptococcal pyrogenic exotoxin type A (SPE A) was determined by the dideoxy chain termination method. The first 30 residues of the translation product represented a hydrophobic signal peptide. The mature protein was 220 amino acids in length and had a molecular weight of 25,805. It has significant protein sequence homology with Staphylococcus aureus enterotoxin B but not with other proteins in the Dayhoff library.

Lysogenic conversion has been suggested as a mechanism of control of group A streptococcal pyrogenic exotoxin type A production. Digestion of DNA from two converting bacteriophages, 3GL16 and T12, with a variety of restriction endonucleases yielded identical DNA fragments upon electrophoresis in agarose gels. Several known A toxin-positive strains that did not appear to produce converting phage upon induction were analyzed for toxin and phage DNA. Strains, including NY5, 594, and C203S, were shown by hybridization studies to carry the A toxin gene (speA) adjacent to chromosomally inserted phage fragments, homologous to phage T12 DNA, which may represent defective converting phages. The phage T12 aft site mapped adjacent to speA. These data suggest that phage T12 acquired the A toxin gene from the bacterial genome. All streptococcal strains tested that were A toxin negative by Ouchterlony immunodiffusion failed to show any hybridization to speA-specific probes.Transfer of virulence factors among bacteria may contribute significantly to the generation of strains with increased pathogenicity. One such virulence factor produced by several group A streptococcal strains is streptococcal pyrogenic exotoxin type A (SPE A). SPE A is a low-molecular-weight toxin (molecular weight, 25,805) that induces scarlet fever and may play a role in the early events in delayed sequelae such as rheumatic fever.Previously, it was shown that expression of SPE A by strains of streptococci was regulated by bacteriophage. Zabriskie (28) first demonstrated that SPE A production could be transferred by bacteriophages T12 and 3GL16, obtained from donor streptococci, to a nontoxigenic bacterial strain designated T253. Subsequently, others have confirmed and extended this work. Johnson et al. (11) suggested that production of SPE A was transferrable to recipient host T253 by lysogenic conversion. Nida and Ferretti (17) indicated that transfer of the genetic determinant by bacteriophage was a general phenomenon. Further, it was shown that a clear-plaque mutant of phage T12 could cause expression of toxin during infection (15). Recently, Johnson and Schlievert (9) provided a restriction endonuclease map of phage T12 and demonstrated that the structural gene for SPE A (speA) is carried on the phage genome (10). Weeks and Ferretti (27) (9). Other strains previously reported to be positive for SPE A included the following: T28 and H44AA (7), obtained from D. W. Watson, and GT9094, GT70.1500, GT8627, and GT9316 (17), provided by L. W. Wannamaker, University of Minnesota.Three group A streptococcal strains which do not make SPE A were used as indicator (recipient) strains. These strains included the following: T253c (4), provided by A. E. Colon-Whitt and R. M. Cole, Laboratory of Streptococcal Diseases, National Institute of Allery and Infectious Diseases, Bethesda, Md.; K56, provided by P. P. Cleary, University of Minnesota; and T18P, provided by D. W. Watson.Escherichia coli K-12 RR1 (F-hsdS20 ara-14 proA2 lac YJ galK2 rpsL20 xyl-5 mtl-l supE44 ...