L forms induced from two strains of group A Streptococcus were inoculated intraperitoneally and intravenously into conventional and germ-free mice. After intravenous injection, streptococcal L forms disappeared very quickly from the blood, whereas, after intraperitoneal injection, it was possible to isolate them as long as 25 days after inoculation. Observations suggest that a certain degree of reversion to a bacterial form may occur spontaneously in animals.

1 2 019 localization with no or very slight diffusion and also a simple and direct method for demonstration of this enzyme without the need for a coupling reaction.We are indebted to Mrs. Georgette M. Dunlalp for stenogriaphic assistance.

Human diploid cells in culture, infected with a balanced amount of living group A streptococci, were able to survive the infection and could be divided and propagated normally thereafter. The streptococci were rapidly phagocytized by the tissue culture cells. At the beginning, they kept their typical appearance, as well as their ability to fix dyes and group-specific immunoglobulins. After 1 to 2 days, the number of detectable streptococci decreased and they underwent important morphological changes. After some subsequent divisions of the cell line, streptococci persisted in cells only as large, isolated, swollen cocci, and no longer grew on suitable media. After six to eight divisions, a noticeable percentage of the tissue culture cells were very similar in appearance to the same cell line experimentally infected with “stable” L-variants. Cultures on L-phase media of supernatant fraction and cells, made 24 to 48 hr after inoculation, showed typical L-colonies. These grew well on media without antibiotics, as well as on media containing penicillin or vancomycin. They could be propagated on media with penicillin for months and were able to revert to group A streptococci after several subcultures on antibiotic-free media. Controls of uninoculated tissue culture cells never showed the presence of any microorganism. Group A streptococci inoculated into Eagle's basal medium, which was used for the tissue cultures, did not grow and never gave rise to L-colonies, even though the medium contained penicillin. Previous data suggest a biochemical explanation for this conversion, which otherwise is an occasional phenomenon.

The subunit composition of cell-internal and surface prosomes during phorbol myristate acetate (PMA)-induced differentiation of human leukemic T lymphocytes (CCRF-CEM cell line) was studied in relation to clusters of differentiation (CD) markers. PMA inhibited cell growth and decreased the amounts of CD1a and CD4 while CD3, CD8, CD25, CD45, CD57 and MHC1 increased it; the p53 anti-oncogene increased while actin levels remained constant. Cells incubated with the inducer PMA for 3 days and placed in fresh inhibitorfree medium resumed growth at a low rate, while the CD values slowly reverted to those of the initial phenotype. The presence and relative amounts of prosome subunits were analyzed by flow cytometry, light and fluorescent microscopy and Western blotting using 3 monoclonal antibodies (p25K, p27K and p30-33K MAbs). The decrease in cytoplasmic antigens on day 3 was remarkable (cells followed for 7 days) while increased surface antigens were observed. Changes in the subcellular distributions of prosome antigens, particularly the p25K and p30-33K subunit, were correlated with a partial arrest of the cell cycle. Interestingly, the composition of cell internal and surface prosomes showed different patterns of change. Int.

No abstract

DNAs isolated from four strains of Brucella bacteriophages were studied by restriction endonuclease mapping and Southern blot analysis. In all strains the genome was composed of a 38 kb (25.1 × 106 dalton) double‐stranded circular DNA. The physical map was the same for the four genomes and Southern blot hybridization of restriction endonuclease fragments with the Tbilissi strain DNA as a probe showed complete homology between the four DNAs. Thus, the four phage strains appear to be identical, the specific host range of each originating from minor changes in phage or Brucella receptors or both.

A simple fluorometric assay requiring only a single sample of cells to determine the number of cells, from the DNA linked to the fluorochrome 4',6-diamidino-2-phenylindol (DAPI), and the uptake of ciprofloxacin, a natural fluorescent quinolone is described. Mouse peritoneal macrophages were found to concentrate ciprofloxacin up to 12.7 (+/- 1.5)-fold. Combined fluorometry provides a precise, sensitive method for determining the intracellular concentration of fluoroquinolones, as well as that of naturally fluorescent or fluorochrome-linked drugs.

20S proteasomes (prosomes/multicatalytic proteinase) are protein particles built of 28 subunits in variable composition. We studied the changes in proteasome subunit composition during the differentiation of U937 cells induced by phorbol‐myristate‐acetate or retinoic acid plus 1,25‐dihydroxy‐cholecalciferol by western blot, flow cytometry and immuno‐fluorescence. p25K (C3), p27K (IOTA) and p30/33K (C2) subunits were detected in both the nucleus and cytoplasm of undifferentiated cells. Flow cytometry demonstrated a biphasic decrease in proteasome subunits detection during differentiation induced by RA+VD. PMA caused an early transient decrease in these subunits followed by a return to their control level, except for p30/33K, which remained low. Immuno‐fluorescence also showed differences in the cytolocalization of the subunits, with a particular decrease in antigen labeling in the nucleus of RA+VD‐induced cells, and a scattering in the cytoplasm and a reorganization in the nucleus of PMA‐induced cells. Small amounts of proteasomal proteins were seen on the outer membrane of non‐induced cells; these membrane proteins disappeared when treated with RA+VD, whereas some increased on PMA‐induced cells. The differential changes in the distribution and type of proteasomes in RA+VD and PMA‐induced cells indicate that, possibly, 20S proteasomes may play a role in relation to the mechanisms of differentiation and the inducer used.