Novel segregation patterns of infecting-mutant genotypes in plate complementation tests among amber mutants of bacteriophage BF23

Amber mutants of bacteriophage BF23 were classified into two functional groups, types I and II, by the yields of the infecting-mutant genotypes in plate complementation tests. Type I mutants produced their genotypes at levels more than 20% of the total progeny phages, and type II mutants did so at levels of less than 5%. Comparison of the results of plate complementation tests with those of extract complementation tests revealed that all the type I mutants were defective in the tail formation, while most type II mutants were defective in the formation of either mature heads (type IIa) or both mature heads and tails (type IIb). Since in extract complementation tests the activated phages are always of genotypes corresponding to mutations defective in only the tail formation, the plate complementation test is comparable with the extract complementation test when judged on the basis of the yield of the mutant genotypes. Of 29 complementation groups, 8 type I, 14 type IIa, and 5 type IIb mutants were identified. Previously, amber mutations of BF23 were mapped on four genetic segments. These segments were ordered in one linkage map by crosses between deletion and amber mutants.     

(6R)-Tetrahydrobiopterin Increases the Activity of Tryptophan Hydroxylase in Rat Raphe Slices

The effects of (6R)- and (6S)-tetrahydrobiopterin (BPH4), tetrahydroneopterin, and 6-methyltetrahydropterin on the activity of tryptophan hydroxylase were investigated in rat raphe slices. The activity of tryptophan hydroxylase was estimated by measurement of 5-hydroxytryptophan (5-HTP) formation under inhibition of aromatic L-amino acid decarboxylase with use of HPLC-fluorometric detection. (6R)-BPH4 (the naturally occurring form) at 42 microM, tetrahydroneopterin at 50 microM, and 6-methyltetrahydropterin at 100 microM increased tryptophan hydroxylase activity to 350, 145, and 146% of control values, respectively. (6S)-BPH4, however, had no significant effects on tryptophan hydroxylase activity. These results suggest that tryptophan hydroxylase is subsaturating in vivo for the naturally occurring cofactor, (6R)-BPH4, and that the concentration of (6R)-BPH4 may play an important role for the regulation of tryptophan hydroxylase activity in vivo.     

Gland formation induced in the allantoic and small-intestinal endoderm by the proventricular mesenchyme is not coupled with pepsinogen expression

Abstract. Allantoic and small-intestinal endoderms of chick and quail embryos were associated with the proventricular mesenchyme of chick embryos and then cultivated on chorioallantoic membrane. This resulted in the induction of complex glands, but the recombinates never produced embryo-specific pepsinogens; also, glandular cells developed a brush border, expressed sucrase antigen on their apical surface, and sometimes differentiated into goblet cells, thus indicating that both endoderms have the tendency to differentiate into an intestinal epithelium. In the recombinates composed of allantoic endoderm and proventricular mesenchyme, acid-protease activity was detected, but biochemical analysis revealed that this activity was not due topepsinogens. These results indicate that the gland formation induced in allantoic and small-intestinal endoderms by the proventricular mesenchyme is not accompanied by the expression of pepsinogens, suggesting that independent mechanisms are responsible for the morphogenesis and cyto chemical differentiation of the endoderm.     

Organization of the replication control region of plasmid ColIb-P9.

We identified a 1,845-base-pair sequence that contains essential information for the autonomous replication and regulation of the 93-kilobase-pair IncI alpha group ColIb-P9 plasmid. Biochemical and genetic analyses revealed that this sequence specifies at least two structural genes, designated repZ and inc. The repZ gene encodes a protein with a molecular weight of 39,000, which probably functions as an initiator for the ColIb-P9 replicon. The inc gene that phenotypically governs the incompatibility encodes an RNA with a size of about 70 bases. This small RNA acts in trans to repress the expression of repZ, thereby functioning to maintain a constant copy number of the ColIb-P9 replicon in host cells.     

Corelike structures in a stable L-form ofStreptococcus pyogenes

Corelike structures, which were interpreted as straight, large cylinders containing ribosome-like particles surrounded by an amorphous substance of low electron density, were found in a stable L-form ofStreptococcus pyogenes grown in the absence of antibiotics.     

Pumpkin (Cucurbita sp.) seed globulin I. Purification, characterization, and subunit structure

A heat stable globulin present in the cotyledons of pumpkinseeds was prepared as crystals which were soluble in a dilutesaline solution below pH 4.5 or in a solution with a high ionicstrength at neutral pHs. The protein was nearly homogeneousby ultracentrifuge analysis, and had a molecular weight of about112,000 daltons. Sodium dodecyl sulfate-polyacrylamide gel electrophoresisseparated the globulin into two subunits, and ß,corresponding to molecular weights of about 63,000 and 56,000daltons, respectively. By reduction of disulfide bonds, thetwo subunits were each separated into two polypeptide chainswith molecular weights of around 36,000 and 22,000 daltons,judged by gel electrophoresis. The amino acid composition ofwhole globulin indicated high contents of arginine, glutamicacid and aspartic acid. The total number of half-cystine residuewas nine and only one residue was shown to be free. The subunitstructure of the globulin is discussed. The protein has beenshown to have oxaloacetate decarboxylase activity, and thisfact was confirmed. However, the activity decreased markedlyat pH 4.5 in a fairly short period. It did not require Mn⁺⁺,and the K_m for oxaloacetate was determined to be 4.1 mM. (Received April 9, 1976; )     

Role of leader peptide synthesis in repZ gene expression of the ColIb-P9 plasmid

The frequency of replication initiation of the ColIb-P9 plasmid depends on the level of repZ expression, which has been shown to be negatively regulated by inc RNA, the approximately 70-base-long product of the inc gene. To further understand the regulatory mechanism of repZ gene expression, we isolated mutants defective in ColIb-P9 replication using a lambda:ColIb-P9 hybrid phage. Among six mutants isolated, one amber mutant, rep57, failed to synthesize the RepZ protein. The mutation occurred in the repZ leader sequence that encodes a 29-amino-acid reading frame, designated as repY. We also isolated mutants that suppressed the rep57 phenotype. These mutations were single base insertions between the repY initiation codon and the rep57 mutation site and resulted not only in a frame shift of repY but also in the formation of repY-repZ fusions without changing the amino acid sequence of RepZ. Thus, repY is not directly involved in the replication reaction but rather functions as a positive regulator for repZ expression. We propose that repZ expression is coupled with repY translation, which acts to disrupt a secondary structure sequestering the repZ translation initiation signal. The positive and negative regulations of repZ expression were discussed. The other mutants were mapped in repZ, confirming that repZ is essential for ColIb-P9 replication.     

Granulocyte colony-stimulating factor potentiates anti-Candida albicans growth inhibitory activity of polymorphonuclear cells

Abstract Granulocyte colony-stimulating factor (G-CSF) stimulates a subset of granulocyte colony forming cells and when administered to neutropenic individuals results in recovery of blood neutrophil numbers to normal levels. Therefore, G-CSF may be a useful therapeutic agent for infections in immunocompromised hosts. However, to date there has been only limited information that G-CSF activates the antimicrobial activity of neutrophils. In the present study, we found that recombinant G-CSF promotes the anti- Candida albicans activity of normal human blood polymorphonuclear (PMN) cells in vitro using both a ³H-glucose uptake procedure and a Candida colony counting assay. As little as 0.1 ng/ml G-CSF induced significant anti-Candida activity in the PMN cultures. G-CSF treatment also enhanced superoxide anion production by the PMNs in response to f-MLP as determined by the superoxide dismutase inhibitable cytochrome C reduction method. Such results show that G-CSF can promote the antimicrobial activity of peripheral blood PMNs against C. albicans .     

Comparison of the Virulence of Methicillin-Resistant and Methicillin-Sensitive Staphylococcus aureus

The virulence of methicillin-resistant Staphylococcus aureus (MRSA) was compared with that of methicillin-sensitive S. aureus (MSSA), using 13 MRSA and 7 MSSA strains isolated from clinical specimens. The infectivity and lethality of the two groups were examined as to the inoculum required to infect 50% of guinea pigs (ID₅₀) and to kill 50% of mice (LD₅₀), respectively. The mean ID₅₀ [log₁₀ colony forming units (CFU)] for MRSA strains was 7.1 ± 0.60 standard deviation, which was 1.5 higher than that for MSSA strains (P < 0.001). The mean LD₅₀ (log₁₀ CFU) for MRSA strains was 9.0 ± 0.42, being 1.1 higher than that for MSSA strains (P = 0.001). Pretreatment of mice with cyclophosphamide decreased the mean LD₅₀ for MRSA strains more than that for MSSA strains, resulting in the difference in the mean LD₅₀ being insignificant (P = 0.502). These results indicate that MRSA is less virulent than MSSA in normal hosts, but that they are equally virulent in immunocompromised hosts. The growth of MRSA strains was much slower than that of MSSA strains in the lag phase, although their growth rates were almost the same in the exponential growth phase, suggesting that the difference in virulence between them may be at least partly due to such a difference in growth.