Maternal origin of a unique extra chromosome, der(9)(pter-->q13::q13-->q12:) in a girl with typical trisomy 9p syndrome.

We report on a girl with the typical trisomy 9p syndrome who had an additional E-sized metacentric chromosome. On the basis of GTG- and CBG-banding, her karyotype was considered to be 47,XX,+der(9)(pter-->q13::q13-->q12:) de novo. Results of a fluorescence in situ hybridization study using a chromosome 9-specific painting probe were compatible with this cytogenetic interpretation. Molecular analyses of six highly polymorphic dinucleotide repeat loci on the short arm and the proximal long arm of chromosome 9 demonstrated that the girl inherited one allele from her father and two identical or different alleles from the mother. We speculated that the extra chromosome may have resulted from either nondisjunction of chromosome 9 followed by a U-type exchange and a crossing-over between different sister chromatids during maternal meiosis I and subsequent breakage and malsegregation during meiosis II, or nondisjunction during meiosis II followed by isochromosome formation in one of the two maternal chromosomes 9 and subsequent breakage.

Effect of AD-5423 on animal models of schizophrenia: phencyclidine-induced behavioral changes in mice.

The antipsychotic efficacy of AD-5423, which has the properties of both a serotonin 5-HT(2) and a dopamine D(2) receptor antagonist, was evaluated using animal models of schizophrenia. Sensitization to phencyclidine (PCP)-induced hyperlocomotion is considered a model of the positive symptoms of schizophrenia, and was significantly antagonized by AD-5423 and haloperidol. The PCP-induced enhancement of immobility induced by the forced swimming test, a model of the negative symptoms of schizophrenia, was attenuated by AD-5423 but not by haloperidol. Since this attenuated effect of AD-5423 was antagonized by DOI, a serotonin 5-HT(2) receptor agonist, it is postulated to be mediated by serotonin 5-HT(2) receptors. These findings suggest that AD-5423 would be clinically effective against both the positive and negative symptoms of schizophrenia.

Purification and some properties of a hemolysin from the poisonous mushroom Rhodophyllus rhodopolius.

A hemolytic protein which causes diarrhea and death to mice was purified from the fruit bodies of a poisonous mushroom species Rhodophyllus rhodopolius (Fries) by DEAE-cellulose column chromatography, ammonium sulfate precipitation, gel filtration on Sephadex G-200, and DEAE-Sephadex A-25 column chromatography. The mol. wt of the purified hemolysin was estimated to be 40,000 by SDS-polyacrylamide slab gel electrophoresis. The hemolytic activity of the purified hemolysin was destroyed by heating at 60 degrees C for 10 min, and partially reduced by pepsin, papain and 2-mercaptoethanol. Cholesterol and phosphatidylcholine did not inhibit the activity. The hemolysin was unstable below pH 7.0 but stable at pH 8.0. The optimal pH for hemolysis was 6.0. Hemolysis did not occur below 4 degrees C even though the hemolysin bound to the erythrocyte. Mouse, chicken, rat, horse and human erythrocytes were sensitive in this order, but sheep and cow erythrocytes were not lysed by the toxin.

Studies on the pathogenicity of Yersinia enterocolitica. I. Experimental infection in rabbits.

Analysis of the pathogenicity of Yersinia enterocolitica was performed with an experimental model successfully produced in rabbits by intraduodenal inoculation with strains isolated from various sources. Pathogenic strains easily penetrated the epithelial linings of the intestinal mucous membrane into the target reticuloendothelial tissues of the intestine, such as the lamina propria and lymph follicles, where they multiplied within mononuclear cells and produced granuloma. Granuloma, in severe infections, underwent necrobiosis and sometimes progressed to ulceration accompanied by colony formation of the organisms. In mild infections, granulomatous lesions were localized in lymph follicles and never progressed to ulceration. Nonpathogenic strains were rapidly excreted without penetration of epithelial linings. Y. enterocolitica should be within the category of invasion type enteropathogenic bacteria such as Shigella and Salmonella. Pathogenic behavior of Y. enterocolitica is discussed in comparison with that of Shigella and Salmonella.

Studies on the pathogenicity of Yersinia enterocolitica. II. Interaction with cultured cells in vitro.

Analysis of the pathogenicity of Yersinia enterocolitica was performed by means of cell culture studies on the interaction of the organisms with HeLa cells and rabbit peritoneal macrophages based on observations of the pathogenic behavior of the organisms in vivo (II). The pathogenic strains, which successfully produced experimental enterocolitis in rabbits (II), had the ability to penetrate HeLa cells and to survive or multiply within the macrophages. The nonpathogenic strains, lacking the ability to produce pathological changes in rabbits (II), failed to penetrate HeLa cells, except for one strain, and also to survive totally or multiply within the macrophages. It was evident that the abilities of the organisms to penetrate epithelial linings which serve as the barrier of intestinal mucosa and to survive or multiply within the host cells appears to be closely related to the pathogenicity of Y. enterocolitica.

Studies on the pathogenicity of Yersinia enterocolitica. III. Comparative studies between Y. enterocolitica and Y. pseudotuberculosis.

Comparative studies on pathogenicity between Yersinia enterocolitica and Yersinia pseudotuberculosis were performed using experimental infection systems in vivo and in vitro. All of thestra ins of both species successfully produced experimental enterocolitis in rabbits although the severity varied with the strains challenged. The changes were characterized by granulomatous lesions with necrobiotic centers in reticuloendothelial tissues of the intestine, mesenteric lymph nodes, liver and spleen. These strains uniformly had the ability to penetrate HeLa cells and to survive or multiply within cultured rabbit peritoneal macrophages. In addition, in infections with strain TP-2 or PST-III of Y. pseudotuberculosis, catarrhal inflammation all over the small intestine and/or focal necrosis and parenchymatous degeneration in the liver were observed, along with the granulomatous lesions. These strains, at the same time, exhibited cytotoxic effects on the cultured cells. The pathogenic factors of Y. enterocolitica are discussed in comparison with those of Y. pseudotuberculosis.

Antichlamydial activity of ofloxacin.

The in vitro activity of ofloxacin, a new pyridone-carboxylic acid, against 11 strains of Chlamydia trachomatis and six strains of Chlamydia psittaci was determined. All test strains of both species were inhibited by 0.39 micrograms of ofloxacin per ml. The antichlamydial activity of ofloxacin was comparable to that of doxycycline and four- to eightfold less than that of minocycline. The results of this susceptibility test, coupled with those of previous pharmacokinetic analyses of ofloxacin, warrant further evaluation of its clinical usefulness against chlamydial infections.

Penetrability of ofloxacin into cultured epithelial cells and macrophages.

It is well known that the penetration of drugs into host cells is the minimal requirement to exhibit their efficacy against infections with intracellular bacteria. Thus the penetrability of new quinolones including ofloxacin, norfloxacin and ciprofloxacin was evaluated by comparing their intracellular and extracellular activities by the use of cell infection systems in vitro. It was evidenced that the new quinolones tested were penetrable into both epithelial cells and macrophages, however, ofloxacin was more penetrable than norfloxacin and ciprofloxacin into both types of cells which serve the nest for proliferation of intracellular bacteria.

Antimicrobial evaluation of cefoxitin: a new semisynthetic cephamycin. Comparative studies with cefazolin and cefalotin.

Antimicrobial activity of 3-carbamoyloxymethyl-7-alpha-methoxy-7-[2-(2-thienyl)-acetamido]-3-cephem-4-carboxylic acid (cefoxitin), a new semisynthetic cephamycin antibiotic, was studied in comparison with that of cefazolin and cefalotin. Cefoxitin exhibited antibacterial activity against both gramnegative and gram-positive bacteria, and its action was bactericidal. Against gram-negative bacteria, cefoxitin was highly active as well as cefazolin, and more active than cefalotin. Especially, cefoxitin was highly active not only against strains of clinical isolates of indole-positive Proteus and S. marcescens but also against those of E. coli and P. mirabilis which were resistant to cefazolin and/or cefalotin, respectively. In addition, cefoxitin was effective against the strains resistant to beta-lactam antibiotics including cefazolin and cefalotin. Cefoxitin was hardly active against the strains of E. cloacae and P. aeruginosa, similar to cefazolin and cefalotin. Against gram-postive bacteria, cefoxitin was less active than cefazolin and cefalotin. In protection tests in mice, cefoxitin and cefazolin were more effective against infection with E. coli than cefalotin. Furthermore, cefoxitin was more active against infections with S. marcescens and P. morgani than the other antibiotics. Cefoxitin, like cefalotin, was less effective against infection with S. aureus than cefazolin. Cefoxitin was highly resistant to hydrolysis by beta-lactamases derived from the organisms insusceptible to the antibiotic. This fact revealed that the resistance of the organisms to cefoxitin may be in part due to factors other than beta-lactamase inactivation.

In vivo comparison of avirulent Vwa- and Pgm- or Pstr phenotypes of yersiniae.

The abilities of Yersinia pestis to undergo restriction in Ca2+-deficient medium with concomitant production of V and W antigens (Vwa+) and to absorb exogenous pigments (Pgm+) are established virulence factors. Mutation of Y. pestis to Pgm- is known to promote resistance to pesticin (Pstr) and reduced lethality by peripheral routes of injection. Vwa+ Pgm- isolates of Y. pestis were shown in this study to retain virulence in mice when injected intravenously. Although Pgm- in appearance, wild-type cells of Yersinia pseudotuberculosis and Yersinia enterocolitica may also be sensitive to pesticin. Pstr mutants of Vwa+ strains of these species were similarly of reduced virulence, especially by peripheral routes of injection. The consequences of mutation to Vwa- and Pgm- or Pstr on growth and persistence in vivo were determined. After intravenous injection, Vwa+ yersiniae of all species exhibited sustained growth in mouse spleen, liver, and lung and accumulated in blood. Septicemia was not observed after similar injection of Vwa- mutants which were unable to maintain comparable rates of net increase in tissues. Mutation to Pgm- or Pstr did not influence proliferation but resulted in enhanced clearance from organs. It is known that reticuloendothelial cells serve as favored sites of replication for all wild-type yersiniae. Our results are consistent with the hypothesis that the Vwa+ phenotype favors growth within macrophages and that the Pgm+ and pesticin-sensitive phenotypes permit long-term, probably extracellular, retention within organs. Virulence in standard animal models (mice, rats, and guinea pigs) was not correlated with resistance to the bactericidal action of serum.

Improvement of 5-HT3 receptor binding assay: enhancement of specific [3H]quipazine binding with Triton X-100-treated membranes from rat cortex.

The 5-hydroxytryptamine (5-HT)3 receptor binding assay using [3H]quipazine was examined. It was impossible to obtain specific [3H]quipazine binding with the membrane fractions from rat cortex prepared by the usual procedure. When the membranes were pretreated with detergent Triton X-100, the ratio of specific [3H]quipazine binding markedly increased, depending upon the concentration of Triton X-100 in the range of 0.01-0.1% (w/v). At a concentration of more than 0.05%, the specific binding reached a maximum of 55 to 60% of the total binding. The specific [3H]quipazine binding to the Triton X-100-treated membranes was reversible and was potently inhibited by several 5-HT3 antagonists, while 5-HT1, 5-HT2 receptor antagonists and other receptor-specific ligands had no effect on the binding. Scatchard analysis indicated a single class of binding sites with a Kd of 0.62 nM and Bmax of 97 fmol/mg protein. Thus, the Triton X-100-treated membranes retained the characteristics of 5-HT3 binding sites, making it possible to use [3H]quipazine for a 5-HT3 receptor binding assay with a high ratio of specific binding.

Ascending pyelonephritis with Pseudomonas aeruginosa in mice.

Elastase- and protease- producing strain of Pseudomonas aeruginosa induced ascending pyelonephritis in mice by intracystic challenge. The pelvis was the site of primary foci development and necrotic, purulent lesions spread from the pelvis to the perihilar area and to the cortex. Severe necrosis was a characteristic of the present infection and caused systemic infection and host death without the development of chronic lesions. In animals challenged with inocula great enough to destroy the cystic mucosa, immediate hematogenous systemic infection without cellular responses led to host death.

Experimental pyelonephritis in mice following ascending infection with C. coli. Acute phase.

The initial interaction between uroepithelial cells and Escherichia coli which has adhesive or invasive activity for cultured cells was studied ultrastructurally at the in situ of infection in the model of ascending pyelonephritis in mice. The densely piliated adhesive strain E77156 isolated from the urine of a patient with urinary tract infection adhered to the pelvic and renal tubular epithelial cells and colonized on their cell surfaces and thereafter in the cytoplasm. The non-piliated invasive strain 633-65 isolated from a patient with dysentery-like syndrome did not colonize on the uroepithelial cell surfaces but easily penetrated into the cytoplasm of these cells. Thereafter multiplication was observed in their cytoplasm. Neither strain scarcely penetrated into the interstitium via the basement membrane of the renal tubules.

Experimental pyelonephritis in mice following ascending infection with E. coli. Chronic phase.

Adhesive and invasive strains of Escherichia coli induced chronic pyelonephritis in mice following the acute phase. The pathological features of the induced chronic pyelonephritis were different between the groups of mice infected with these strains. In piliated adhesive strain (E77156)-infection, the kidneys with viable bacilli showed pyonephrosis with incomplete obstruction or atrophy with coarse scar. Mice with these renal lesions showed high serum antibody levels, but histologically recurrent infection was frequent. On the other hand, in non-piliated invasive strain (633-65)-infection, sterile pyelonephritis developed. This chronic lesion was characterized by the migration of antigen-bearing macrophages and lymphocytes and by a negative serum antibody response. In infections with either strain the predominant lymphocytes in the renal lesions were smooth-surfaced T-lymphocytes.

Cefoxitin: synergism with aminoglycosides in vitro.

In vitro synergistic effect of cefoxitin with aminoglycosides such as gentamicin, tobramycin, amikacin and dibekacin against 15 Escherichia coli, 15 Staphylococcus aureus and 105 Pseudomonas aeruginosa strains of fresh clinical isolates was examined. The combination of cefoxitin with the aminoglycosides showed distinct synergy against many strains of P. aeruginosa which were highly resistant to cefoxitin alone, while it failed in synergism against E. coli and S. aureus. A possible mechanism of in vitro synergy between cefoxitin and aminoglycosides against P. aeruginosa was discussed.

Sub-inhibitory and post-antibiotic effects of an optically active isomer of ofloxacin.

The sub-inhibitory and post-antibiotic suppressive effects of DR-3355, an optically active isomer (S-(-) form) of (+/-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methylpiperazin-1-yl)-7- oxo-7H- pyrido[1,2,3-de]-1,4-benzoxazine-6-carboxylic acid (ofloxacin) were compared with those of ofloxacin, ciprofloxacin and ceftazidime by observing changes in bacterial multiplication and morphology in vitro. DR-3355 affected the proliferation of Staphylococcus aureus E46, Escherichia coli E77156 and Pseudomonas aeruginosa PI-III at 1/4 to 1/2 times the minimal inhibitory concentration (MIC) for each strain. The compound also affected the morphology of these strains at 1/32 times the MICs. Upon MIC basis, these sub-inhibitory effects of DR-3355 were almost identical to those of the reference compounds. On the other hand, exposure of E. coli to 1 and 4 times the MIC of DR-3355 for 3 h produced post-antibiotic effects of 0.7 and 1.9 h, respectively, and the effects of the compound were almost equal to those of ofloxacin and ciprofloxacin and much greater than that of ceftazidime.

Stimulation of non-specific resistance to infection by muroctasin.

The profile of the stimulant activity of a muramyl dipeptide (MDP) analog, N2-[(N-acetylmuramoyl)-L-alanyl-D-isoglutaminyl]-N6-stearoyl-L-lysine (MDP-Lys(L18), muroctasin) on the host resistance to infection was clarified through the following results. A comparative study of parent MDP and MDP-Lys(L18) in relation to the protection against infection with various pathogens revealed the same spectrum of antiinfectious activity, but a different potency: the greater strength of MDP-Lys(L18) was demonstrated both by the smaller influence of bacterial inoculum size on its activity and by the smaller minimal dosage required for inducing significant activity. The superiority of the oral application of MDP-Lys(L18) over MDP was also demonstrated. The protective activity of the compound was substantially influenced by the timing of treatment, the greatest activity being achieved by treatment 1 day before infection. Furthermore, treatment with MDP-Lys(L18) restored the depressed resistance induced by cyclophosphamide to systemic infection with E. coli in mice, and that induced by cortisone acetate to bacteremic pneumonia with P. aeruginosa in guinea pigs. The chemotherapeutic efficacy of antibiotics on E. coli infection was potentiated by combined use of MDP-Lys(L18) for normal mice and mice immunosuppressed with cyclophosphamide. From these findings, enhancement of the host resistance to infection by MDP-Lys(L18) may be an important aspect in the future evaluation of therapy for infection in immunocompromised patients. Finally, since the compound augmented 1. the function of polymorphonuclear leukocytes, such as chemotaxis, phagocytosis, and superoxide anion generating capacity.(ABSTRACT TRUNCATED AT 250 WORDS)

Roles of V antigen in promoting virulence and immunity in yersiniae.

It is established that yersiniae harboring an approximately 45-megadalton Vwa-plasmid can produce V and W antigens (Vwa+), and that sera containing anti-V provides passive protection to mice against Yersinia pestis. This observation was extended by the use of monospecific anti-V prepared by injecting rabbits with partially purified V, absorption of antisera with a Vwa- extract, and then separation of gamma-globulin by traditional processes of fractionation or by affinity chromatography. These preparations provided passive protection against 10 minimum lethal doses of virulent Y. pestis KIM, Yersinia pseudotuberculosis PB1, and Yersinia enterocolitica WA. Kinetics of elimination of these Vwa+ yersiniae from organs and blood of passively immunized mice closely resembled those of avirulent Vwa- mutants from normal mice. Injection into mice of sterile crude extracts of Y. pseudotuberculosis PB1 containing V promoted significant survival and retention of Vwa- mutants of Y. pestis, Y. pseudotuberculosis, and Y. enterocolitica. This effect was eliminated by the removal of V before injection by precipitation with monospecific antibody. These results indicate that V antigen per se is the major virulence factor mediated by Vwa-plasmids.