RESUMO
Choleragen, a diarrheagenic protein enterotoxin elaborated by Vibrio cholerae, has been isolated from the supernate of fermenter cultures by steps involving ammonium sulfate precipitation, DEAE cellulose, Sephadex G-75, and Agarose A-5m chromatography. The resulting product appears to be pure according to immunoelectrophoretic, disc electrophoretic, ultracentrifugal, and immunologic criteria. Sephadex gel filtration and membrane filtration studies suggest a molecular size of 61,000. The isolated product is highly active in inducing experimental cholera in infant and adult rabbit models. It also elicits, in small dosage, an increased vascular permeability in skin. These observations indicate that choleragenicity and increased vascular permeability are intimately associated phenomena and may be manifestations of the same basic mechanism. An additional, antigenically identical, protein has also been isolated by the same procedures. The latter substance, termed "choleragenoid", lacks the permeability effect and choleragenicity of the choleragen moiety. Its size (estimated from Sephadex gel filtration at 42,000) is smaller than that of choleragen and it also differs in charge. Choleragenoid may prove useful as a nontoxic immunogen to protect against pathologic effects of V. cholerae infection.
Assuntos
Cólera/etiologia , Enterotoxinas/isolamento & purificação , Vibrio/metabolismo , Animais , Antígenos/isolamento & purificação , Cólera/imunologia , Cromatografia , Eletroforese Descontínua , Soros Imunes/isolamento & purificação , Imunodifusão , Imunoeletroforese , Peso Molecular , CoelhosRESUMO
The exo-enterotoxin of Vibrio cholerae has been obtained in crystalline form. A solution of the crystalline protein was equal in potency to the parent pure toxin in both choleragenicity and skin reactivity. Crystals of the natural toxoid, choleragenoid, resemble those of the toxin in appearance. A solution of crystalline choleragenoid was equivalent to the parent preparation in the flocculation test.
Assuntos
Enterotoxinas/isolamento & purificação , Toxoides/isolamento & purificação , Vibrio/análise , Animais , Cólera , Cromatografia , Cristalização , Eletroforese Descontínua , Enterotoxinas/administração & dosagem , Testes de Floculação , Microscopia , Coelhos , Testes Cutâneos , Solubilidade , Toxoides/administração & dosagemRESUMO
Matings between strains of Vibrio cholerae differing in toxinogenicity, nutritional requirements, and antibiotic susceptibilities were performed in order to determine the location of the gene tox that controls production of cholera enterotoxin. Segregation analysis shows that tox is linked to a gene required for histidine biosynthesis. Our data indicate that the tox gene is located on the bacterial chromosome and not on a plasmid in the strains of V. cholerae studied.
Assuntos
Conjugação Genética , Enterotoxinas/biossíntese , Genes , Vibrio cholerae/metabolismo , Mapeamento Cromossômico , Cromossomos Bacterianos , Cruzamentos Genéticos , Ligação Genética , Genótipo , Histidina/biossíntese , Recombinação GenéticaRESUMO
Large crystals of the cholera toxin were grown; their dimensions, symmetry (P21), order, and resistance to radiation make them ideally suited for a high-resolution x-ray structure determination. There is one molecule (approximately 84,000 daltons) per asymmetric unit, and therefore the lattice reveals no molecular symmetry. Two distinct bioassays indicate that the protein from dissolved crystals retains full biological activity.
Assuntos
Toxinas Bacterianas , Vibrio cholerae , Cristalização , Peso Molecular , Conformação Proteica , Difração de Raios XRESUMO
Pasteurella multocida is the commonest organism infecting pet bites. Anecdotal reports tend to overemphasize dramatic outcomes. We aimed to study a large database of P. multocida infections. This retrospective survey of P. multocida infections in Israeli hospitals refers to the y 2000-2005. Clinical microbiologists were contacted by email and asked to perform a back-search of their hospital's records for isolates of P. multocida. The charts of patients growing P. multocida were abstracted into a structured questionnaire. 77 cases were identified in 12 hospitals, yielding an annual incidence of 0.19/100,000. The mean age was 49.2+/-26.5 y and the mortality rate was 2.6%. Those who died were >65 y of age, had diabetes mellitus or cirrhosis and were bacteraemic. One-third of the cases occurred in people aged > or =65 y. Cats caused most of these infections (54%). Surgery for debridement was common (53.7%), but no-one required amputation; a second- and third-look operation was necessary for these patients. Bacteraemia was found in 32.5% of patients and was significantly more common among those aged >60 y (p =0.044). Hospitalized patients with P. multocida have a favourable prognosis, apart from elderly and bacteraemic patients with comorbidities. Surgery and reoperations may be required in about half of the patients.
Assuntos
Infecções por Pasteurella/epidemiologia , Pasteurella multocida/isolamento & purificação , Adolescente , Adulto , Idoso , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Bacteriemia/epidemiologia , Distribuição de Qui-Quadrado , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Israel/epidemiologia , Masculino , Pessoa de Meia-Idade , Infecções por Pasteurella/diagnóstico , Infecções por Pasteurella/tratamento farmacológico , Estudos Retrospectivos , Inquéritos e QuestionáriosRESUMO
Many of the genes responsible for axial patterning have been conserved throughout evolution. Recent studies of invertebrates and vertebrates are extending our understanding of this molecular conservation into the anterior region of the animal embryo, including the developing brain. These studies suggest that this domain is specified according to a genetic paradigm that is different from that governing trunk development, and are also beginning to provide insights into the structures that underlie the rostral brain.
Assuntos
Evolução Biológica , Drosophila/genética , Mamíferos/genética , Prosencéfalo , Animais , Regulação da Expressão Gênica no Desenvolvimento , CabeçaRESUMO
The ability of potential pathogens to acquire iron in a host is an important determinant of both their virulence and the nature of the infection produced. Virulent gram-negative bacteria are capable of acquiring sufficient iron from the host because their virulence (for chick embryos) is unaffected by exogenous iron. Avirulent mutants which are apparently limited in their ability to acquire iron could be isolated from the virulent strains. The lethality of these mutants was significantly enhanced by exogenous iron. Reduction of the relatively high serum iron saturation of chick embryos (to levels more closely approximating those in man) by pretreatment with iron-binding proteins or endotoxin inhibits the lethality of some virulent bacteria. Those bacteria whose virulence was reduced include the Shigella, Vibrio cholerae and strains of Neisseria gonorrhoeae, all of which are nondisseminating pathogens in the normal human host. Pathogens which produce septicemic and disseminating infections such as Neisseria meningitidis, Haemophilus influenzae type B, Escherichia coli possessing K-1 antigen, Pseudomonas aeruginosa and Salmonella typhimurium and disseminating strains of N. gonorrhoeae were, in general, unaffected by reduced serum iron saturation. These disseminating bacteria appeared to produce greater quantities of compounds (siderophores) which stimulated microbial growth in low-iron media than did the nondisseminating pathogens. Thus, the gram-negative bacteria tested can be divided into four major classes according to their responses to modifications in iron levels in the chick embryo model and these results correlate with the nature of the infections which they typically produce in man.
Assuntos
Infecções Bacterianas/metabolismo , Ferro/metabolismo , Animais , Bactérias/efeitos dos fármacos , Bactérias/metabolismo , Bactérias/patogenicidade , Infecções Bacterianas/imunologia , Embrião de Galinha , Conalbumina/farmacologia , Ferro/farmacologia , Quelantes de Ferro/metabolismoRESUMO
Spontaneous and chemically induced mutants with reduced ability to produce cholera enterotoxin (choleragen) as an extracellular protein were isolated from Vibrio cholerae strains 569B Inaba, a classical cholera vibrio, and 3083-2 Ogawa, an El Tor vibrio. By qualitative and quantitative immunological assay in vitro such mutants could be separated into different classes characterized either by production of no detectable choleragen (tox minus), or of small quantities of extracellular choleragen, or of large quantities of cell-associated choleragen but little extracellular choleragen. Analysis of proteins in concentrated culture supernates by electrophoresis in polyacrylamide gels showed that cultures from tox minus strains lacked proteins with electrophoretic mobilities corresponding with choleragen or the spontaneously formed toxoid (choleragenoid). Infant rabbits infected with the tox minus strains remained asymptomatic or developed milder symptoms than rabbits infected with the tox+ parental strains. When symptoms of cholera developed after inoculation with tox minus mutants, detectable numbers of tox+ revertants could be isolated from the intestines of the infected animals. Two tox minus strains, designated M13 and M27, caused no sumptoms and showed no evidence of reversion to tox+ during single passage in infant rabbits, and mutant M13 also remained avirulent and stably tox minus during six cycles of serial passage in infant rabbits. Strains M13 and M27 were also noncholeragenic in acult rabbit ileal loops. Quantitative cultures of the intestines from infected infant rabbits demonstrated that the avirulent mutant M13 can multiply in vivo and can persist in the intestinal tract for at least 48 h.
Assuntos
Cólera/microbiologia , Enterotoxinas/biossíntese , Mutação , Vibrio cholerae/metabolismo , Animais , Bioensaio , Bovinos , Cólera/genética , Modelos Animais de Doenças , Eletroforese em Gel de Poliacrilamida , Íleo/microbiologia , Imunodifusão , Enteropatias/genética , Enteropatias/microbiologia , Fenótipo , Coelhos , VirulênciaRESUMO
To assess the ion transport mechanism by which cholera causes the small bowel to secrete, ion transport rates and electrical potential difference (PD) were determined simultaneously in the normal and choleragen-treated dog ileum in vivo. The results indicate that, during cholera, HCO(3) is actively secreted (i.e., against both an electrical and a concentration gradient); Cl is also actively secreted, against a modest electrochemical gradient. Electrogenic pumping of one or both of these anions is probably responsible for an observed PD change of approximately 13 mv (lumen negative). Na secretion can be accounted for entirely by passive ion movement. K secretion can be partly explained by passive diffusion secondary to the negative intraluminal PD; however, its concentration in the secreted fluid is two to three times higher than expected on the basis of passive forces, suggesting a component of active K secretion. The PD response of the choleragen-treated ileum is normal in response to glucose, but there was no PD response to saline-free mannitol perfusion. This suggests that the normal differential permeability of the ileum to anions and cations may be altered by choleragen, although other explanations of this finding are also possible.
Assuntos
Transporte Biológico Ativo , Cólera/metabolismo , Íleo/metabolismo , Animais , Bicarbonatos/metabolismo , Cloretos/metabolismo , Cães , Glucose/farmacologia , Íleo/efeitos dos fármacos , Absorção Intestinal , Manitol/farmacologia , Potássio/metabolismo , Potenciometria , Sódio/metabolismo , Toxoides/farmacologiaRESUMO
PURPOSE: To assess the changes observed in surgical site infection (SSI) rates following total joint arthroplasty (TJA) after the introduction of an infection control programme and evaluate the risk factors for the development of these infections. DESIGN: Prospective cohort study. SETTING: Large tertiary medical centre in Israel. METHODS: Data about SSIs and potential prophylaxis-, patient-, and procedure-related risk factors were collected for all patients who underwent elective total hip and total knee arthroplasty during the study period. Multivariant analyses were conducted to determine which significant covariates affected the outcome. RESULTS: During the 76-month study period, SSIs (superficial and deep) occurred in 64 (4.4%) of 1554 patients. As compared with the 34 (7.7%) SSIs that occurred in the first 25 months, there were 23 (4.7%) SSIs in the following 25 months, and only 7 (1.3%) SSIs in the last third of the study (p = 0.058 and <0.001, respectively). A multiple logistic regression model indicated that risk factors for prosthetic joint infection were a National Nosocomial Infections Surveillance (NNIS) System surgical patient risk index score of 1 (OR 1.8; 95% CI 1.1-3.1) or 2 (OR 2.8; 95% CI 1.2-11.8). The incidence of SSI was not correlated with the timing, nor the duration of antibiotic prophylaxis. CONCLUSIONS: The introduction of preventive measures and surveillance coincided with a significant reduction in SSIs following TJA in our institution. The risk of infection correlated with higher scores in the NNIS System surgical patient risk.
Assuntos
Artroplastia de Quadril , Artroplastia do Joelho , Infecção da Ferida Cirúrgica/prevenção & controle , Idoso , Idoso de 80 Anos ou mais , Antibioticoprofilaxia , Feminino , Humanos , Incidência , Controle de Infecções , Israel/epidemiologia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Vigilância da População , Estudos Prospectivos , Fatores de Risco , Infecção da Ferida Cirúrgica/epidemiologia , Infecção da Ferida Cirúrgica/etiologia , Infecção da Ferida Cirúrgica/microbiologia , Centros de Atenção Terciária/estatística & dados numéricosRESUMO
In the original article, one of the co-author's family name has been published incorrectly.
RESUMO
The homeobox gene orthodenticle (otd) specifies anterior head development in the Drosophila embryo, otd-related genes are also found in vertebrates, with expression patterns suggesting that they are important for the development of anterior regions of the head and brain. Here, we analyze the molecular mechanisms by which otd expression is activated within its normal domain in the head and repressed outside this region. We demonstrate that, contrary to early models of embryonic pattern formation, high levels of the bicoid morphogen are not required for otd activation or for the establishment of anterior head structures. We also show that the terminal system contributes to otd activation in the head primordium. Finally, we identify a novel pathway mediated by the gap gene huckebein through which three maternal systems cooperate to repress otd expression at the anterior terminus of the embryo.
Assuntos
Proteínas de Drosophila , Drosophila melanogaster/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Genes Homeobox , Proteínas de Homeodomínio/biossíntese , Transativadores , Animais , Proteínas de Ligação a DNA/fisiologia , Drosophila melanogaster/genética , Desenvolvimento Embrionário , Cabeça/embriologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/fisiologia , Hormônios de Inseto/fisiologia , Morfogênese/genética , Proteínas Nucleares/fisiologia , Fosfoproteínas/fisiologia , Receptores Proteína Tirosina Quinases/fisiologia , Fatores de Transcrição/fisiologiaRESUMO
To analyze the molecular mechanism of pattern formation in the anteriormost regions of the zebrafish embryo, we isolated two zebrafish sequences, zOtx1 and zOtx2, related to the Drosophila orthodenticle (otd) and two murine Otx genes. zOtx1 and zOtx2 encode predicted gene products which are 82% and 94% identical to the corresponding mouse proteins. Transcripts of both zebrafish genes appear abruptly at high levels in a triangular patch at the animal pole of the mid-gastrula, a region which contains cells fated to become midbrain and forebrain. Between 9 and 14 h of development, zOtx transcripts disappear from forebrain regions in a manner characteristic for each gene, and from 14 to 24 h, particular regions of the forebrain and midbrain express one or both genes. The posterior limit of expression of both genes in 10-30-h embryos forms a sharp boundary at the posterior border of the midbrain. As in the mouse, the early expression patterns of the zOtx genes are consistent with a role in defining midbrain and forebrain territories. However, there are a number of interesting differences between the forebrain and midbrain regions which express the genes in the two species.
Assuntos
Encéfalo/embriologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Peixe-Zebra/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Encéfalo/metabolismo , Drosophila , Gástrula/fisiologia , Mesencéfalo/embriologia , Mesencéfalo/metabolismo , Camundongos , Dados de Sequência Molecular , Prosencéfalo/embriologia , Prosencéfalo/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Peixe-Zebra/embriologiaRESUMO
Three abscisic acid (ABA)-controlled responses (seed dormancy, inhibition of germination by applied ABA, and stomatal closure) were compared in wild-type versus homo- and heterozygotes of two Arabidopsis thaliana ABA-insensitive mutants, abi1 and abi2. We found that sensitivity of seeds to applied ABA is partially maternally controlled but that seed dormancy is determined by the embryonic genotype. The effects of the abi1 and abi2 mutations on ABA sensitivity of seed germination ranged from recessive to nearly fully dominant, depending on the parental source of the mutant allele. This maternal effect disappeared during vegetative growth. Stomatal regulation in heterozygotes showed substantial variability, but the average water loss was intermediate between that of homozygous mutants and wild type.
RESUMO
In the context of a prospective, longitudinal, and controlled nosologic study, the characteristics and diagnostic validity of major depressive disorder, dysthymic disorder, and adjustment disorder with depressed mood were examined in a school-aged cohort. The entities were distinguishable on several dimensions such as age at onset and pattern of recovery. Time to recovery from onset was most favorable for the adjustment disorders (90% remission rate during nine months) and least so for the dysthymias (89% remission rate during six years). Major depression and dysthymia were similar with respect to the prevalence of concurrent nonaffective disorders. For both, early age at onset predicted a more protracted illness. Treatment contacts, none of which were under the control of the investigators, had no clear impact on recovery from the depressions.
Assuntos
Transtorno Depressivo/diagnóstico , Logro , Transtornos de Adaptação/complicações , Transtornos de Adaptação/diagnóstico , Transtornos de Adaptação/psicologia , Adolescente , Adulto , Fatores Etários , Transtorno do Deficit de Atenção com Hiperatividade/complicações , Transtorno do Deficit de Atenção com Hiperatividade/diagnóstico , Transtorno do Deficit de Atenção com Hiperatividade/psicologia , Criança , Ensaios Clínicos como Assunto , Transtorno Depressivo/complicações , Transtorno Depressivo/psicologia , Feminino , Humanos , Masculino , Avaliação de Processos e Resultados em Cuidados de Saúde , Estudos ProspectivosRESUMO
As part of a longitudinal nosologic study of major depressive disorder, dysthymic disorder (DD), and adjustment disorder with depressed mood ( ADDM ) in a school-aged cohort, the predictive validity of each diagnosis was examined. Using all available data on the course of the disorders, the criterion was the first subsequent major depressive episode. Major depressive disorder and DD signaled a similarly high risk of a new bout of depressive illness. For the children who recovered from their first episode of major depression and then had their second one (40%), the free interval did not exceed two years; an underlying dysthymia increased the risk of recurrence. Major depression and dysthymia were distinct from ADDM and a set of control disorders; the latter two diagnostic groups were associated with a minimal risk for major depression.
Assuntos
Transtorno Depressivo/diagnóstico , Transtornos de Adaptação/diagnóstico , Transtornos de Adaptação/psicologia , Adolescente , Adulto , Fatores Etários , Criança , Transtornos do Comportamento Infantil/diagnóstico , Transtornos do Comportamento Infantil/psicologia , Transtorno Depressivo/psicologia , Feminino , Seguimentos , Humanos , Estudos Longitudinais , Masculino , Avaliação de Processos e Resultados em Cuidados de Saúde , Recidiva , Risco , Fatores de TempoRESUMO
Continuous overlapping synthetic hexapeptides representing the entire 103 amino acid sequence of the immunodominant B-subunit protein of cholera enterotoxin were used to examine reactivities of a variety of antisera in attempts to detect and define sequence-related (continuous) antigenic regions. The validity of the methods was established by the reactions of polyclonal antisera raised against longer synthetic peptides with appropriate synthetic hexapeptides. An unexpected cross-reaction is attributed to the presence of three identical amino acids (Gln16-Ile17-His18)--although in different order (Gln56-His57-Ile58)--in two parts of the B-subunit chain. Adsorption studies using polyclonal rabbit antisera revealed that, in many instances, denatured B-subunit protein more effectively removed reactivity with hexapeptides than did the native protein. Native holotoxin was more effective than native B-subunit. Sera from human cholera convalescents gave diffuse patterns of reactivity with synthetic hexapeptides--primarily against regions of reactive hexapeptides rather than with clearly defined continuous epitopes. Among many epitopic regions encountered, a strongly reactive tetramer, Ser-Gln-His-Ile (SQHI), was discovered in a highly conserved region, residues 55-58, of the B-subunit amino acid sequence. Adsorption studies revealed that this epitope is apparently exposed on the surface of the native protein. Amino acid substitution revealed the essentiality of Gln and His residues to this epitope. Gly54 was not part of the epitope but substitution of acidic residues Glu and Asp for Gly eliminated reactivity with antibody. The results suggest that continuous epitopes may contribute to the antigenicity of the native toxin protein and may be potentially useful for development of a peptide vaccine.
Assuntos
Toxina da Cólera/genética , Epitopos/genética , Fragmentos de Peptídeos/genética , Sequência de Aminoácidos , Animais , Toxina da Cólera/imunologia , Epitopos/imunologia , Soros Imunes/imunologia , Técnicas In Vitro , Dados de Sequência Molecular , Fragmentos de Peptídeos/imunologia , RatosRESUMO
As part of a prospective, longitudinal study of school-aged children with newly diagnosed insulin-dependent diabetes mellitus (IDDM), we examined how the parents adjusted to the illness. The present article documents this process for the first year of IDDM. We found no support for earlier claims that most parents resort to blatantly neurotic or psychopathologic behavior to cope. Instead, the initial strain of living with IDDM generally elicited mild and subclinical depression, anxiety, and overall distress. Mothers were more affected than fathers: they were more symptomatic (about one of four developed a mild grief reaction) and the bulk of them worried considerably about their children. However, the parents' initial emotional upheaval resolved in approximately equal to 6 mo; most mothers came to terms with IDDM by the end of the first year; and other areas of parental functioning (e.g., quality of their marriage) were not affected. Therefore, along with our previous report on how the children coped initially, the findings document the emotional resiliency of families during the first year of IDDM.
Assuntos
Adaptação Psicológica , Diabetes Mellitus Tipo 1 , Pais/psicologia , Adolescente , Adulto , Ansiedade/etiologia , Criança , Depressão/etiologia , Diabetes Mellitus Tipo 1/diagnóstico , Feminino , Humanos , Masculino , Casamento , Pessoa de Meia-Idade , Estresse Fisiológico/etiologiaRESUMO
El Tor biotype Vibrio cholerae strains express a cell-associated mannose-sensitive hemagglutinin (MSHA) which is a putative attachment factor. Several MSHA-negative mutants from V. cholerae strain JBK70 were previously generated by Tn5 mutagenesis [Finn et al., Infect. Immun. 55 (1987) 942-946]. The chromosomal DNA regions containing the Tn5 insertions were isolated from eight strains for further analysis. Nucleotide sequencing of the insertional junctions and corresponding clones containing the intact chromosomal region from the parental strain revealed the presence of several contiguous ORFs. Only two ORFs of this region had received insertions, and these showed remarkable homology to genes involved in the general secretory pathway found in several Gram- bacterial species. Proteins corresponding to the observed ORFs were visualized with the T7 promoter/RNA polymerase expression system. Marker exchange mutagenesis was used to insert kanamycin-resistance cassettes and TnphoA insertions into different locations of this region in the chromosome of wild-type V. cholerae strains. The phenotypes of these mutants showed that this DNA region is involved in MSHA production, but is not required for general extracellular protein secretion.
Assuntos
Elementos de DNA Transponíveis , Hemaglutininas/genética , Vibrio cholerae/genética , Sequência de Aminoácidos , Clonagem Molecular , Genoma Bacteriano , Manose/farmacologia , Dados de Sequência Molecular , Mutagênese , Fases de Leitura Aberta , Fenótipo , Homologia de Sequência de AminoácidosRESUMO
A simple technique, checkerboard immunoblotting (CBIB), is described, which facilitates the examination of multiple antigen/antibody interactions, conveniently and reproducibly, using minimal amounts of reactants. Antigens, immobilized on a solid-phase membrane in parallel lanes, are allowed to react with primary antibodies, applied in lanes perpendicular to the antigens, and the reactions are developed with appropriately labeled secondary antibody and substrate. Positive reactions, at the intersections of antigen/antibody lanes, are small squares, giving a checkerboard appearance to the blot. The results are easily read visually and presented in the form of a permanent record. CBIB has wide range of applications, including the screening of hybridomas for monoclonal antibody production. With the cholera toxin (CT)-related antigens used, homologous reactions were markedly stronger then heterologous reactions.