RESUMO
OBJECTIVES: The Z0011 trial questioned the role of axillary ultrasound (AxUS) in preoperative staging of breast cancer in patients with ≤2 positive sentinel lymph nodes (SLN). The purpose of this study was to correlate the number of abnormal nodes on AxUS with final nodal burden and determine the utility of AxUS with sampling (AxUS + S) in preoperative staging. METHODS: Six hundred and seventy-nine patients underwent pre-operative AxUS. Suspicious nodes were sampled. Negative axillae proceeded to SLN biopsy. The number of abnormal nodes identified on ultrasound and final histology as well as sensitivity and specificity for AxUS + S were calculated. Subgroup analysis was performed on Z0011 eligible patients. RESULTS: Two hundred and ninety-six patients had positive axillary nodes on final histology with 169 detected by AxUS + S (sensitivity 86.2%, specificity 100%, PPV 100 %, NPV 71.9%). Patients with nodal metastases identified by AxUS had a mean burden of 7.3 nodes on histology (1 node on AxUS = 5.2 nodes on histology, 2 nodes on AxUS = 7.5 nodes, >2 nodes = 10.1 nodes). Patients diagnosed on SLNB had a mean burden of 2.2 nodes. CONCLUSION: A single nodal metastasis detected on AxUS + S correlated with a mean of 5.2 nodes on final histology highlighting that AxUS remains essential in guiding appropriate management of the axilla in breast cancer. KEY POINTS: ⢠Axillary ultrasound +/- sampling is an essential technique in preoperative axillary staging. ⢠Axillary ultrasound findings correlate with final histological axillary node disease burden. ⢠Axillary ultrasound can help triage patients who require axillary lymph node dissection. ⢠The role of axillary ultrasound in breast cancer staging continues to evolve.
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Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/patologia , Linfonodos/diagnóstico por imagem , Linfonodos/patologia , Cuidados Pré-Operatórios , Adulto , Idoso , Idoso de 80 Anos ou mais , Axila , Estudos de Coortes , Bases de Dados Factuais , Feminino , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Estudos Retrospectivos , Sensibilidade e Especificidade , Biópsia de Linfonodo Sentinela/métodos , Ultrassonografia , Adulto JovemRESUMO
Injury to the spleen is a recognised complication of colorectal resections involving mobilisation of the splenic flexure. Bleeding from the spleen is difficult to control and not infrequently requires splenectomy with its attendant lifelong potential haematological and immunological complications. Furthermore, conversion from a laparoscopic to an open procedure may be required as splenic haemorrhage is more difficult to control laparoscopically. We describe a technique for control of bleeding from the inferior pole of the spleen, used during laparoscopic splenectomy, which may be applied to either open or laparoscopic surgery to achieve haemostasis thereby obviating splenectomy and in laparoscopic cases, conversion to open.
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Perda Sanguínea Cirúrgica/prevenção & controle , Colo Transverso/cirurgia , Complicações Intraoperatórias/cirurgia , Laparoscopia/efeitos adversos , Baço/cirurgia , Esplenectomia/métodos , Colo Transverso/irrigação sanguínea , Conversão para Cirurgia Aberta , Humanos , Laparoscopia/métodos , Baço/irrigação sanguínea , Baço/lesões , Resultado do TratamentoRESUMO
BACKGROUND: Intussusception in adult patients is uncommon and appendiceal lead points are particularly rare. CASE PRESENTATION: We present the case of a 42-year-old male with a history of ulcerative colitis, presenting with sudden onset abdominal pain and bloody diarrhoea. Endoscopy revealed grossly normal mucosa in the descending colon with a congested polypoid mass in the proximal transverse colon. Computed tomography revealed ileocecal intussusception at the hepatic flexure. A right hemicolectomy was performed, where a grossly dilated appendix was noted, resected and sent for histopathological evaluation. Results revealed low-grade appendiceal mucinous neoplasm. Post-operatively, the patient remained symptom free, however required reintroduction of biologic therapy due to relapse of his ulcerative colitis 12 weeks later. CONCLUSION: This case depicts a rare acute surgical presentation and reminds physicians and surgeons of the importance of 'thinking outside the box' in clinical practice.
RESUMO
Kaposi sarcoma (KS), a multifocal neoplasm of the skin that can spread to visceral organs, is the most prevalent malignant tumor in acquired immuno deficiency syndrome (AIDS) patients. KS-associated herpesvirus (KSHV or HHV8) is considered the primary etiological factor of this malignancy, as well as of primary effusion lymphoma and multicentric Castleman's disease. KS lesions are characterized by proliferating spindle cells of endothelial cell (EC) origin. The action of the insulin-like growth factor (IGF) system has been implicated in many malignancies, and recent data have demonstrated that the IGF-I receptor (IGF-IR) is required for in vitro growth of the KS-derived KSIMM cell line. To examine whether the IGF pathway is also involved in KSHV-mediated transformation of ECs, we examined the expression and function of the IGF system in KSHV-infected, immortalized dermal microvascular EC (E-DMVEC). The expression of the insulin receptor (IR) was strongly induced in latently infected E-DMVEC, whereas the expression levels of the IGF-IR remained unchanged. Gene knockdown of IR, but not IGF-IR, prevented the characteristic focus formation seen in KSHV-infected E-DMVEC. Similarly, treatment with the IR-specific small-molecule inhibitor HNMPA-(AM(3)) inhibited postconfluent growth. These data suggest a role for the IR, but not the IGF-IR, in KSHV-induced transformation of vascular ECs.
Assuntos
Transformação Celular Viral/genética , Receptor de Insulina/fisiologia , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/virologia , Linhagem Celular Transformada , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células Endoteliais/virologia , Herpesvirus Humano 8/fisiologia , Humanos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Naftalenos/farmacologia , Organofosfonatos/farmacologia , RNA Interferente Pequeno/farmacologia , Receptor IGF Tipo 1/antagonistas & inibidores , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismo , Receptor de Insulina/antagonistas & inibidores , Receptor de Insulina/genética , Sarcoma de Kaposi/patologiaRESUMO
Differences in the activated partial thromboplastin time (aPTT) were shown when blood taken from patients receiving intravenous heparin therapy was collected into 5 ml and 1 ml citrate containers. Mean aPTTs were 27% shorter with the plasma from the 1 ml citrate containers (n = 23). These results were paralleled by a 37% reduction in the mean heparin concentration (n = 11) and a 77% increase in the mean platelet factor 4 (PF4) concentration (n = 7). This phenomenon is due to increased platelet activation and subsequent increased heparin neutralization in the 1 ml citrate container. In an attempt to overcome this, the citrate was removed from a 1 ml container and replaced with a buffered tri-sodium citrate solution containing theophylline, adenosine and dipyridamole anticoagulant (CTAD). Blood from heparinized patients taken into both 5 ml citrate and 1 ml CTAD showed a correction of the shortening artefact in the low volume container. The mean aPTT of plasmas from the 1 ml CTAD container showed an increase of 10% compared with the 5 ml citrate. There was no significant difference in the mean heparin or PF4 concentrations of blood taken into either container. The 1 ml CTAD tube described is a suitable collection container for monitoring heparin in neonates or patients who are difficult to venepuncture and overcomes the neutralization of heparin in part filled low volume containers.
Assuntos
Coleta de Amostras Sanguíneas/instrumentação , Heparina/uso terapêutico , Tempo de Tromboplastina Parcial , Citratos , Ácido Cítrico , Heparina/sangue , Humanos , Fator Plaquetário 4/análiseRESUMO
Butylated hydroxyanisole (BHA) was found to induce chromosome aberrations in Chinese hamster ovary (CHO) cells in the presence of Aroclor-induced rat-liver S9. The effects were more marked when washed microsomes were employed and chromosome damage was considerably reduced in the presence of catalase, suggesting that hydrogen peroxide was involved. Stimulation of H2O2 production by BHA in S9 or microsome incubation mixtures was demonstrated using the catalase-mediated production of formaldehyde from methanol. One of the major microsomal metabolites of BHA, tert.-butyl hydroquinone (t-BHQ), which autoxidises in solution producing H2O2 also induced extensive catalase-sensitive chromosome damage in the absence of metabolic activation. These observations suggest that extracellular generation of reactive oxygen species may be implicated in the mechanism of BHA clastogenicity in vitro. However, chromosome damage was not completely abolished by catalase and the end product of t-BHQ oxidation, tert.-butyl quinone, was also weakly clastogenic, suggesting that intracellular effects of quinone metabolites may also be involved in the clastogenicity of BHA.
Assuntos
Hidroxianisol Butilado/toxicidade , Aberrações Cromossômicas/efeitos dos fármacos , Microssomos/efeitos dos fármacos , Animais , Biotransformação , Hidroxianisol Butilado/farmacocinética , Catalase/farmacologia , Linhagem Celular , Sobrevivência Celular , Cricetinae , Cricetulus , Peróxido de Hidrogênio/metabolismo , Hidroquinonas/toxicidade , Dose Letal Mediana , Masculino , Microssomos/metabolismo , Testes de Mutagenicidade , Ratos , Ratos EndogâmicosRESUMO
Butylated hydroxyanisole (BHA) was found to react readily with nitrite in acidified physiological saline. With low concentrations of reactants at pH 2, HPLC analysis demonstrated the formation of two products, tert-butylquinone (BQ) and a second, unidentified compound. Neutralized BHA/nitrite reaction mixtures were highly toxic to cultured Chinese hamster ovary (CHO) cells. At non-lethal concentrations, causing some cell cycle delay, there was a statistically significant but variable induction of endoreduplication and tetraploidy and a very weak induction of chromosome breakage. The effects were similar to those of pure BQ. It is suggested that the formation of toxic products from BHA, by an oxidative reaction such as that described with nitrite, might be involved in the mechanism of BHA carcinogenesis in the rat forestomach.
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BACKGROUND: This study evaluates the combined role of axillary ultrasound (Ax US), fine needle aspiration (FNAC) and intraoperative frozen section analysis of the sentinel node (FS SN) in a practical, time efficient algorithm to reduce the requirement for reoperation for axillary clearance in breast cancer in a busy tertiary unit. METHODS: Between October 2007 and June 2009 188 women underwent Ax US as a first investigation for nodal status. Suspicious nodes were biopsied, negative axillae proceeded to FS SN at time of primary breast surgery. All confirmed positive cases proceeded to immediate axillary clearance. RESULTS: 93 women had positive axillary nodes at final histology. Ax US + FNAC identified 59 positive axillae and had a sensitivity of 63.4% and specificity of 100%. FS SN identified a further 26 cases with a sensitivity of 76.5% and specificity of 100%. Overall, only 8 women required reoperation for axillary clearance. Sensitivity for the combined procedures was 91.4%. Commencement of adjuvant therapy was significantly less in those women identified earlier compared to those requiring a second operation (23.3 days vs 49.0 days, p < 0.005). CONCLUSION: 95.7% of cases were diagnosed accurately in the perioperative period, preventing delay to triage to definitive oncological care and reducing requirement for costly reoperation.
Assuntos
Algoritmos , Axila/patologia , Neoplasias da Mama/patologia , Metástase Linfática/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Axila/diagnóstico por imagem , Neoplasias da Mama/terapia , Feminino , Secções Congeladas , Humanos , Metástase Linfática/patologia , Pessoa de Meia-Idade , Sistema de Registros , Sensibilidade e Especificidade , Biópsia de Linfonodo Sentinela , UltrassonografiaRESUMO
As marketing budgets for physician liaison departments increase, health care marketers are being held more accountable for their efforts by managers. Market researchers at The Cleveland Clinic Foundation (CCF) have developed a health care organizational information dissemination model that provides an understanding of how referring physicians choose a referral center for their patients. Interviews with 89 new referring physicians show patient influence and interpersonal media to be the two most influential channels of information. Financial analysis of referrals shows that a true physician referral generates significantly more revenue than a patient-influenced referral. CCF managers and marketers have used the data to understand better the effectiveness of their current programs targeted at physicians.
Assuntos
Tomada de Decisões , Marketing de Serviços de Saúde/métodos , Médicos/psicologia , Encaminhamento e Consulta/economia , Humanos , Entrevistas como Assunto , Modelos Teóricos , Ohio , Médicos/normasRESUMO
In the context of the third UKEMS collaborative trial on cell mutation assays, three chemicals have been tested for mutagenicity using the Chinese hamster ovary (CHO) cell/HPRT assay. The protocol employed was based on the re-spreading of monolayer cultures, maintaining one million cells in growth for an expression time of 7 days. Ethyl methanesulphonate gave clear and consistent positive results in the absence of S9 and a clear response to benzo[a]pyrene was obtained with 1% S9. Benzidine gave no evidence of mutagenicity either with or without S9. The results show that the CHO/HPRT assay protocol used in this study is adequate for the detection of potent mutagens. The failure to detect benzidine as a mutagen may be due either to a basic unresponsiveness of the system or to a lack of sensitivity because of the limitation on the numbers of cells which can be maintained during expression. Statistical analysis showed that significant variation occurred between cell survival and mutagenicity estimates for replicate cultures, demonstrating the need for duplicate cultures.
Assuntos
Benzidinas/toxicidade , Benzo(a)pireno/toxicidade , Metanossulfonato de Etila/toxicidade , Testes de Mutagenicidade/métodos , Mutagênicos , Animais , Benzidinas/metabolismo , Benzo(a)pireno/metabolismo , Biotransformação , Linhagem Celular , Sobrevivência Celular , Glucose-6-Fosfato , Glucofosfatos/metabolismo , Hipoxantina Fosforribosiltransferase/biossíntese , Microssomos Hepáticos/metabolismoRESUMO
We used the Vibrio cholerae Fur protein as a model of iron-sensitive repressor proteins in gram-negative bacteria. Utilizing manganese mutagenesis, we isolated twelve independent mutations in V. cholerae fur that resulted in partial or complete loss of Fur repressor function. The mutant fur genes were recovered by PCR and sequenced; 11 of the 12 contained point mutations (two of which were identical), and one contained a 7-bp insertion that resulted in premature truncation of Fur. All of the mutants, except that containing the prematurely truncated Fur, produced protein by Western blot (immunoblot) analysis, although several had substantially smaller amounts of Fur and two made an immunoreactive protein that migrated more rapidly on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Nine of the 11 point mutations altered amino acids that are identical in all of the fur genes sequenced so far, suggesting that these amino acids may play important structural or functional roles in Fur activity. Eight of the point mutations occurred in the amino-terminal half of Fur, which is thought to mediate DNA binding; most of these mutations occurred in conserved amino acids that have been previously suggested to play a role in the interaction between adjacent alpha-helices of the protein. Three of the point mutations occurred in the carboxy-terminal half of Fur, which is thought to bind iron. One mutation at histidine-90 was associated with complete loss of Fur function; this amino acid is within a motif previously suggested as being involved in iron binding by Fur. The fur allele mutant at histidine-90 interfered with iron regulation by wild-type fur in the same cell when the mutant allele was present at higher copy number; wild-type fur was dominant over all other fur mutant alleles studied. These results are analyzed with respect to previous models of the structure and function of Fur as an iron-sensitive repressor.
Assuntos
Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Proteínas Repressoras/genética , Vibrio cholerae/genética , Sequência de Aminoácidos , Sequência de Bases , Primers do DNA/química , DNA Bacteriano/genética , Proteínas de Ligação a DNA/genética , Genes Bacterianos , Ferro/metabolismo , Dados de Sequência Molecular , Mutagênese , Mutação , Relação Estrutura-AtividadeRESUMO
We have identified a gene in Vibrio cholerae (epd) which encodes an erythrose-4-phosphate dehydrogenase activity and is located immediately downstream of an iron-regulated virulence gene, irgA, and immediately upstream of a gene encoding phosphoglycerate kinase (pgk). Expression of epd in V. cholerae is not regulated by iron, nor is it required for virulence in an infant mouse model.
Assuntos
Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Proteínas de Escherichia coli , Genes Bacterianos/genética , Receptores de Superfície Celular , Vibrio cholerae/genética , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Sequência de Bases , Clonagem Molecular , Regulação Bacteriana da Expressão Gênica/fisiologia , Gliceraldeído-3-Fosfato Desidrogenases/genética , Ferro/fisiologia , Dados de Sequência Molecular , Fosfoglicerato Quinase/genética , Mapeamento por Restrição , Homologia de Sequência de Aminoácidos , Vibrio cholerae/enzimologia , Vibrio cholerae/patogenicidade , VirulênciaRESUMO
Iron is an essential nutrient to support the growth of most bacterial species. However, iron is not easily available to microorganisms infecting mammalian hosts, because it is largely sequestered by iron-binding proteins, such as transferrin or lactoferrin, or complexed to heme. In response to environmental iron stress, Vibrio cholerae produces the siderophore vibriobactin as well as a number of iron-induced outer membrane proteins. Previous data on the role of iron acquisition systems for the intraintestinal growth of mucosal pathogens such as V. cholerae are conflicting. In this report, we isolated mutants of V. cholerae with TnphoA fusions in each of viuA, hutA, and irgA, as well as strains mutant in each pair of these genes and all three simultaneously, to analyze the role of these iron-induced outer membrane protein receptors for in vivo growth of V. cholerae. The fusion between hutA and TnphoA in a single copy on the chromosome allowed the study of in vitro regulation of hutA in response to iron, fur, and irgB; transcription of hutA was tightly iron regulated (70-fold) and dependent on a functional Fur but did not require IrgB. To investigate the effects of mutations in these iron-induced outer membrane proteins on in vivo growth, we inoculated ileal loops in a rabbit model of infection. This avoids exposure of organisms to the potential killing effects of gastric acid, allows several logarithmic increases in growth in the in vivo environment, and facilitates direct comparison of multiple strains in the same animal to avoid any differences between animals. We grew each mutant to be tested in competition with the wild-type strain in the same loop, to provide an internal control. We confirmed that the inocula for these experiments were grown under conditions of iron stress prior to in vivo inoculation, by measuring the alkaline phosphatase activity of the iron-regulated fusion in each strain. The results confirmed that mutation of irgA produced a much more substantial in vivo growth defect than mutation of either hutA or viuA alone. Double mutants of irgA with either viuA or hutA, or the strain mutant in all three genes, showed an in vivo growth defect comparable to the strain mutant in irgA only, suggesting that mutation of irgA was the most relevant for in vivo growth. The strain mutant in both hutA and viuA was also markedly impaired for in vivo growth, suggesting that mutation of both of these iron uptake systems simultaneously can also produce a substantial in vivo growth defect.
Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Transporte/metabolismo , Ferro/metabolismo , Proteínas de Membrana Transportadoras , Vibrio cholerae/crescimento & desenvolvimento , Vibrio cholerae/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Proteínas da Membrana Bacteriana Externa/genética , Transporte Biológico Ativo , Proteínas de Transporte/genética , Expressão Gênica , Genes Bacterianos , Heme/metabolismo , Masculino , Camundongos , Mutagênese Insercional , Mutação , Coelhos , Vibrio cholerae/genéticaRESUMO
Epidemic strains of Vibrio cholerae O1 are divided into two biotypes, classical and El Tor. In both biotypes, regulation of virulence gene expression depends on a cascade in which ToxR activates expression of ToxT, and ToxT activates expression of cholera toxin and other virulence genes. In the classical biotype, maximal expression of this ToxR regulon in vitro occurs at 30 degrees C at pH 6.5 (ToxR-inducing conditions), whereas in the El Tor biotype, production of these virulence genes only occurs under very limited conditions and not in response to temperature and pH; this difference between biotypes is mediated at the level of toxT transcription. In the classical biotype, two other proteins, TcpP and TcpH, are needed for maximal toxT transcription. Transcription of tcpPH in the classical biotype is regulated by pH and temperature independently of ToxR or ToxT, suggesting that TcpP and TcpH couple environmental signals to transcription of toxT. In this study, we show a near absence of tcpPH message in the El Tor biotype under ToxR-inducing conditions of temperature and pH. However, once expressed, El Tor TcpP and TcpH appear to be as effective as classical TcpP and TcpH in activating toxT transcription. These results suggest that differences in regulation of virulence gene expression between the biotypes of V. cholerae primarily result from differences in expression of tcpPH message in response to environmental signals. We present an updated model for control of the ToxR virulence regulon in V. cholerae.
Assuntos
Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/genética , Proteínas de Fímbrias , Proteínas de Membrana , Óperon , Regulon , Fatores de Transcrição/genética , Transcrição Gênica , Vibrio cholerae/genética , Concentração de Íons de Hidrogênio , Temperatura , Vibrio cholerae/patogenicidadeRESUMO
Part I of this study explores whether clothing today contains formaldehyde levels likely to cause contact allergy in formaldehyde-allergic patients. Part II of this study examines whether current reactions to textiles may be due to allergy to textile resins and whether individuals with formaldehyde-related textile allergy will react to the newer low formaldehyde resins used in the textile industry. Part I: free formaldehyde was measured in 16 fabric specimens produced in the US and overseas. Additionally, since the textile industry has moved to the use of newer methods for measuring fabric formaldehyde content, the newer methodology was compared with the older methods used in the medical literature. Part II: 10 subjects with known textile contact allergy were patch tested to available Chemotechnique textile resins and 6 new low-formaldehyde resins used by the textile industry. Part I: 8 fabric specimens yielded no detectable formaldehyde and 7 specimens yielded <200 ppm free formaldehyde, using Schiff's reagent and Merck testing methods. 1 specimen showed approximately 2000 ppm formaldehyde, as measured by the Merck test, but only 24 ppm free formaldehyde when retested by the method described in Japanese Law #112. Part II: all subjects reacted strongly to formaldehyde and DMDHEU (the predominant resin currently used in textiles). 6 subjects reacted to EUMF. 2 subjects had mild reactions to the newer low-formaldehyde resins and 1 to the non-formaldehyde Fixapret NF. Our results suggest that most clothing today yields free formaldehyde levels unlikely to cause contact allergy in formaldehyde-allergic individuals. Japanese method #112 is the recommended methodology to measure free formaldehyde in future studies. DMDHEU may now represent the main cause of textile allergy and may be a better screen than EUMF for this problem. Newer resins yielding fabrics with <75 ppm free formaldehyde may cause occasional reactions, but are more likely to be tolerated by individuals with textile contact allergy. Treatment of these individuals should be directed at identification of reliable sources of garments utilizing these newer resins.
Assuntos
Dermatite Alérgica de Contato/etiologia , Formaldeído/efeitos adversos , Formaldeído/análise , Têxteis/efeitos adversos , Adulto , Idoso , Dermatite Alérgica de Contato/diagnóstico , Dermatite Alérgica de Contato/prevenção & controle , Feminino , Formaldeído/análogos & derivados , Humanos , Imidazóis/administração & dosagem , Imidazóis/efeitos adversos , Masculino , Pessoa de Meia-Idade , Resinas Vegetais/administração & dosagem , Resinas Vegetais/efeitos adversos , Indústria Têxtil , Têxteis/análise , Estados UnidosRESUMO
We examined the utility of a PCR-based method termed cleaved amplified polymorphic sequences (CAPS) to type 35 well-characterized isolates of Staphylococcus epidermidis. The results were compared with detailed epidemiologic information and typing obtained by using pulsed-field gel electrophoresis (PFGE). To identify CAPS markers for this study, eight pairs of oligonucleotide primers corresponding to five previously sequenced S. epidermidis genes were synthesized and then used to amplify DNA sequences from the S. epidermidis strains by using PCR. Amplified products were reproducibly obtained for seven of eight primer pairs from chromosomal DNA of 33 of the 35 isolates. Seven restriction site polymorphisms were found in five of the amplified products when they were subjected to digestion with a panel of restriction endonucleases. Each fragment-enzyme combination that was polymorphic demonstrated only two alleles in the 33 S. epidermidis isolates analyzed, corresponding to the presence or absence of a single restriction site. Overall, five distinct combinations of alleles were detected and were designated CAPS types A through E. There was a close correlation between the CAPS grouping, the epidemiologic information for the strains, and grouping by PFGE following SmaI digestion of chromosomal DNA. Although PFGE analysis was more discriminatory than typing based on the limited number of CAPS markers used in this study (isolates from the same CAPS group were sometimes distributed into more than one PFGE group), no isolates from the same PFGE group were found in more than one CAPS group. The CAPS procedure was highly reproducible, in contrast to published experience with arbitrarily primed PCR. These preliminary data suggest that CAPS represents a PCR-based technique for strain typing that is highly reproducible, rapid, utilizes widely available technologies, and provides results that are relatively easy to interpret and express.
Assuntos
Técnicas de Tipagem Bacteriana , Reação em Cadeia da Polimerase/métodos , Polimorfismo Genético , Staphylococcus epidermidis/classificação , Staphylococcus epidermidis/genética , Técnicas de Tipagem Bacteriana/estatística & dados numéricos , Sequência de Bases , Primers do DNA/genética , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Eletroforese em Gel de Campo Pulsado , Estudos de Avaliação como Assunto , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/estatística & dados numéricos , Reprodutibilidade dos Testes , Especificidade da Espécie , Staphylococcus epidermidis/isolamento & purificaçãoRESUMO
OBJECTIVE: The Ciba Corning 512 coagulation monitor (CC512) can be used to monitor heparin therapy by performing an activated partial thromboplastin time (APTT) at the patient's bedside. This study was designed to compare the CC512 results to results using the laboratory system. The relative sensitivities of both systems to the effect of oral anticoagulant therapy also was investigated. METHODS: Activated partial thromboplastin times were performed with both the CC512 and laboratory system on 74 specimens from patients receiving i.v. heparin therapy, and on 14 specimens from patients on warfarin only. Heparin assays were performed on 43 of the specimens from the heparinized patients. RESULTS: When a patient was receiving heparin only, the APTT results of the CC512 proved to be similar to existing laboratory methods. The CC512 APTT results of patients on warfarin only were markedly prolonged, whereas the laboratory APTTs were only slightly affected. CONCLUSION: The CC512 results were comparable to the laboratory system. However, the CC512 APTT was more sensitive to the effect of warfarin than the laboratory APTT system used in this study. CC512 APTT results on a patient receiving both oral and intravenous anticoagulation could be misleading.
Assuntos
Coagulação Sanguínea , Heparina/uso terapêutico , Monitorização Fisiológica/instrumentação , Administração Oral , Coagulação Sanguínea/efeitos dos fármacos , Heparina/administração & dosagem , Humanos , Tempo de Tromboplastina Parcial , Varfarina/uso terapêuticoRESUMO
Clostridium difficile causes pseudomembranous colitis through the action of Rho-modifying proteins, toxins A and B. Antibodies directed against C. difficile toxin A prevent or limit C. difficile-induced colitis. We engineered plasmid pETR14, containing the hlyB and hlyD genes of the Escherichia coli hemolysin operon, to express a fusion protein containing 720 amino acid residues from the nontoxic, receptor-binding, carboxy terminus of C. difficile toxin A and the secretion signal of E. coli hemolysin A. We introduced pETR14 into Vibrio cholerae and found that the toxin A-HlyA fusion protein was secreted by a number of V. cholerae strains and recognized by both monoclonal and polyclonal anti-C. difficile toxin A antibodies. We introduced pETR14 into an attenuated V. cholerae strain, O395-NT, and inoculated rabbits orally with this construct. Colonization studies disclosed that the V. cholerae vector containing pETR14 was recoverable from rabbit ilea up to 5 days after oral inoculation. Vaccination produced significant systemic anti-C. difficile toxin A immunoglobulin G and anti-V. cholerae vibriocidal antibody responses. Vaccination also produced significant protection against toxin A in an ileal loop challenge assay, as assessed by determination of both fluid secretion and histological changes. These results suggest that the hemolysin system of E. coli can be used successfully in V. cholerae vector strains to effect secretion of large heterologous antigens and that a V. cholerae vector strain secreting a nontoxic, immunogenic portion of C. difficile toxin A fused to the secretion signal of E. coli HlyA induces protective systemic and mucosal immunity against this toxin.