Analysis of the genomic sequence of a human metapneumovirus

We recently described the isolation of a novel paramyxovirus from children with respiratory tract disease in The Netherlands. Based on biological properties and limited sequence information the virus was provisionally classified as the first nonavian member of the Metapneumovirus genus and named human metapneumovirus (hMPV). This report describes the analysis of the sequences of all hMPV open reading frames (ORFs) and intergenic sequences as well as partial sequences of the genomic termini. The overall percentage of amino acid sequence identity between APV and hMPV N, P, M, F, M2-1, M2-2, and L ORFs was 56 to 88%. Some nucleotide sequence identity was also found between the noncoding regions of the APV and hMPV genomes. Although no discernible amino acid sequence identity was found between two of the ORFs of hMPV and ORFs of other paramyxoviruses, the amino acid content, hydrophilicity profiles, and location of these ORFs in the viral genome suggest that they represent SH and G proteins. The high percentage of sequence identity between APV and hMPV, their similar genomic organization (3'-N-P-M-F-M2-SH-G-L-5'), and phylogenetic analyses provide evidence for the proposed classification of hMPV as the first mammalian metapneumovirus.     

Symptomatic peripheral arterial disease: the value of a validated questionnaire and a clinical decision rule

BACKGROUND: If a validated questionnaire, when applied to patients reporting with symptoms of intermittent claudication, could adequately discriminate between those with and without peripheral arterial disease, GPs could avoid the diagnostic measurement of the ankle brachial index. AIM: To investigate the Edinburgh Claudication Questionnaire (ECQ) in general practice and to develop a clinical decision rule based on risk factors to enable GPs to easily assess the likelihood of peripheral arterial disease. DESIGN OF STUDY: An observational study. SETTING: General practice in The Netherlands. METHOD: This observational study included patients of > or =55 years visiting their GP for symptoms suggestive of intermittent claudication or with one risk factor. The ECQ and the ankle brachial index were performed. The prevalence of peripheral arterial disease, defined as an ankle brachial index <0.9, was related to risk factors using logistic regression analyses, on which a clinical decision rule was developed and related to the presence of peripheral arterial disease. RESULTS: Of the 4790 included patients visiting their GP with symptoms suggestive of intermittent claudication, 4527 were eligible for analyses. The prevalence of peripheral arterial disease in this group was 48.3%. The sensitivity of the ECQ was only 56.2%. The prevalence of peripheral arterial disease in a clinical decision rule that included age, male sex, smoking, hypertension, hypercholesterolemia, and a positive ECQ, increased from 14% in the lowest to 76% in the highest category. CONCLUSION: This study indicates that the ECQ alone has an inadequate diagnostic value in detecting patients with peripheral arterial disease. The ankle brachial index should be performed to diagnose peripheral arterial disease in patients with complaints suggestive of intermittent claudication, although our clinical decision rule could help to differentiate between extremely high and lower prevalence of peripheral arterial disease.     

How can the evaluation of genetic tests be enhanced? Lessons learned from the ACCE framework and evaluating genetic tests in the United Kingdom.

Advances in genetic technology are increasing the availability of genetic tests, not only for rare single gene disorders, but also for common diseases such as breast and colo-rectal cancer. Before there can be widespread uptake of these tests, they must be evaluated to confirm the benefits of their use. But how should genetic tests be evaluated, given the speed at which new tests are emerging? One highly influential approach is the analytic validity, clinical validity, clinical utility and ethical, legal and social issues (ACCE) framework, which has provided a benchmark for the evaluation of genetic tests. The approach has been adopted and adapted by the United Kingdom Genetic Testing Network, with the help of the Public Health Genetics Unit in Cambridge, to evaluate new genetic tests for use in the National Health Service. We discuss a number of conceptual, methodological, and practical issues concerning the evaluation of genetic tests, based on lessons learned from applying the ACCE framework and from the UK experience, and make a number of recommendations to further strengthen the evaluation of genetic tests.     

Genomic instability occurs in colorectal carcinomas but not in adenomas

Genomic instability, as demonstrated by the presence of additional alleles at short tandemly repeated (STR) loci, has recently been observed in colorectal tumours from individuals with hereditary non-polyposis colorectal cancer (HNPCC), and in some sporadic tumours. These neoplasms have been called replication error positive (RER⁺). In this study, we confirm the presence of genomic instability in a proportion of unselected colorectal carcinomas but find no evidence of instability in adenomas. We further report replication errors in a tetranucleotide sequence, and in STRs within two tumour suppressor genes. 108 colorectal adenocarcinomas and 46 adenomas were analysed for the presence of variant bands at 4–15 microsatellite markers. Seven (6.5%) of carcinomas were RER⁺, four of which originated from the proximal colon. Analysis of the adenomas and of matched adenoma-carcinoma and carcinoma-metastatic samples from four patients suggests that the replication errors may occur during the development of carcinomas but are rare in adenomas. © 1993 Wiley-Liss, Inc.     

CD28-dependent killing by human YT cells requires phosphatidylinositol 3-kinase activation

CD28/B7 interactions have been demonstrated to provide a co-stimulatory signal for the generation of CD8⁺ cytotoxic T lymphocytes in the absence of CD4⁺ T helper cells. The CD28 signals required for induction of cytotoxicity have yet to be described. To investigate further the biochemical signaling pathways associated with CD28-dependent cytotoxicity, we have studied the human thymic leukemia cell line, YT. YT cells kill B7⁺ targets in a non-major histocompatibility complex (MHC)-restricted, CD28-dependent manner. CD28 ligation on the surface of YT cells caused a rapid increase in the tyrosine phosphorylation of four major cellular substrates with masses estimated to be 110, 95, 85, and 44 kDa. The 110 and 85 kDa substrates were identified as the catalytic and regulatory subunits, respectively, of phosphatidylinositol 3-kinase (PI3-K). Engagement of CD28 caused the rapid receptor association and activation of PI3-K but did not activate phospholipase C_γ. CD28-induced tyrosine phosphorylation and PI3-K activation was independent of p56^lck protein tyrosine kinase (PTK) activity (previously reported to be associated with CD28) and was insensitive to inhibition by the PTK inhibitor herbimycin A. Two structurally and mechanistically dissimilar inhibitors of PI3-K, wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002) also failed to block CD28-dependent tyrosine phosphorylation events or the association of PI3-K with the CD28 receptor. However, both drugs inhibited CD28-dependent cytotoxicity and CD28 receptor associated PI3-K activity with IC₅₀ values similar to the reported IC₅₀ values for PI3-K inhibition. Although herbimycin A did not significantly block the observed CD28-dependent tyrosine phosphorylation or PI3-K activation, herbimycin did block CD28-dependent cytotoxicity in a dose-dependent manner. These data support a role for PI3-K activation in the CD28-dependent initiation of cytotoxic effector function and suggest that a herbimycin sensitive step(s) is either CD28-independent, resides within a PI3-K-independent CD28 signaling pathway, or is downstream of CD28-dependent PI3-K activation.     

A meta-analytic review of HIV behavioral interventions for reducing sexual risk behavior of men who have sex with men

This meta-analysis examines the efficacy of international HIV prevention interventions designed to reduce sexual risk behavior of men who have sex with men (MSM). We performed a comprehensive search of published and unpublished English-language reports of HIV prevention interventions that focus on MSM and evaluated changes in risky sexual behavior or biologic outcomes related to sexual risk. Data from 33 studies described in 65 reports were available as of July 2003. Studies with insufficient data to calculate effect sizes were excluded from the meta-analysis. Interventions were associated with a significant decrease in unprotected anal intercourse (odds ratio [OR] = 0.77, 95% confidence interval [CI]: 0.65-0.92) and number of sexual partners (OR = 0.85, 95% CI: 0.61-0.94) and with a significant increase in condom use during anal intercourse (OR = 1.61, 95% CI: 1.16-2.22). Interventions successful in reducing risky sexual behavior were based on theoretic models, included interpersonal skills training, incorporated several delivery methods, and were delivered over multiple sessions spanning a minimum of 3 weeks. Behavioral interventions provide an efficacious means of HIV prevention for MSM. To the extent that proven HIV prevention interventions for MSM can be successfully replicated in community settings and adapted and tailored to different situations, the effectiveness of current HIV prevention efforts can be increased.     

Screening for Escherichia coli O157:H7 in Illinois

Objective: To determine the incidence of infection with Escherichia coli O157:H7 in a tertiary referral center in Chicago, where a similar study had been performed in 1984, to evaluate cases of disease reported to the Illinois Department of Public Health (IDPH) in 1993, and to determine laboratory practices used to detect this infection throughout the state.
Methods: During a 6-month period in 1993, all stool specimens at Rush-Presbyterian-St Luke's Medical Center (RPSLMC) were tested for E. coli O157:H7. Reports of diagnosed E. coli O157:H7 cases investigated by IDPH were also reviewed. A survey of 73 hospitals in the Chicago area was performed to determine routine culturing practices, specifically, the selection of stool specimens for evaluation for this pathogen.
Results: In the RPSLMC survey, two cases were identified among 1985 samples (incidence 0.1%), similar to the 0.08% incidence detected in a similar study conducted at the same institution in 1984. Through passive surveillance, the IDPH received 44 reports of E. coli O157:H7 in 1993. The hospital survey revealed that, in the seven labs testing all stool specimens for E. coli O157:H7, an incidence of 16/8137 specimens (0.2%) was determined.
Conclusions: These data suggest that sporadic E. coli O157:H7 remains uncommon in Illinois and that the incidence may not have changed over a 9-year period. The low yield and substantial cost of culturing all stools suggest that only specimens from patients with bloody diarrhea should be evaluated routinely in areas of low endemicity.     

Minimal α1- and α2-adrenoceptor-mediated coronary vasoconstriction in the anaesthetized swine

Summary -Adrenoceptor-mediated coronary vasoconstriction contributes to the initiation and aggravation of experimental and clinical myocardial ischaemia. However, the extent of ₁- and ₂-adrenoceptor-mediated constriction has not been characterized in the porcine coronary circulation despite the frequent use of this experimental model.Fifteen swine were anaesthetized with either -chloralose, enflurane or isoflurane to determine the amount of -adrenoceptor-mediated coronary constriction elicited by either the selective ₁-adrenoceptor agonist methoxamine or the selective ₂-adrenoceptor agonist azepexole. The left anterior descending coronary artery was cannulated and perfused by an external pump delivering constant blood flow from the carotid artery. Following bilateral cervical vagotomy and ß-adrenoceptor blockade with propranolol (2 mg kg^–1), graded dosages of either one of the -adrenoceptor agonists (9–45 g kg^–1 min^–1) were infused into the coronary perfusion line while coronary arterial pressure (CAP) was measured through a distal side arm of the cannula to detect changes in coronary vascular resistance. Infusion of the -adrenoceptor agonists was terminated when systemic arterial pressure increased. Sonomicrometers were used to measure anterior left ventricular wall thickening for the assessment of regional contractile function. During methoxamine infusion, no increase in vascular resistance was observed during -chloralose, enflurane or isoflurane anaesthesia, whereas the infusion of azepexole increased CAP from 103 ± 31 mmHg to 120 ± 35 mmHg (-chloralose), from 101 ± 16 mmHg to 122 ± 11 mmHg (enflurane) and from 84 ± 20 mmHg to 94 ± 19 mmHg (isoflurane), respectively. In four additional swine anaesthetized with enflurane, the intracoronary infusion of the full catecholamine agonist noradrenaline in the presence of propranolol (6 mg kg^–1) increased CAP from 98 ± 10 to 105 ± 10 mmHg prior to an increase in regional left ventricular function or systemic arterial pressure.These results indicate that there are no ₁- and relatively little ₂-adrenoceptor-mediated coronary constrictive effects in swine. Furthermore, neither -adrenoceptor agonist produced any detectable change in regional myocardial contractile function, regardless of the anaesthesia used.Supported by the German Research Foundation (He 1320/3-2). Dr. Guth is the recipient of a scholarship from the Alexander von Humboldt-Foundation. Send offprint requests to G. Heusch at the above address