How to switch the motor on: RNA polymerase initiation steps at the single-molecule level.

RNA polymerase (RNAP) is the central motor of gene expression since it governs the process of transcription. In prokaryotes, this holoenzyme is formed by the RNAP core and a sigma factor. After approaching and binding the specific promoter site on the DNA, the holoenzyme-promoter complex undergoes several conformational transitions that allow unwinding and opening of the DNA duplex. Once the first DNA basepairs (∼10 bp) are transcribed in an initial transcription process, the enzyme unbinds from the promoter and proceeds downstream along the DNA while continuously opening the helix and polymerizing the ribonucleotides in correspondence with the template DNA sequence. When the gene is transcribed into RNA, the process generally is terminated and RNAP unbinds from the DNA. The first step of transcription-initiation, is considered the rate-limiting step of the entire process. This review focuses on the single-molecule studies that try to reveal the key steps in the initiation phase of bacterial transcription. Such single-molecule studies have, for example, allowed real-time observations of the RNAP target search mechanism, a mechanism still under debate. Moreover, single-molecule studies using Förster Resonance Energy Transfer (FRET) revealed the conformational changes that the enzyme undergoes during initiation. Force-based techniques such as scanning force microscopy and magnetic tweezers allowed quantification of the energy that drives the RNAP translocation along DNA and its dynamics. In addition to these in vitro experiments, single particle tracking in vivo has provided a direct quantification of the relative populations in each phase of transcription and their locations within the cell.

Correlation of cancer risk evaluation and early detection (CADET) scores with abnormal ultrasonographic ovarian findings.

OBJECTIVE: To determine the utility of a modified version of ovarian cancer-focused cancer risk evaluation and early detection (CADET) scores as a screening tool for ultrasonographic ovarian findings. STUDY DESIGN: Prospective pilot study. MAIN OUTCOME MEASURES: CADET scores were compared with abnormal ultrasonographic ovarian findings of peri- and postmenopausal women who attended their gynecologist for a routine check-up. The women filled in the CADET questionnaire before seeing their gynecologists who were blinded to the CADET results. The women whom they referred for pelvic transvaginal ultrasonographic examination comprised the study group. The results of their scans were compared with their CADET scores. RESULTS: Of the 181 peri- and postmenopausal women who were candidates for this study, 154 were referred for ultrasonography, of whom 38 (24%, Group A) had abnormal ovarian scans (30 simple cysts and 8 complex findings). The other 116 (76%) women had normal sonograms (Group B). Demographic characteristics were similar for both groups. Thirteen Group A women (34%) and 52 Group B women (45%) had positive CADET scores (p = NS). The average group CADET scores were also not significantly different (0.8 +/- 1.7 for Group A and 1.7 +/- 2.5 for Group B). CONCLUSION: CADET scores did not correlate with abnormal ultrasonographic ovarian findings.

Improvement of detection of bacterial pathogens in normally sterile body sites with a focus on orthopedic samples by use of a commercial 16S rRNA broad-range PCR and sequence analysis.

A new commercially available universal 16S and 18S rRNA gene PCR test, which is followed by sequence analysis of amplicons (SepsiTest), was evaluated for rapid identification of pathogens in the diagnosis of bone and joint infections. Eighty-three orthopedic samples and 21 specimens from other normally sterile body sites collected from 84 patients were analyzed in parallel by culture and PCR for detection of bacteria and fungi. Compared to culture, the diagnostic sensitivity and specificity of PCR were 88.5% and 83.5%, respectively. The detection rate of PCR (34.6%) was higher than that of bacterial culture (25.0%) as a consequence of the presence of fastidious and noncultivable species in samples and antibiotic treatment of patients. Thirteen culture-negative infections were identified by PCR, and PCR was able to detect culture-proven polymicrobial infections. On the other hand, three samples were culture positive but PCR negative. SepsiTest was demonstrated to be a valuable supplemental tool in the rapid detection of bacteria, especially for fastidious and noncultivable organisms, allowing earlier initiation of pathogen-adapted therapy in patients with bone and joint infections.

Congestive heart failure in pregnancy: a case of peripartum cardiomyopathy.

A healthy 28-year-old woman developed full-blown pulmonary oedema in the 36th week of gestation. Echocardiography revealed a globally enlarged heart with reduced systolic function. A remarkable dinical response with regain of normal ventricular function was noted with early medical intervention. This case report illustrates peripartum cardiomyopathy, a unique form of dilated cardiomyopathy affecting women during/following gestation. Clinician familiarity with this entity increases the probability of prompt appropriate treatment, offering patients the best possible prognosis.

Serotonin (5-HT) enhances the activity of amphotericin B against Aspergillus fumigatus in vitro.

We investigated the in vitro synergistic antifungal potential of combining serotonin (5-HT) and sertraline with amphotericin B and itraconazole against clinical isolates of Aspergillus spp. Synergy tests were performed using the chequerboard microdilution method. Activity was measured against Aspergillus fumigatus (n = 7), Aspergillus flavus (n = 3) and Aspergillus terreus (n = 2), and compared with that for Candida albicans and Candida parapsilosis. The fractional inhibitory concentration (FIC) indices ranged between 0.25 and 3 for the various isolates tested. 5-HT was shown to enhance the activity of amphotericin B against Aspergillus spp. Combination studies with 5-HT and itraconazole and with sertraline and itraconazole or amphothericin B showed different activities for the various strains, including synergism (FIC < 1.0), additivity (FIC = 1), and indifference (FIC between 1.0 and 2.0). 5-HT and sertraline showed antagonistic activity (FIC > 2) with amphotericin B and itraconazole against C. parapsilosis.

TB or not TB: cavitary bronchiolitis obliterans organizing pneumonia mimicking pulmonary tuberculosis.

Two patients with subacute symptoms and signs compatible with pulmonary tuberculosis (TB) had right upper lobe cavitary infiltrates shown on chest radiography. In both patients, purified protein derivative and microbiologic testing excluded TB, and tissue examination yielded typical histologic changes of bronchiolitis obliterans organizing pneumonia (BOOP). Glucocorticoid therapy led to clinical and radiologic resolution. Though probably rare in this situation, BOOP should be considered in the differential diagnosis of patients presenting with clinical and radiologic features of pulmonary TB.

The value of stress thallium-201 single photon emission CT imaging as a predictor of outcome and long-term prognosis after CABG.

This study examined the capability of rest-redistribution 201Tl single photon emission CT imaging (TSPECT) done shortly after coronary artery bypass grafting (CABG) to predict outcome and long- term prognosis. Results of TSPECT at 5.5+/-0.7 (range 4 - 8) months postoperatively were correlated during a 4-year follow-up period wit mortality, major surgical and nonsurgical cardiac events and cardiac event-free survival in 170 patients who underwent CABG. Ten (5.8%) patients died during follow-up: 4 had a documented cardiac death (all of them with large or moderate reversible filling defects). In the survivors, there were 10 nonfatal myocardial infarctions and 11 revascularization interventions. Patients who eventually required and underwent revascularization procedures demonstrated large or moderate reversible filing defects, in contrast with those who had no need for an additional procedure (15.7% versus 2.5%, p < 0.001). Postoperative TSPECT carried out soon after CABG had definite prognostic value and should be performed routinely to help decide treatment protocol.

Dysphagia with multiple autoimmune disease.

Myasthenia gravis (MG) and polymyositis (PM) are organ-specific autoimmune diseases. Occasional reports describe patients with clinical and pathologic features of both. Achalasia is idiopathic in nature, but autoimmune and inflammatory mechanisms have been proposed for this disorder as well. We describe a patient with dysphagia who was diagnosed at different points in time with all these three rare conditions. Despite at least putatively having immune mechanisms in common, an association between the three has not been previously described.

The impact of myocardial viability as determined by rest-redistribution 201Tl single photon emission CT imaging and the choice of therapy on prognosis in patients with left ventricular dysfunction.

While thallium-201 (201Tl) single-photon emission CT (TSPECT) scintigraphy is a commonly used method for determining the viability of the myocardium, its value for predicting outcome is limited. The diagnosis of myocardial viability in patients with ischemic systolic left ventricular dysfunction might indicate which of them will benefit more from surgical revascularization. Forty patients (mean +/- SD aged 64.5 +/- 12 years, 33 males and 7 females) with impaired systolic left ventricular function (ejection fraction < or = 45%) underwent TSPECT examination. Twenty patients were surgically revascularized and 20 were treated medically. The patients were followed-up for a 34 +/- 10-month period and the cardiac long-term prognosis was evaluated. The significant viability percentage (SVP), defined as the percentage of the total number of segments showing a normal uptake of 201Tl redistribution divided by the total number of segments evaluated, was > or =55: this was observed in 18 patients. Among them, the cardiac event-free survival was 100% in the surgical group versus 22% in the medical group. However, in patients with non-SVP, the survival was lower and not significantly different in the two treatment groups. Surgical revascularization is the preferred method of treatment in patients with ischemic systolic left ventricular dysfunction and myocardial viability as defined by TSPECT scintigraphy.

Graphic representation of sequential Bayesian analysis.

Previously undescribed algebraic transforms of Bayes' theorem define families of operating points in the receiver operating characteristic (ROC) space which, at given pre-test probability, produce constant post-test probabilities. These isopredictive operating points form straight lines in the ROC space. The lines can be used to emulate Bayesian sequential analysis in a strictly graphic procedure, which can be applied in clinical work and medical education.

Long-term follow-up after coronary artery bypass grafting reoperation.

BACKGROUND: Coronary artery bypass grafting (CABG) reoperation is being performed with increasing frequency. OBJECTIVE: To determine the clinical outcome and the long-term results of a second CABG. SETTING: An 1100-bed urban university-affiliated hospital. DESIGN: Retrieval of data on selected parameters from medical records before surgery and prospective follow-up afterwards. PATIENTS AND METHODS: We studied the outcomes of 498 consecutive patients who underwent CABG reoperation in our institution from January 1978 to December 1989 and who were followed postoperatively. Their perioperative mortality, morbidity, and long-term follow-up results were re-evaluated. The end points of the study were December 1997, 15 years of follow-up, or the patient's death. RESULTS: The perioperative mortality rate was 3%. The cumulative survival rates were 90.1%, 74%, and 63.4% at the 5-year, 10-year, and 15-year follow-ups, respectively. The cardiac event-free survival rates were 91.5%, 83.4%, and 67.8% at the 5-year, 10-year, and 15-year follow-ups, respectively. The risk factors adversely affecting long-term survival were advanced age, hypertension, and a low left ventricular ejection fraction (LVEF). CONCLUSIONS: The long-term results of cumulative survival and cardiac event-free survival in patients who underwent CABG reoperation are good. Although this reoperation is safe overall, advanced age, hypertension, and a decreased LVEF significantly increase the surgical risk.

CaDet, a computer-based clinical decision support system for early cancer detection.

Cancer risk evaluation and early detection are subject to serious limitations mainly related to human factors and to characteristics of the data involved. To help overcome these problems, a computer-based system was designed to provide the physician with a clearer clinical picture and aid in directing patients to appropriate measures. Clinical and epidemiological data related to early cancer detection and to cancer risk factors was collected from the literature and incorporated in a database, together with heuristic rules for evaluating this data. Individual data obtained from patients through a questionnaire are input into CaDet, a computerized clinical decision support system. A report summarizing patient data and cancer hypotheses, with a scoring system that reflects degrees of alarm, is generated. The CaDet system, as well as some preliminary results of the clinical experience accumulated in its use, are described. These preliminary results suggest that the approach may be useful in improving cancer risk assessment and screening in primary care setups.

Short- and long-term follow-up after coronary bypass grafting for single-vessel coronary artery disease.

Short-term outcome and 10-year clinical outcome were reviewed in 114 consecutive patients after coronary artery bypass grafting (CABG) for single-vessel coronary artery disease (CAD). Gated equilibrium radionuclide cineangiography was performed soon after CABG in all cases, and revealed very good early graft patency rates. There was no perioperative mortality, and very low morbidity. During follow-up there were seven late deaths, two from cardiac disease and five from non-cardiac causes. Cumulative survival at 10 years was 93%. Cumulative freedom from additional cardiac invasive procedures was 96%, 93% and 80% at 1, 5, and 10 years, respectively, and cumulative freedom from angina was 93%, 80% and 73%. Conventional single-vessel CABG thus can be safely performed, with minimal postoperative morbidity and no mortality, providing good long-term relief of angina and circumventing need for additional invasive procedures.

Impact of early exercise radionuclide cineangiography on long-term prognosis after CABG.

BACKGROUND: The immediate benefits of coronary artery bypass grafting might be only transient. This prospective study examined the capability of exercise radionuclide cineangiography done shortly after coronary artery bypass grafting to predict outcome and long-term prognosis. METHODS: Results of exercise radionuclide cineangiography at 5.5 +/- 0.7 months (range, 4 to 8 months) postoperatively were correlated with mortality, major surgical and nonsurgical cardiac events, and cardiac event-free survival in 100 consecutive patients who underwent coronary artery bypass grafting. Stepwise logistic regression analysis was used to evaluate the incremental value of radionuclide cineangiography beyond the commonly used variables. RESULTS: Left ventricular ejection fraction at rest was normal (> or = 0.45) in 72 patients and increased on exercise in 58. The exercise radionuclide variables that correlated with future cardiac events were change and fractional change in heart rate, ST segment changes, anginal pain and congestive heart symptoms during exercise, rest ejection fraction, and change and fractional changes in ejection fraction. Predictors of event-free survival were exercise heart rate, rest ejection fraction, and change and fractional change in ejection fraction during exercise. Logistic regression analysis revealed that change in ejection fraction was an independent predictor of cardiac death and surgical interventions, whereas resting ejection fraction was a predictor of nonsurgical cardiac events. CONCLUSIONS: Postoperative exercise radionuclide cineangiography carried out soon after coronary artery bypass grafting had definite independent prognostic value and should be performed routinely to help decide treatment protocol.

Very early onset of acute amiodarone pulmonary toxicity presenting with hemoptysis.

A case of amiodarone pulmonary toxicity (APT) occurring within days of the start of therapy and presenting with hemoptysis is reported. Both the very early onset and the presenting symptom are highly unusual in this entity. Implications in the diagnosis of APT are discussed.