Antigenic and genetic analyses of isolate APMV/wigeon/Italy/3920-1/2005 indicate that it represents a new avian paramyxovirus (APMV-12).

Isolate wigeon/Italy/3920-1/2005 (3920-1) was obtained during surveillance of wild birds in November 2005 in the Rovigo province of Northern Italy and shown to be a paramyxovirus. Analysis of cross-haemagglutination-inhibition tests between 3920-1 and representative avian paramyxoviruses showed only a low-level relationship to APMV-1. Phylogenetic analysis of the whole genome and each of the six genes indicated that while 3920-1 grouped with APMV-1 and APMV-9 viruses, it was quite distinct from these two. In the whole-genome analysis, 3920-1 had 52.1 % nucleotide sequence identity to the closest APMV-1 virus, 50.1 % identity to the APMV-9 genome, and less than 42 % identity to representatives of the other avian paramyxovirus groups. We propose isolate wigeon/Italy/3920-1/2005 as the prototype strain of a further APMV group, APMV-12.

Quantitative assessment of the extent of myocardial infarction in the conscious dog by means of analysis of serial changes in serum creatine phosphokinase activity.

This study was designed to develop a method for quantitative assessment of infarct size in the conscious animal based on serial changes of serum creatine phosphokinase (CPK) activity. From 11 experiments in which myocardial CPK was injected intravenously in conscious dogs, the average CPK distribution space and average CPK fractional disappearance rate from serum were found to be 11.4% of body weight and 0.48% min respectively. In other experiments, myocardial infarction was produced in 22 conscious dogs by constriction of a left coronary artery snare and serum CPK activity was determined at frequent intervals for 24 hr. Since myocardial CPK depletion reflects infarct size, infarct size was determined directly by analysis of myocardial CPK content in the same animals 24 hr after coronary artery occlusion. CPK released from the infarct was determined from observed changes in serum CPK activity analyzed according to a model taking into account the fraction of CPK released from an infarct and the rates of appearance and disappearance of CPK activity from serum. Infarct size was calculated on the basis of observed changes in serum CPK and compared to infarct size determined directly by analysis of myocardial CPK depletion. Agreement was close and results from all experiments fit the equation: [infarct size (g) determined from serum CPK] = 1.13 x [infarct size (g) determined from myocardial CPK] - 1.3, r = 0.96, n = 22. The method described is useful for accurate assessment of infarct size in the conscious animal and for detection of modification of infarct size produced by pharmacologic interventions.

Early estimation of myocardial damage in conscious dogs and patients with evolving acute myocardial infarction.

To estimate the ultimate extent of myocardial damage during evolving myocardial infarction in conscious dogs and patients, we analyzed early serum creatine phosphokinase (CPK) changes with nonlinear curve-fitting techniques. In experiments with dogs, serial serum CPK changes were fit to a log-normal function by the least squares method; the extent of the completed infarct was calculated by analysis of observed serum CPK changes and verified by measurement of myocardial CPK depletion 24 h after coronary occlusion. Early prediction of myocardial damage was based on projected serum CPK values from best fit curves based on data obtained during the first 5 h after initial elevation of enzyme activity. The correlation between predicted and observed values was close (r > 0.96, n = 11). In 11 additional conscious animals subjected to coronary occlusion, isoproterenol was administered continuously as soon as damage had been estimated from projected serum CPK values. The extent of the completed infarct was assessed by analysis of all serial serum CPK values and verified by analysis of myocardial CPK depletion 24 h after coronary occlusion. In each experiment the calculated completed infarct size exceeded infarct size projected before administration of isoproterenol (average increase = 44+/-10 [SE]%). When similar calculations were applied in experiments with eight dogs treated with propranolol, myocardial salvage was detected in 50% of the animals. In 30 patients with uncomplicated acute myocardial infarction the extent of the completed infarct, measured by analysis of CPK activity in serum samples obtained every 2 h, was compared with damage estimated from CPK values projected by the best fit log-normal curve derived from data obtained during the first 7 h after the initial serum CPK elevation. The estimate of damage based on early data correlated closely with the extent of infarction calculated from all available serial serum CPK values (r = 0.93, n = 30). Thus, the extent of the completed infarct could be estimated accurately during the early evolution of infarction. In patients with spontaneous extension of infarction manifested by chest pain and electrocardiographic changes, the calculated extent of the completed infarct exceeded that predicted. Conversely, salvage of myocardium, after reduction of myocardial oxygen requirements by administration of trimethaphan, was reflected by reduction of the extent of the calculated completed infarct with respect to that predicted from early serum CPK changes.

Coronary artery reperfusion. I. Early effects on local myocardial function and the extent of myocardial necrosis.

The effects of coronary artery reperfusion 3 hr after coronary occlusion on contractile function and the development of myocardial damage at 24 hr was studied experimentally. In 14 control and 6 reperfused dogs, relationships between epicardial ST segment elevation 15 min after coronary occlusion and myocardial creatine phosphokinase activity (CPK) and histologic appearance 24 hr later were examined. The electrocardiograms were recorded from 10 to 15 sites on the left ventricular epicardium and transmural samples for CPK and histology were obtained from the same sites where epicardial electrocardiograms had been recorded. An inverse relation existed between ST segment elevation (mv) 15 min after occlusion and log CPK activity (IU/ mg of protein) 24 hr later, log CPK = - 0.06ST + 1.26. In dogs subjected to coronary artery reperfusion, there was significantly less CPK depression (log CPK = - 0.01ST + 1.31, [P < 0.01]) than that expected from the control group. In the control group 97% of specimens showing ST segment elevations over 2 mv at 15 min showed abnormal histology 24 hr later. In contrast, in the reperfused group 43% of sites exhibiting elevated ST segment at 15 min showed abnormal histology 24 hr later. In six additional dogs it was shown that the paradoxical movement of the left ventricular wall could be reversed within 1 hr of perfusion. Therefore, by enzymatic and histologic criteria, as well as by functional assessment, coronary artery reperfusion 3 hr after occlusion resulted in salvage of myocardial tissue.

Validated H5 Eurasian real-time reverse transcriptase-polymerase chain reaction and its application in H5N1 outbreaks in 2005-2006.

Real time reverse transcriptase (RRT)-polymerase chain reaction (PCR) for the detection of Eurasian H5 avian influenza virus (AIV) isolates was adapted from an existing protocol, optimized, and validated using a number of genetically diverse H5 isolates (n = 51). These included 34 "Asian lineage" H5N1 highly pathogenic avian influenza (HPAI) viruses (2004-2006), plus 12 other H5 isolates from poultry outbreaks and wild birds in the Eastern Hemisphere (1996-2005). All 51 were positive by H5 Eurasian RRT-PCR. Specificity was assessed by testing representative isolates from all other AL virus subtypes (n = 52), non-AI avian pathogens (n = 8), plus a negative population of clinical specimens derived from AI-uninfected wild birds and poultry (n = 604); all were negative by H5 Eurasian RRT-PCR. RNA was directly extracted from suspect HPAI H5N1 clinical specimens (Africa, Asia, and Europe; 2005-2006; n = 58) from dead poultry and wild birds, and 55 recorded as positive by H5 Eurasian RRT-PCR: Fifty-one of these 55 were in agreement with positive AIV isolation in embryonated chickens' eggs. H5 Eurasian RRT-PCR was invaluable in H5 outbreak diagnosis and management by virtue of its rapidity and high degree of sensitivity and specificity. This method provides a platform for automation that can be applied for large-scale intensive investigations, including surveillance.

Recent epidemiology and ecology of influenza A viruses in avian species in Europe and the Middle East.

There have been at least ten distinct outbreaks of LPAI or HPAI in poultry caused by H5 or H7 viruses in the last eight years in Europe and the Middle East. There appears to be an increased occurrence of such episodes consistent with global trends. As a result, surveillance systems have been enhanced to facilitate early detection of infection in poultry, together with active surveillance of wild bird populations. These complementary activities have resulted in the detection of a number of viruses in wild bird populations, including some with high genetic similarity to newly detected viruses in poultry, for example, H7N3 in Italy and H7N7 in the Netherlands. Furthermore, there is evidence for continued circulation of H5 and H7 viruses in wild Anseriformes, thereby presenting a real and current threat for the introduction of viruses to domestic poultry, especially those reared in outdoor production systems. Viruses of H9N2 subtype continue to circulate widely in the Middle East and are associated with significant disease problems in poultry. The epidemiology has the potential to be complicated further by introduction of novel viruses through illegal importation of captive birds, such as was detected with H5N1 in Belgium in 2004. Continual genetic exchange in the avian virus gene pool and independent evolution of all gene segments either within an individual host species or among wild bird hosts suggests that these viruses are not in evolutionary stasis in the natural reservoir.

Evaluation of a radioimmunoassay for alpha 1-acid glycoprotein to monitor therapy of cancer patients.

A new radioimmunoassay for alpha 1-acid glycoprotein (AGP) for monitoring the therapy of cancer patients was evaluated. Plasma levels of this glycoprotein were measured in 49 normal healthy volunteers, 71 patients with illnesses other than cancer, 190 patients with solid tumors, and 58 patients with hematologic neoplasms. Plasma levels of AGP were elevated in 89% of the solid tumor patients and 87% of the patients with hematologic neoplasms who had newly diagnosed, locally recurrent, or metastatic cancer. Only 18% of patients with illnesses other than cancer and normal renal function had elevations of plasma AGP. Serial measurements of AGP may be useful for monitoring therapy in several tumor types, including small-cell lung cancer, colon cancer, lymphoma, and non-small-cell lung cancer.

Prostaglandin E1 coronary venous retroperfusion in acute myocardial ischemia: effects on regional left ventricular function and infarct size.

Prostaglandin E1 was administered by means of coronary venous synchronized retroperfusion and the effectiveness of the combined (prostaglandin-retroperfusion) system was examined during acute myocardial ischemia in 10 closed chest anesthetized dogs. Such treatment was administered between 30 minutes and 3 hours after occlusion of the proximal left anterior descending coronary artery. An equivalent series of 10 dogs with arterial blood retroperfusion alone and 9 untreated dogs served as control subjects. Standardized two-dimensional echocardiographic measurements of global and regional left ventricular function were performed in five short-axis cross sections. The global low left ventricular section and its profoundly ischemic anterolateral region exhibited distinctly improved systolic fractional area changes as a result of the prostaglandin E1 retroperfusion treatment between 30 minutes and 3 hours after occlusion (22.9 +/- 1.5 to 41.2 +/- 4.0% and 1.8 +/- 3.6 to 29.4 +/- 5.6%, respectively). In contrast, further deterioration in function was noted during an untreated equivalent coronary occlusion period (16.3 +/- 2.7 to 10.0 +/- 3.3% and 12.6 +/- 6.1 to 4.1 +/- 6.9%). Although arterial blood retroperfusion alone provided distinct benefits in the ischemic region of a midpapillary echo section (from 13.4 +/- 3.9 to 32.1 +/- 10.4%, p less than 0.05), no improvements were observed in profoundly jeopardized segments at the low left ventricular level (5.6 +/- 6.0 to 0.9 +/- 5.7%). Triphenyltetrazolium chloride delineation of infarction revealed significant myocardial salvage with prostaglandin E1 retroperfusion as compared with findings in untreated control dogs (3.7% +/- 1.3% of the left ventricle versus 9.3 +/- 1.9%, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Differences in beta-adrenoceptor binding in anterior and inferior myocardial wall microsomes of normal canine left ventricle.

Myocardial beta-adrenoceptor binding was investigated, with (-)3H dihydroalprenolol as radioligand, in microsomes derived from anterior (ALV) and inferior (ILV) myocardial wall sections of the canine left ventricle. Characterisation of specific binding sites revealed a hierarchy of myocardial beta-adrenoceptor binding with greater binding occurring to the anterior than the inferior wall of the left ventricle, under identical experimental conditions. Equilibrium analysis by Scatchard plots suggested a significant (P less than 0.01) difference in the number of receptors (Bmax ALV = 70 fmol . mg-1 protein vs Bmax ILV = 37 fmol . mg-1 protein) with no alteration in the binding affinity of the receptors (KDALV = 10.1 nmol . litre-1 vs KDILV = 6.7 nmol . litre-1). Such differences in the extent of binding of beta-adrenoreceptors in cardiac muscle may be of physiological and pathological significance and may account for the heterogeneity of regional autonomic responses in the heart.

Protein deprivation in primates. XI. Determinants of weight change during and after pregnancy.

Individual differences in weight gain of rhesus monkeys during pregnancy far exceed the variation in infant weight. Weight gain thus reflects maternal adjustment to her physiological state in relation to her environment rather than growth of the infant. Diets high in protein lead to great weight gain. Skeletal length is positively correlated and conception weight negatively correlated with weight gain. Season of pregnancy, length of gestation and sex of the infant are unrelated to weight gain. None of the monkeys exhibited edema during gestation. Postpartum changes in body weight were small during the first 6 weeks.

Early appearance of MB-creatine kinase activity in nontransmural myocardial infarction detected by a sensitive assay for the isoenzyme.

The early release patterns of MB-creatine kinase (CK-MB) in myocardial ischemia and infarction are largely unknown. We utilized a sensitive column chromatographic assay of CK-MB activity (precision = 1.1 IU/liter) and sequential CK-MB samples were obtained during the first 6 hours of illness to define the early time course of enzyme release. The average CK-MB in 39 normal subjects was 2.4 +/- 0.93 (mean +/- standard deviation (SD)). Twenty-two patients with ischemic chest pain, in whom myocardial infarction did not develop, were characterized by normal CK-MB's (2.4 +/- 1.0). Of 39 patients in whom transmural myocardial infarction developed, 28 (72 percent) were found to have abnormal CK-MB either initially or over a 20-minute sampling period. In contrast, 100 percent of the patients considered to have sustained a nontransmural myocardial infarction had abnormal initial CK-MBs and subsequently demonstrated significant increases in CK-MB from 28 +/- 19 initially to 41 +/- 30 IU/liter (P less than 0.01, N = 16) over the 20-minute sampling period. Thus, CK-MB appears earlier in plasma following nontransmural myocardial infarction than transmural myocardial infarction, probably reflecting perfusion to ischemic myocardium.

Mechanisms and therapy of silent myocardial ischemia and the effect of transdermal nitroglycerin.

Studies of the mechanisms and characteristics of ischemic heart disease have increasingly documented evidence of myocardial ischemia in the absence of symptoms. Recent work using objective criteria of ischemic events has confirmed that angina pectoris or its equivalents need not accompany true myocardial ischemia, and this appears to be quite common. The impact of these findings on prognosis awaits further study, but preliminary data suggest an improved prognosis for persons in whom coronary artery disease remains asymptomatic compared with symptomatic patients. Further, reduction of silent ischemic events with nitrate therapy may be associated with a more benign subsequent course. Preliminary trials show a reduction of the number, duration and magnitude of silent ischemic events by transdermal nitroglycerin. Ongoing technical innovations in monitoring systems should allow more complete characterization of this syndrome and lead to definition of medical therapy for it.

Dissociation of exercise tolerance and total myocardial ischemic burden in chronic stable angina pectoris.

Exercise treadmill tests and ambulatory monitoring were used in a double-blind, placebo-controlled, double-dummy crossover comparison of nifedipine (10 mg, 3 times daily) and transdermal nitroglycerin (15 mg). All patients (n = 20) had chronic stable angina with symptomatic and silent events. All patients had 3 episodes of angina/week and 3 episodes of ischemia/24 hr. The protocol was made up of 2 weeks of placebo followed by 2 weeks of active drug, then crossed over for 2 weeks of placebo followed by the other active drug. At the end of each 2-week period, patients had ambulatory monitoring and exercise treadmill testing. All ambulatory monitoring reports were read blind and entered into an independent data base. The results were the following: on transdermal nitroglycerin, the duration of ischemia decreased by 57% from 140 min/24 hr to 60 min/24 hr (p = 0.0054). The exercise time increased by 5.5% from 4.8 to 5.0 minutes (p = 0.16). With nifedipine, the duration of ischemia decreased by 22% from 175 min/24 hr to 137 min/24 hr (p = 0.16). The exercise tolerance time increased by 13% from 4.5 to 5.0 minutes (p = 0.0264). Nifedipine increased exercise time without altering total ischemic time, while transdermal nitroglycerin decreased total ischemic time without increasing exercise time. Thus, changes in exercise time do not necessarily predict changes in total ischemic time.

Sensitivity and specificity of MB creatine kinase activity determined with column chromatography.

Several analytic techniques are used to estimate the activity of creatine kinase (CK) isoenzymes. These techniques are undergoing continual refinement because the MB CK molecule appears to be nearly cardiospecific. An improved column chromatographic assay is presented with a lower limit of analytic detection of 0.8 mlU/ml. The assay was applied to normal subjects and to patients with myocardial infarction and with complex disease states. Results of the column chromatography were compared with those of agarose and cellulose acetate electrophoresis. There was MB CK activity in all normal sera (no. = 19, mean +/- standard deviation = 2.4 +/- 7). After acute myocardial infarction the MB CK activity increased by 650 percent (no. = 86, mean +/- standard error of the mean = 158.2 +/- 12.6 mlU/ml). The sensitivity for myocardial infarction was 100 percent. The specificity, evaluated in multiple clinical states, approached 100 percent. The column assay was compared with both agarose and cellulose acetate electrophoresis, and poor correlations with these semiquantitative systems were demonstrated. Finally, peak MB CK activity was closely correlated with kinetic analyses used to estimate infarct size (r = 0.94). Thus, column chromatography is a sensitive and specific method of estimating MB creatine kinase activity and is more precise than current electrophoretic methods.

Coronary vascular responses to nitroglycerin following aorta-coronary saphenous vein grafting in dogs.

Phasic coronary arterial flow responses to nitroglycerin were evaluated in 15 open-chest greyhounds before and after aorta-coronary saphenous vein grafting. Coronary flow was measured with electromagnetic probes proximal and distal to the vein graft site. Simultaneous measurements of branchiocephalic flow, aortic and left atrial pressure, left ventricular pressure, and dp/dt were made. Grafts were placed during normothermic hemodilution cardiopulmonary bypass and induced ventricular fibrillation. Before grafting, intravenous nitroglycerin (0.6 mg.) elicited an abrupt rise in diastolic coronary flow which decreased as aortic pressure fell and systolic coronary flow increased. However, after grafting, nitroglycerin elicited no change in diastolic coronary flow, whereas systolic coronary flow increased and aortic pressure fell. In 6 control dogs treated similarly except for aorta-coronary grafting, coronary flow responses to nitroglycerin were unchanged. These acute experiments suggest that, in dogs with aorta-coronary saphenous vein grafts, the coronary dilating effect of nitroglycerin is virutally abolished.

CK-MB release following coronary artery bypass grafting in the absence of myocardial infarction.

Elevation of levels of the myocardial-specific isoenzyme of creatine kinase (CK-MB) in the immediate postoperative period in patients undergoing coronary artery bypass grafting is usually associated with myocardial necrosis. However, mean isoenzyme elevations of 18 +/- 2 IU/L (standard error of the mean) were recently observed in 6 patients in the absence of electrocardiographic or scintigraphic (technetium 99m stannous pyrophosphate) evidence of perioperative myocardial infarction. To test the hypothesis that surgical trauma of the atrium and aorta during cannulation for cardiopulmonary bypass might contribute to elevated CK-MB levels, biopsy of the right atrial appendage and aorta of 7 patients was done at operation, the tissue samples were assayed for total creatine kinase (CK) activity using the Rosalki technique, and for CK-MB using column chromatography. The results indicate that the human atrium is a rich source of CK, with the proportion of CK-MB similar to that present in the ventricle (20%). In addition, technical considerations inherent in the performance of coronary bypass surgery may result in release of CK-MB, causing elevated serum enzyme levels in the post-coronary artery bypass patient in the absence of myocardial infarction.

Radioimmunoassay of inactive creatine kinase B protein in human plasma.

We describe a rapid, sensitive radioimmunoassay for enzymatically inactive creatine kinase B protein (CK-Bi) in plasma. 125I-CK-Bi of high specific activity and good stability was prepared by oxidant-based iodination. A 12 minute first antibody incubation was used. Bound and free antigen were separated by a second antibody system. Large excesses of purified CK-MM from human skeletal muscle did not react in the assay. Cross reactivity to CK-MB purified from the plasma of patients with acute myocardial infarction was negligible. The 95th percentile of plasma CK-Bi in 150 adults was 145 microgram equivalents/ml. Within-assay and between-assay precision ranged from 5% to 9% and 6% to 10%, respectively. Evidence is presented indicating that the assay measures inactive creatine kinase B protein, a protein not measured by current assay systems dependent on biological activity.

Treatment of silent myocardial ischemia with transdermal nitroglycerin added to beta-blockers and alprazolam.

Investigations into the mechanisms and characteristics of ischemic heart disease have increasingly documented evidence of myocardial ischemia in the absence of symptoms. Recent work using objective criteria of ischemic episodes confirmed that angina pectoris or its equivalents need not accompany myocardial ischemia and noted that these episodes appear to be quite common. The impact on prognosis awaits further study, but preliminary data suggest an adverse prognosis for patients with recurrent spontaneous silent vasoconstrictive ischemia. Furthermore, treatment of silent ischemic episodes with nitrates may be associated with reduced ischemia. Preliminary trials show reduction of the number, duration, and magnitude of silent ischemic episodes by transdermal nitroglycerin given to patients receiving beta-blockers. Therapy of acute ischemic syndromes should be designed to eliminate ischemia completely, not merely ameliorate pain.

Increased MB-creatine kinase isoenzymes in an alcoholic population.

Using agarose gel electrophoresis MB-creatine kinase (MB-CK) activity was examined in the serum of 120 patients with acute alcohol intoxication admitted to a detoxification ward. Total CK activity was increased in 67 percent of patients and MB-CK activity was increased in 8.3 percent. Alcoholic patients also were studied by Sephadex chromatography and, in seven cases, MB-CK was greater than three standard deviations from the normal. Thus, this study demonstrates that acute alcohol intoxication is associated with increased MB-CK activity. These findings raise the possibility that excessive alcohol ingestion may lead to acute myocardial injury.

Phylogenetic and molecular characteristics of Eurasian H9 avian influenza viruses and their detection by two different H9-specific RealTime reverse transcriptase polymerase chain reaction tests.

Avian influenza viruses (AIVs) of the H9 haemagglutinin subtype are endemic in many Asian and Middle-East countries, causing mortality and morbidity in poultry. Consequently there is a need for accurate and sensitive detection of Eurasian H9 subtype viruses. Two H9 RealTime reverse transcriptase polymerase chain reaction (RRT-PCR) tests, developed by Monne et al. (2008) and Ben Shabat et al. (2010), were originally validated with a limited number of H9 specimens. In the present study, the two tests have been assessed using 66 diverse H9 isolates and 139 clinical specimens from six H9 poultry outbreaks in four geographically disparate Eurasian countries. The Monne et al. (2008) test was modified and successfully detected all H9 viruses from all three Eurasian H9 lineages. Bayesian analysis of the clinical specimens' results revealed this test to be more sensitive (97%) than the Ben Shabat et al. (2010) test (31%). The latter test detected most H9 isolates of the G1 lineage, but no isolates from other H9 lineages. Mismatches in the primer/probe binding sequences accounted for sensitivity differences between the two H9 RRT-PCRs. Genetic analysis of 34 sequenced H9 haemagglutinin genes showed the South Asian and Middle-East H9 isolates to belong to the H9 G1 lineage, and possessed residues that appear to preferably bind alpha 2,6-linked sialic acid receptors which indicate a potential for human infection. European H9s clustered phylogenetically in a broader geographical group that includes recent North American H9 wild bird isolates and contemporary Asian viruses in the Y439 H9 lineage.