A multi-centre retrospective study of Nocardia speciation and antimicrobial susceptibility in Queensland, Australia.

The study aims to characterise the species identification and antimicrobial susceptibility testing (AST) results of Nocardial isolates from adult patients across major public hospitals in Queensland, Australia, over a 15-year period. A multi-centre retrospective observational study of Nocardia sp. isolates was conducted from 7 major public hospitals in Queensland, Australia, over a 15-year period. Clinical samples from patients aged ≥ 18 years that isolated Nocardia sp. were included. Demographic and clinical data were collected, along with species identification and AST results. Overall, 484 Nocardia sp. were isolated. Most patients were male (297, 61%) with a mean (IQR) age of 60 (51-75) and a median (IQR) Charlson Comorbidity Index of 4 (2-6). Of these, 239 (49%) patients were immunosuppressed. Organisms were most frequently isolated from sputum (174, 36%), and superficial swabs (102, 21%). Patients presented with pulmonary infections (165, 35%) and superficial skin and soft tissue infections (87, 18%) most commonly. One hundred (21%) isolates were deemed pulmonary colonisation and were not treated. Of the speciated organisms, N. nova complex was the most common (93, 19%), followed by N. farcinica complex (79, 16%). Organisms were reliably susceptible to linezolid (240/245, 98%), amikacin (455/470, 97%), and trimethoprim/sulfamethoxazole (459/476, 96%), but less so to imipenem (243/472, 51%) and ceftriaxone (261/448, 58%). This is the largest Australian description of Nocardia sp. to date. Given antimicrobials are often commenced prior to AST results and sometimes even speciation, characterisation of local species and antibiogram data is important to guide empiric choices and local guidelines.

Reporting guidelines and journal quality in otolaryngology.

OBJECTIVES: Journals increasingly use reporting guidelines to standardise research papers, partly to improve quality. Although defining journal quality is difficult, various calculated metrics are used. This study investigates guideline adoption by otolaryngology journals and whether a relationship exists between this and journal quality. DESIGN, SETTING, PARTICIPANTS: Retrospective MEDLINE database review for English language, Index Medicus, journals of interest to otolaryngologists (October 2013). MAIN OUTCOME MEASURES: The resulting journals were examined for the number of guidelines endorsed and then tabulated against surrogate measures of journal quality (Impact factor, Eigenfactor, SCImago, Source-Normalised rank). The primary outcome measure was the number of recognised reporting guidelines endorsed per journal. This was then correlated against journal quality scores. For comparison, a further small sample correlation was performed with 6 randomly selected and 6 high-profile clinical non-otolaryngology journals. RESULTS: 37 otolaryngology journals were identified. Number of guidelines used and quality scores were not normally distributed. Mean guideline usage was 1.0 for otolaryngology journals, 1.5 for randomly selected, and 5.5 for the high-profile journals. Only 18/37 (49%) otolaryngology journals endorsed any guidelines, compared with 11/12 non-otolaryngology journals. Within otolaryngology, Eigenfactor positively correlated with guideline use (r = 0.4, n = 44, p < 0.01) otherwise no correlation was found between guideline endorsement and journal quality. CONCLUSIONS: Reporting guideline endorsement within otolaryngology journals is low. Although it might be expected that use of reporting guidelines improved quality, this is not reflected in the derived quality scores in otolaryngology. This may reflect low levels of use/enforcement, that quality indicators are inherently flawed, or that generalised guidelines are not always appropriate or valued by editors.

Endothelium-derived relaxing factor.

This article reviews what is known of endothelium-derived relaxing factor and its possible physiologic and pathophysiologic roles. This relaxing factor is now thought to be nitric oxide or a ready source of it. It acts as an endogenous nitrovasodilator, stimulating soluble guanylate cyclase to increase cyclic guanosine monophosphate (GMP) levels in vascular smooth muscle and platelets, with consequent relaxant and anti-aggregatory effects (predominantly when stimulated through receptor-operated channels). Its actions are thus synergistic with those of cyclic adenosine monophosphate (AMP)-mediated stimulation (for example, adenosine, prostacyclin). Endothelium-derived relaxing factor is unstable and is thought to act only very locally in vivo. Its release is continuous in the basal state and is stimulated by a number of neuropeptides and by agents released during platelet activation and thrombosis--with large differences in activity among different vessels. Endothelium-derived relaxing factor activity is also flow related, thereby coordinating vasomotor behavior in an intact vascular tree in response to changes in flow. Endothelium-derived relaxing factor activity is reduced in several pathologic states, including atherosclerosis.

Unstimulated release of endothelium derived relaxing factor is independent of mitochondrial ATP generation.

Rabbit aortic strips were used to investigate the effect of mitochondrial inhibitors on basal (unstimulated) endothelium dependent relaxation. Since haemoglobin inhibits and the cyclic GMP phosphodiesterase inhibitor, MB22948, amplifies endothelium dependent relaxation they were used to provide evidence of basal activity of endothelium derived relaxing factor (EDRF). Basal activity was not inhibited by incubation with any of three differently acting inhibitors of mitochondrial ATP generation. The mechanism underlying basal EDRF production may thus differ from that of stimulated EDRF production, which is abolished by these mitochondrial inhibitors.

Effect of lipid feeding on endothelium dependent relaxation in rabbit aortic preparations.

The effect of a cholesterol enriched diet on concentration responses to 5-hydroxytryptamine, ergometrine, and acetylcholine and on endothelium dependent relaxation was investigated in rabbit aortic strip preparations. Sensitivity to ergometrine was slightly reduced after both two and 10 weeks of cholesterol feeding. Sensitivity to 5-hydroxytryptamine was slightly reduced after two but not after 10 weeks of cholesterol feeding. Sensitivity to acetylcholine was not altered at two or 10 weeks in the absence of endothelium, but the endothelium dependent lowering of the constrictor concentration-response was almost abolished after 10 weeks of cholesterol feeding. Likewise the endothelium dependent relaxant response of preconstricted preparations to acetylcholine was almost abolished after 10 weeks of cholesterol feeding. Thus this model of hyperlipidaemia leads after 10 weeks to loss of endothelium dependent relaxation.

Determinants of hypoxic contracture in isolated heart muscle preparations.

Further studies on hypoxic contracture are described, using resting and contracting isolated muscle preparations from several species. The effect of temperature, pH, substrate, calcium concentration, osmolality and inotropic interventions was explored. The 'protective' effect of acidosis was not contingent on its negative inotropic influence. No evidence was adduced that hypoxic contracture can be modulated other than by altering energy supply or demand. The study does not discriminate between rigor and rised cytosolic calcium as mechanisms causing hypoxic contracture, but demonstrates that hypoxic contracture is not directly dependent on the availability of intracellular calcium.

Myocardial reoxygenation damage: can it be circumvented?

We investigated whether reoxygenation damage could be prevented by interventions directed towards reducing calcium influx only during the reoxygenation period. We measured reoxygenation contracture and recovery of contractile performance, using isolated papillary muscle preparations from cat and rabbit, pretreated with ouabain so as to exaggerate the phenomenon of reoxygenation contracture. Reoxygenation contracture was abolished and contractile recovery achieved by lowering extracellular calcium during early reoxygenation and then gradually replacing it. Gradual reoxygenation only postponed contracture and contractile failure. The slow channel blocker, diltiazem, but not verapamil or lidoflazine--in similarly negative inotropic concentrations of 10(-4) mol X litre-1, 10(-4) mol X litre-1 and 2 X 10(-5) mol X litre-1 respectively--reduced early reoxygenation contracture, as did Mg2+ (30 mmol X litre-1), Mn2+ (8 mmol X litre-1), or metabolic acidosis (pH 6.5), without in any case allowing contractile recovery. These observations indicate that reoxygenation damage is not an irrevocable consequence of the preceding hypoxic insult. They imply that calcium entry during early reoxygenation contributes both to contracture and contractile failure, that this occurs through paths other than the slow calcium channel, and that diltiazem may have properties additional to those of blocking the slow calcium channel.

Haptoglobin-haemoglobin complex in human plasma inhibits endothelium dependent relaxation: evidence that endothelium derived relaxing factor acts as a local autocoid.

Endothelium dependent relaxation of isometrically mounted rabbit aortic strip preparations was rapidly inhibited by human plasma at dilutions down to 1:1000. Gel filtration and ion exchange chromatography were used to demonstrate that this inhibitory activity was present in fractions containing haptoglobin. Purified haptoglobin itself possessed no inhibitory action against endothelium dependent relaxation, but the haptoglobin-haemoglobin complex did, consistent with the documented ability of haemoglobin to inhibit this phenomenon. The concentration of haemoglobin normally bound to haptoglobin is sufficient to account for the inhibitory properties of human plasma. This suggests that endothelium derived relaxing factor exerts no downstream intravascular effect in vivo and thus that its physiological dilator role is that of a local autocoid acting on subjacent smooth muscle.

Production of endothelium derived relaxant factor is dependent on oxidative phosphorylation and extracellular calcium.

The production (synthesis or release or both) of endothelium derived relaxant factor was studied in rabbit aortic strip preparations and an aortic-coronary artery bioassay system. Production of endothelium derived relaxant factor was rapidly inhibited by agents that inhibit mitochondrial electron transport or F1-ATPase, or which uncouple oxidative phosphorylation, but was only slowly impaired by inhibition of glycolysis. It was dependent also on the presence of extracellular calcium with a rapid on-off response time. This study shows that production of endothelium derived relaxant factor appears to be dependent on both oxidative phosphorylation and extracellular calcium.

Myocardial stiffness during hypoxic and reoxygenation contracture.

Isolated rat papillary muscle preparations were used to study hypoxic contracture, and cat papillary muscle preparations with ouabain to study reoxygenation contracture. Electronic analysis of the response to rapid small sinusoidal perturbations gave a continuous measurement of the elastic and viscous components of total stiffness. Increased resting force during hypoxic contracture was characterised by an increase in resting elastic and viscous stiffness relative to the control stiffness-active force relationships. During reoxygenation contracture the stiffness-force relationships followed those of active force development. The linear active force-elastic stiffness relationship (dt/dl=kT+c) was also reversibly altered during hypoxic contracture, predominantly by an increase in intercept c. These data imply that hypoxic contracture unlike reoxygenation contracture is not due solely to a rise in intracellular calcium, but is associated with a component of stiffness not participating an active force development, for example rigor.

Preparation of human saphenous vein for coronary artery bypass grafting impairs its capacity to produce prostacyclin.

Prostacyclin production was measured from freshly isolated human saphenous vein and from vein subjected to routine surgical preparation for coronary bypass grafting. Surgical preparation had no effect on spontaneous prostacyclin production but significantly reduced stimulated rates from 16.9(1.1) to 7.1(0.5) pg.min-1 per mg wet weight (n = 27). Stimulated prostacyclin production was not reduced by storage of vein for 2 h at 23 degrees C in blood or saline nor by distension, but it was reduced to 5.0(0.6) pg.min-1 per mg (n = 10) by de-endothelialisation. Reduced prostacyclin production, which might in itself contribute to vein graft occlusion, provides a quantitative biochemical estimate of endothelial integrity.

Endothelial function is impaired in fit young adults of low birth weight.

OBJECTIVE: Non-insulin-dependent diabetes, hypertension and ischaemic heart disease, with insulin resistance, are associated with low birth weight (the 'Small Baby Syndrome'). Common to these adult clinical conditions is endothelial dysfunction. We tested the hypothesis that endothelial dysfunction could precede their development in those of low birth weight. METHODS: Endothelial function was measured by ultrasonic 'wall-tracking' of flow-related brachial artery dilatation in fit 19-20 year old subjects randomly selected (blind to the investigators throughout the study) from low (< 2.5 kg) and normal (3.0-3.8 kg) birth weight subjects in the 1975-7 cohort of the Cardiff Births Survey and with no known cause for endothelial dysfunction. RESULTS: Flow-related dilatation was impaired in low birth weight relative to normal birth weight subjects (median 0.04 mm [1.5%] [n = 22] cf. 0.11 mm [4.1%] [n = 17], p < 0.05; 0.04 mm [1.5%] [n = 15] cf. 0.12 mm [4.4%] [n = 12], p < 0.05 after exclusion of inadvertently included ever-smokers). CONCLUSION: The findings suggest that endothelial dysfunction is a consequence of foetal malnutrition, consistent with contributing to the clinical features of the 'Small Baby Syndrome' in later adult life.

Syndrome X and endothelial dysfunction.

OBJECTIVE: Syndrome X (angina, normal coronary arteriogram and positive exercise test) remains an enigma with unexplained features and apparent conflicts of evidence. The present study addressed whether (i) the Syndrome is characterised by generalised flow-related endothelial dysfunction, (ii) myocardial thallium201 defects reflect myocardial or microvascular dysfunction, (iii) endothelial dysfunction and its consequences can be improved by oral L-arginine. METHODS: Flow-mediated brachial artery dilatation was measured by ultrasonic 'wall-tracking' in 7 Syndrome X patients, further characterised as having thallium201 defects and no known cause of endothelial dysfunction, and a normal control group. Syndrome X patients entered a 4-week randomised double-blind placebo-controlled cross-over trial of oral L-arginine (7 g twice daily), with brachial artery studies, exercise tests and technetium99 tetrafosmin scans. RESULTS: Flow-mediated dilatation was absent in Syndrome X vs. normal. Stress technetium99 tetrafosmin and thallium201 scans showed similar defects. Flow-mediated dilatation, symptom-limited exercise duration and peak oxygen consumption (VO2max) were increased but rate-pressure-product (RPP) and radionuclide defects were unchanged after L-arginine vs. placebo. CONCLUSIONS: The study supports coronary microvascular rather than myocardial dysfunction and shows loss of flow-mediated dilatation in systemic arteries. Oral L-arginine improved flow-mediated dilatation, exercise capacity and VO2max (by ca. 17%) despite unchanged RPP. The findings support generalised endothelial dysfunction. The arginine effects imply NO-mediated improvement of skeletal muscle perfusion suggesting improved homogeneity of microvascular distribution.

Metabolic damage to human saphenous vein during preparation for coronary artery bypass grafting.

Segments of saphenous vein from patients undergoing coronary artery by-pass graft surgery were frozen in liquid nitrogen immediately on dissection (control), after stripping of the adventitia and side branch ligation (manipulation), after distention with blood (distention), or at completion of the last proximal anastomosis (prepared vein). Vein was stored during the operation in patient's heparinised arterial blood at room temperature. Frozen vein was extracted with perchloric acid. ATP, ADP, and AMP, adenosine, inosine and hypoxanthine concentrations were measured by high pressure liquid chromatography. Prepared vein had ca 50% lower ATP concentrations and ATP/ADP ratio than control vein, higher concentrations of inosine and hypoxanthine and lower concentrations of AMP and adenosine. ATP concentration and ATP/ADP ratio did not correlate with the time elapsed between dissection and freezing of the prepared vein. The characteristic changes seen in prepared vein were not seen when control vein was simply stored in arterial blood at 23 degrees C, in normal saline at 23 degrees C or 4 degrees C, in Krebs-Ringer bicarbonate buffer at 37 degrees C or at St Thomas's Hospital cardioplegic solution at 4 degrees C. Distention with unlimited pressure did not distension at less than 300 mmHg gave rise to the same changes in ATP concentration and ATP/ADP ratio as in the prepared vein. These results show that vein suffered metabolic changes during preparation for bypass grafting and suggest that uncontrolled distention may contribute to these changes. Such biochemical measurements provide a quantitative estimate of tissue damage and allow objective comparison of different preparative techniques.

Endothelium-derived relaxing factor and atriopeptin II elevate cyclic GMP levels in pig aortic endothelial cells.

1. Two directly-acting stimulants of soluble guanylate cyclase, glyceryl trinitrate (0.1 microM) and sodium azide (10 microM), and a receptor-mediated stimulant of particulate guanylate cyclase, atriopeptin II (10 nM), each elevated the cyclic GMP content of primary cultures of pig aortic endothelial cells without affecting the cyclic AMP content. 2. Two receptor-mediated stimulants of adenylate cyclase, glucagon (1 microM) and isoprenaline (10 microM), had no effect on the cyclic AMP or cyclic GMP content of these cells, but the directly acting stimulant, forskolin (30 microM), induced a small increase in cyclic AMP content. 3. Three agents that release endothelium-derived relaxing factor (EDRF); bradykinin (0.1 microM), ATP (10 microM) and ionophore A23187 (0.1 microM), each markedly elevated the cyclic GMP content of pig aortic endothelial cells, but acetylcholine (1 microM) had no effect. None of these agents had any effect on cyclic AMP content. 4. Two agents that potentiate the actions of EDRF; M & B 22948 (100 microM) and superoxide dismutase (30 units ml-1), each elevated the cyclic GMP content of pig aortic endothelial cells without affecting the cyclic AMP content. Pretreating cells with catalase (100 units ml-1) did not affect the rise in cyclic GMP content induced by superoxide dismutase (30 units ml-1). 5. Pretreatment of pig aortic endothelial cells with haemoglobin (10 microM) reduced the resting content of cyclic GMP and blocked the increase in cyclic GMP content induced by glyceryl trinitrate (0.1 microM), sodium azide (10 microM), bradykinin (0.1 microM), ATP (10 microM), ionophore A23187 (0.1 microM), M & B 22948 (100 microM) and superoxide dismutase (30 units ml-1), but not that induced by atriopeptin II (10 nM). 6. Pretreatment of pig aortic endothelial cells with an inhibitor of soluble guanylate cyclase, methylene blue (20 microM), had no effect on the resting content of cyclic GMP. Methylene blue (20 microM) blocked the increase in cyclic GMP content induced by glyceryl trinitrate (0.1 microM), M & B22948 (100 microM) and bradykinin (0.1 microM), but not that induced by atriopeptin II (10 nM). 7. The data show that soluble guanylate cyclase, particulate guanylate cyclase and adenylate cyclase are present in pig aortic endothelial cells. They further suggest that EDRF, produced spontaneously or in response to vasoactive agents, elevates endothelial cyclic GMP content by stimulating soluble guanylate cyclase. It is possible that this may serve as a feedback loop by which the endothelial cell modulates EDRF production.

Evidence that inhibitory factor extracted from bovine retractor penis is nitrite, whose acid-activated derivative is stabilized nitric oxide.

1. Unactivated extracts of bovine retractor penis (BRP) contains 3-7 microM nitrite. Acid-activation of these extracts at pH 2 for 10 min followed by neutralization generates the active form of inhibitory factor (IF; assayed by its vasodilator action on rabbit aorta), and is associated with partial loss of nitrite. 2. Increasing the time of acid-activation at pH 2 from 10 to 60 min with intermittent vortex mixing generates greater vasodilator activity and increases nitrite loss. 3. When acid-activated and neutralized extracts are incubated at 37 degrees C or 30 min or boiled for 5 min, vasodilator activity is lost and nitrite content increased. Reactivation of these samples at pH 2 for 10 min followed by neutralization leads to partial recoveries of vasodilator activity with loss in nitrite content. 4. Addition of sodium nitrite to BRP extracts increases acid-activatable vasodilator activity pro rata. 5. Acid-activation of aqueous sodium nitrite solutions results in less loss of nitrite and generation of less vasodilator activity than BRP extracts. Vasodilatation is only transient and is rapidly abolished on neutralization, whereas responses to acid-activated BRP extracts are more prolonged and activity is stable on ice. 6. Bovine aortic endothelial cells yield vasodilator activity that is indistinguishable from that isolated from BRP. It is activated by acid, stable on ice, abolished by boiling or by haemoglobin, and appears to be due to the generation of nitric oxide (NO) from nitrite. 7. The data provide confirmatory evidence that nitrite in BRP extracts is IF, that acid-activation of BRP extracts yields NO which is responsible for its vasodilator action, and that inactivation occurs by decay of NO to nitrite and nitrate. They further suggest that BRP extracts contain a NO-stabilizing agent which favours conversion of nitrite to NO. 8. The finding that bovine aortic endothelial cells yield an agent indistinguishable from IF suggests that nitrite in endothelial cells may likewise be the precursor of endothelium-derived relaxing factor (EDRF), itself identified as NO.

In vivo EDRF activity influences platelet function.

The administration of carbachol to rabbits to stimulate the release of endothelium derived relaxing factor (EDRF) results in inhibition of platelet aggregation and elevation of platelet cyclic GMP content. These effects are reversed by simultaneous administration of the EDRF inhibitors methylene blue or haemoglobin. The data provide the first direct biochemical evidence of in vivo EDRF activity.

Endothelium-derived relaxing factor inhibits in vitro platelet aggregation.

We studied the effects of endothelium-derived relaxing factor (EDRF), bovine retractor penis muscle inhibitory factor and sodium nitroprusside, three stimulants of guanylate cyclase, on the in vitro aggregation of washed human platelets. Platelet aggregation induced either by collagen or by the thromboxane A2 analogue U46619 was inhibited by all three agents. The anti-aggregatory effect of each agent was inhibited by haemoglobin. The anti-aggregatory effect of EDRF was potentiated by superoxide dismutase. These findings are discussed in relation to a potential role for EDRF in haemostasis.

Effects of delta1-trans-tetrahydrocannabinol on mechanical performance of isolated heart muscle preparations.

1 The effects of four concentrations of delta1-trans-tetrahydrocannabinol (delta1-THC) (0.1, 1, 5 and 20 microgram/ml) were compared to that of the vehicle (ethanol, 0.5 microliter/ml) on the mechanical performance of isolated cardiac muscle of the cat and rat. 2 In rat isolated papillary muscles, delta1-THC (20 microgram/ml) caused a decline in mechanical performance (-8% in developed tension and -11% in Vmax) in contrast to the apparent lack of effect of ethanol. 3 All other parameters of mechanical performance studied in both rat and cat papillary muscles were unaffected by delta1-THC when compared with ethanol. 4 It was concluded that delta1-THC in concentrations up to 20 microgram/ml had negligible effect on contractile performance, the time course of contraction or muscle elasticity in rat and cat isolated papillary muscle preparations under the conditions studied.