Galleria mellonella as an alternative model of Coxiella burnetii infection.

Coxiella burnetii is a Gram-negative intracellular bacterium and is the causative agent of the zoonotic disease Q fever. Several rodent and non-human primate models of virulent phase I C. burnetii [Nine Mile (NM)I] have been developed, and have been used to determine the efficacy of antibiotics and vaccine candidates. However, there are several advantages to using insect models to study host-microbe interactions, such as reduced animal use, lowered cost and ease of manipulation in high containment. In addition, many laboratories use the avirulent phase II C. burnetii clone (NMII) to study cellular interactions and identify novel virulence determinants using genetic manipulation. We report that larvae of the greater wax moth, Galleria mellonella, were susceptible to infection with both C. burnetii NMI and NMII. Following subcutaneous infection, we report that intracellular bacteria were present within haemocytes and that larval death occurred in a dose-dependent manner. Additionally, we have used the model to characterize the role of the type 4 secretion system in C. burnetii NMII and to determine antibiotic efficacy in a non-mammalian model of disease.

Radiographic features of pelvis and hip joint development of English Bulldogs.

OBJECTIVES: To evaluate distinctive features of pelvis and hip joint development of English Bulldogs throughout the first year of life. METHODS: The pelves of 20 English Bulldogs were radiographed at three different ages (<4, 6-8, and 12-14 months). At each time point, the dogs were clinically evaluated and the abnormal hip joints were classified as mild, moderate, or severely deformed. The pelves were compared to a phantom study in which external rotation of a normal hemipelvis around its long axis was artificially created at different degrees, with different pelvic inclinations, and classified as either normal and without deformity, or as mild, moderate, or severely deformed. Hip joints and pelvic scores were statistically compared. RESULTS: Although none of the dogs were considered lame at the end of the study, none of the hips showed normal development; 77.5% were moderately to severely deformed at 12-14 months of age. At this age, 75% of the hemipelves had moderate to severe torsional deformity (>5.2 degrees of external rotation), with retroversion of the acetabulum confirmed by the presence of the crossover sign. An external rotation of the hemipelvis on its long axis >5 degrees was likely associated with a moderate to severely altered hip joint conformation. CLINICAL SIGNIFICANCE: Abnormal hip conformation was common in this series of English Bulldogs. Torsional deformity of the pelves with acetabular retroversion was a common and distinctive feature, which has not yet been thoroughly studied in dogs. These findings need further evaluation in English Bulldogs as well as in other breeds.

T-cell-mediated epineurial vasculitis and humoral-mediated microangiopathy in cryoglobulinemic neuropathy.

We used immunohistochemistry to assess the role of humoral and cellular factors in endoneurial microangiopathy and epineurial vasculitis in 15 nerve biopsies of patients with axonal neuropathy and monoclonal or mixed cryoglobulinemia (CG). Deposition of immunoglobulins and cytolytic complement was detected in endoneurial capillaries of patients with mixed CG. Epineurial inflammatory infiltrates containing beta2-integrin-positive lymphocytes and monocytes surrounded arterioles expressing cell adhesion molecules, thus suggesting a cell-mediated pathogenesis of the epineurial vasculitis. On the other hand, the absence of immune complex deposition and polymorphonuclear elements suggests a minor role for the humoral mechanisms in the formation of the vasculitic lesions. This study indicates that both cell-mediated mechanisms and immune complexes/cryoglobulins are involved, although at different levels, in the pathogenesis of CG neuropathy.

Studies of skin-window exudate human neutrophils: increased resistance to pentoxifylline of the respiratory burst in primed cells.

Human neutrophils were isolated both from peripheral blood (PB) and from aseptic inflammatory exudates obtained by the Senn's skin window technique (SW). The respiratory burst (O2- production) induced by in response to n-formyl-methionyl-lencyl-phenylalanine (fMLP) and by serum-treated zymosan (STZ) was investigated using a microplate assay. SW neutrophils were primed to enhanced fMLP-dependent O2- production in response to fMLP but not to STZ. Pentoxifylline, a cAMP-elevating drug, dose-dependently inhibited the respiratory burst in any experimental condition, but the dose-effect curves were markedly different according the stimulant used and the source of the cells. With fMLP as stimulant, a significant inhibition of the O2- production by PB neutrophils was obtained using doses of 10 micrograms/ml, while SW neutrophils were inhibited only by doses equal or higher than 100 micrograms/ml. With STZ as stimulant, the inhibition of the respiratory burst of PB neutrophils and of SW neutrophils was obtained only with doses higher than 400 micrograms/ml and 1 mg/ml respectively. Pentoxifylline dose-dependently (10 micrograms/ml to 1 mg/ml) increased the intracellular adenosine 3'-5'-cyclic monophosphate (cAMP) to the same extent in SW and in PB neutrophils. These data indicate that the priming of neutrophil oxidative metabolism by in vivo inflammation is associated with an increase in the resistance to the regulating effect of cAMP on the fMLP-dependent activation pathway of NADPH oxidase. The fact that therapeutic doses of pentoxifylline do not inhibit the respiratory burst of primed neutrophils may have relevance in the interpretation of the clinical effects of this drug.

Monitoring of elastase in plasma of burned patients in relation to other inflammation parameters.

Twenty burned patients divided in three groups according to the severity of the lesions were investigated at 1- or 2-day intervals for up to 5 weeks after injury. Plasma elastase levels were elevated during the first day after injury and were correlated with the area of the burns. However, plasma elastase was rapidly bound and inactivated by protease inhibitors. Leucocyte counts, fever and the concentration of alpha-1-proteinase inhibitor were not correlated with the extent of the burn. The rise of plasma elastase was not accompanied by consumption of the elastase inhibitory capacity (EIC) of plasma, which increased to a plateau around day 5. The EIC values were in accord with the rise of alpha-1-proteinase inhibitor, the major anti-elastase agent in plasma. Studies of blister fluid in eight patients showed that the elastase content was higher than that of corresponding plasma, while the concentration of alpha 1-proteinase inhibitor and the EIC were comparable with those of plasma. Measurements of the levels of tumour necrosis factor released by stimulated macrophages in five patients with major burns showed no significant increase compared with controls.

An uncommon cause of pericardial actinomycosis.

Cardiac actinomycosis is rare; the pericardium is the most frequently involved site, but myocardial, endocardial and valvular involvement have all been documented. Most cases originate from a thoracopulmonary site of actinomycosis and spread directly to the pericardium. Widespread dissemination from extrathoracic organs is uncommon; in fact actinomycosis is prevented by anatomical barriers and hematogenous diffusion is rare. We describe an uncommon case of pericardial actinomycosis due to a draining fistula from the liver to the pericardial space across the diaphragm. The massive dissemination through the fistula could explain the peculiar echocardiographic images of macroscopic, echo-reflective, irregular masses, floating in the pericardial space, probably consistent with aggregates of sulfur granules.

Analytical performance of an heterogeneous enzyme immunoassay using the same separation system.

A single "one step" solid phase sandwich enzyme immunoassay was developed. The assay employs two monoclonal antibodies with affinity and specificity for different epitopes of the analyte: one is covalently coupled to peroxidase, while the other one is coupled to biotin. The addition of the reagent to an avidin-coated strip gave a specific and efficient mean for binding the sandwich complex to the solid phase, via the high affinity interaction between biotin and avidin. The methodology was applied to the determination of hypophysis hormones (LH, FSH, PRL), tumor markers (CEA, AFP, FRT, hCG beta free subunit, alpha subunit and others alpha-P Amylase). Analytical performances concerning the developed kits are described, demonstrating the good reproducibility, sensitivity, accuracy and the wide application field for this new technology.

Relation between demographic and epidemiological characteristics and permanency under a dental health care program for HIV infected patients.

The association between factors involved in health care and the health status of the people has been proven. The use of health care services, particularly in the case of patients who suffer from chronic pathologies, has been the object of many studies aimed at establishing factors which contribute to guarantee permanence in treatment and implementation of health care controls. The purpose of the present study was to identify the response of HIV infected or AIDS patients to the oral health care program and establish the association between permanence in treatment and the presence of risk factors, epidemiological or demographic conditions of the users. Ninety patients selected at random from the 300 who attended the Clinic for High Risk Patient Care, School of Dentistry, University of Buenos Aires (CLAPAR, Spanish acronym) during 1994-1995 were included in this study. The patients were assigned to one of seven groups, according to their permanence in treatment and commitment to the program during the phase of maintenance in health. Each of these categories was characterized in terms of age, sex, educational level, place of residence (CIRFS, 1990), type of job, type of health coverage and place where healthcare was received, risk behavior and date of positive serological diagnosis. The frequency of each variable was established. Contingency tables were employed to establish the statistical significance of the association between the different variables and the patient categories. The data revealed that 24.2% of the patients performed occasional or emergency consultation, 57.1% achieved discharge with or without the assistance of the social worker or are still in treatment and 18.7% abandoned the program. Significant association were found between the response to odontological treatment and the following variables: place of residence, date of positive serological diagnosis, and risk behavior. We may conclude that certain demographic, epidemiological or life-related factors would be linked to the response to odontological treatment.

Effect of pelvic inclination and torsional deformity on canine acetabular morphology with computed tomography. A phantom study.

OBJECTIVES: To evaluate how the inclination and torsional deformity of the hemipelvis using extra-rotation as a model affect acetabular angle (AA) and dorsal acetabular rim angle (DARA) assessment with computed tomography (CT). METHODS: A normal canine hemipelvis positioned in dorsal recumbency was scanned with a 16-slice multidetector CT scanner, with different degrees of torsional deformity, using extra-rotation around the long axis (0-5-10-15°) of the hemipelvis. Each degree of extra-rotation was acquired at 0°, +20° and -20° of gantry tilt, to mimic different pelvic inclinations on its transverse axis. Cranial and central individual acetabular angles (IAA) and central DARA were calculated and correlated with inclination and torsional deformity. RESULTS: A very strong negative correlation was found between cranial and central IAA, pelvic inclination, and torsional deformity. A very strong positive correlation was found between DARA, pelvic inclination, and torsional deformity. CLINICAL SIGNIFICANCE: Pelvic inclination and torsional deformity affect acetabular angles assessment with CT. The greater the inclination (ilia far from the tabletop in dorsal recumbency) and torsional deformity of the pelvis, the worse the acetabulum appeared. A standardized scanning protocol for acetabular morphology assessment is needed and it should consider pelvic inclination. The actual relevance of pelvic torsional deformity is not well known and it should be investigated more thoroughly.

[Clotiazepam vs flunitrazepam in premedication in a day hospital regimen]. / Clotiazepam vs flunitrazepam nella premedicazione in regime di day-hospital.

PIP: The authors evaluated preoperative sedation, intraoperative analgesia, and psychometric integrity resiliency of 98 female patients who underwent voluntary interruption of pregnancy. These women, with a mean age of 27.5 years (range of 19-35) received Clotiazepam or Flunitrazepam during the preanesthesia. The patients were administered the Zazzo "deux barrages" test. The 1st 2 parameters did not show any difference between the groups, whereas the patients treated with Clotiazepam recovered their pschometric abilities significantly more quickly than the patients treated with Flunitrazepam. Side effects were mild, equally present in both groups, and likely attributable to the intraoperative medication. The results suggest that short 1/2-life benzodiazepines like Clotiazepam are 1st choice drugs for the preanesthesia of short surgical operations.^ieng

[Trazodone versus flunitrazepam in premedication in day-care surgery]. / Trazodone versus flunitrazepam nella premedicazione in regime di ricovero giornaliero.

A prospective single-blind study was conducted to compare flunitrazepam vs trazodone in the premedication of patients undergoing day-case surgery for termination of pregnancy, with particular regard to the degree of preoperative sedation, intraoperative analgesia and postoperative recovery. 86 patients were randomly allocated to receive orally 45 minutes before the surgical procedure either flunitrazepam 2 mg (group F) or trazodone 50 mg (group T). In both groups anaesthesia was achieved by i.v. fentanyl 2.5 micrograms/kg and ketamina 250 micrograms/kg. Patients in group F showed a deeper degree of preoperative sedation. There were no significant differences in intraoperative analgesia and in the immediate arousal time. In the postoperative period, the incidences of emetic symptoms and dizziness were similar in both groups; the incidence of drowsiness was significantly higher in group F at 120 minutes but not at 180 minutes of observation. Psychomotor performance was assessed preoperatively two days before the surgical procedure and 60, 120 and 180 minutes after surgery, using the Toulouse-Pieron test and the reaction time to a luminous stimulus with the aid of a computerized analogic tachystoscope (Neurometer). Trazodone allowed a more rapid recovery of psychomotor performance and it can represent a valid alternative to the use of benzodiazepines in the premedication of day-case surgical patients.

Alcohol and pregnancy: an international comparison.

A study was set up to compare the drinking habits of women of childbearing age in two cities (Milan and Southampton) in two countries, Italy and the United Kingdom, and to look at the effects of alcohol consumption on the weight of the babies born to these women. The sample consisted of 1516 women in Milan and 996 in Southampton, who were interviewed after delivery; data on their drinking history, before and during pregnancy, were gathered, together with social, demographic and health data. The clinical records of the women and their babies were examined. On average, both groups drank less than one unit/day before pregnancy, but there were differences in drinking habits between the two countries, as well as in the modifications induced by pregnancy. The possible effects of alcohol on birthweight were then considered. After controlling for confounding factors, there was no general association between birthweight and alcohol consumption. The impact on health education policy is discussed.

Molecular aspects of membrane fission in the secretory pathway.

Membrane fission is essential in various intracellular dissociative transport steps. The molecular mechanisms by which endocytic vesicles detach from the plasma membrane are being rapidly elucidated. Much less is known about the fission mechanisms operating at Golgi tubular networks; these include the Golgi transport and sorting stations, the trans-Golgi and cis-Golgi networks, where the geometry and physical properties of the membranes differ from those at the cell surface. Here we discuss the lipid and protein machineries that have so far been related to the fission process, with emphasis on those acting in the Golgi complex.

Column affinity chromatography for bound/free separation in ligand assays. I. Radioimmunoassay of choriomammotropin (human placental lactogen).

A method is described for separating antibody-bound from free fractions in ligand assays by column affinity chromatography, and its application to radioimmunoassay of choriomammotropin. In the method, 70 x 10 mm (i.d.) polypropylene columns containing about 150 mg of immunosorbent (goat anti-rabbit gamma-globulins covalently linked to Sepharose CL-4B) are used. Standards or unknowns, tracer and antiserum, pipetted into bottom-capped columns, are kept separated from the immunosorbent bed by a porous polyethylene disc and allowed to react for 15 min at room temperature. The reaction mixture is then allowed to pass through the columns by removing the bottom caps. Free antigen is eluted by washing the column, and discarded; antibody-bound fractions remain bound to the immunosorbent. The radioactivity in the columns is counted. The major advantages of the present technique, arising from the liquid-phase reaction combined with the solid-phase separation by column affinity chromatography, are the very low nonspecific binding (less than 1%), good sensitivity (0.02 mg/L), good precision (CV 3.4%), and simple and fast (30-min) assay. For 50 clinical samples so assayed (gamma) and compared with a polyethylene glycol precipitation technique (x), the regression equation was: y - 0.14 + 0.98x (r = 0.994). The assay method was clinical validated by 3493 determinations.

[The incidence of stress ulcers and the effectiveness of preventive alkalization treatment in patients scheduled for major vascular surgery. Prospective study]. / Incidenza dell'ulcera da stress ed efficacia del trattamento profilattico alcalinizzante in pazienti sottoposti ad interventi di chirurgia aortica maggiore. Studio prospettico.

BACKGROUND: A prospective double blind study has been carried out in order to evaluate the current incidence of stress ulceration and the efficacy of acid-reducing prophylaxis, in patients scheduled for major vascular surgery, without a documented history of ulcer disease or previous upper GI-tract surgery. METHODS: Seventy patients have been randomly assigned to treatment group with ranitidine 50 mg e.v. every six hours or to control group with placebo at the same intervals. Gastric juice has been evaluated for pH and for occult blood by a slide test four times a day and cultured once a day on a suitable ground in the postoperative period. Data have been analyzed on a "treatment efficacy" and not "intention to treat" basis: patients have been considered alkalinized if more than 80% of pH measurement were > 4.5. Patients have been therefore divided into two groups, the "alkaline" one (43 patients) and the "acid" one (27 patients). RESULTS: Twenty-five patients (58.1%) in the alkaline group vs four patients (14.8%) in the acid group, showed positive gastric-juice cultures (p > 0.01). Twenty-three patients in the alkaline group (53.4%) and nine patients in the acid group (33.3%) showed occult gastric bleeding (p = NS). Overall, nine patients (12.8%) had overt gastrointestinal bleeding, four in the alkaline group (9.3%) and five in the acid group (18.5%) (p = NS). Endoscopic evaluation revealed erosive gastropathy in six patients, gastric ulcer in two patients and duodenal ulcer in one. Neither occult nor overt gastrointestinal bleeding did correlate with gastric pH values. On the contrary, the development of overt gastrointestinal bleeding has been positive associated with length of aortic cross-clamping and with postoperative score index. CONCLUSIONS: This observation suggests that factors other than gastric acidity contribute to mucosal damage and bleeding, as intraoperative mesenteric ischemia/hypoperfusion, or oxygen free radicals damage produced during reperfusion of districts below the level of aortic cross-clamping. The results obtained suggest that routinary alkalisation of gastric juice is not warranted in the prevention of gastrointestinal bleeding in major vascular surgical patients.

Impact of perioperative haemodynamic monitoring on cardiac morbidity after major vascular surgery in low risk patients. A randomised pilot trial.

OBJECTIVE: to evaluate whether perioperative haemodynamic optimisation influences outcome from infrarenal abdominal aortic aneurysm repair. METHODS: a consecutive series of 100 eligible patients were randomised to either haemodynamic optimisation through the use of a pulmonary artery catheter (CI > 3.0 l/min/sqm, PWP > 10 and <18 mmHg, SVR <1450 dyne/sec/cm(-5), DO(2)> 600 ml/min/sqm) or conventional treatment. RESULTS: there were no differences in terms of in-hospital mortality, cardiovascular morbidity, postoperative renal failure or duration of hospital stay between the groups. CONCLUSIONS: in this study perioperative haemodynamic optimisation was not beneficial.

[Ultrasonography-guided identification of the lumbar epidural space]. / Individuazione ecoguidata dello spazio epidurale lombare.

The study was designed to assess the reliability of sonographic evaluation in the prediction of the depth of the lumbar epidural space. Forty males, scheduled for epidural anesthesia for surgical repair of inguinal hernia, were prospectively studied. Patients were placed in a sitting position and sagittal scanning of the lumbar spine was performed with a 5-MHz transducer over the fourth or fifth interspace in order to identify the deeper hyperechogen interface, which represents the landmark between the ligamentum flavum and the epidural space. Ultrasound depth was measured and transducer removed. A Tuohy needle 18 G was then introduced percutaneously according to the standard technique and a rubber slide placed over it, so that it depth of insertion could be accurately measured. The potential for using ultrasounds for prediction of the distance from skin to epidural space was analyzed using a simple linear regression analysis; p values < 0.05 were considered significant. Mean values of ultrasound depth and needle depth were respectively 51 mm (SD 6.3) and 50.9 mm (SD 6.2); the correlation coefficient was 0.99. Ultrasound scanning of the lumbar spine provides an accurate measurement of the depth of the epidural space, which can facilitate the performance of the epidural anaesthesia and may decrease the complication rate, particularly in those patients in which anatomic landmarks are obscured.

Increased serum levels of ICAM-1, ELAM-1 and TNF-alpha in inflammatory disorders of the peripheral nervous system.

Endothelial intercellular adhesion molecule-1 (ICAM-1) and glycoprotein E-selectin (ELAM-1) allow the homing of leukocytes to inflammation sites. A circulating form of ICAM-1 markedly increases in inflammatory CNS disorders. In the present study, the serum levels of ICAM-1, ELAM-1 and tumor necrosis factor-alpha (TNF-alpha) were measured in patients with acute (AIDP) and chronic (CIDP) inflammatory demyelinating polyneuropathies and cryoglobulinemic neuropathy (CGN). Immunoenzymometric assays revealed increased sICAM-1 levels in some of these patients; furthermore, high titres of ELAM-1 and TNF-alpha were detected in two patients with AIDP and one patient with CGN. Our data extend previous observations on inflammatory PNS disorders by showing that, in addition to ICAM-1, ELAM-1 also represents a useful marker of endothelial activation and that, taken together, the two molecules may serve as an indicator of specific pathogenetic mechanisms.