Bacteria detected in the honeybee parasitic mite Varroa destructor collected from beehive winter debris.

AIMS: The winter beehive debris containing bodies of honeybee parasitic mite Varroa destructor is used for veterinary diagnostics. The Varroa sucking honeybee haemolymph serves as a reservoir of pathogens including bacteria. Worker bees can pick up pathogens from the debris during cleaning activities and spread the infection to healthy bees within the colony. The aim of this study was to detect entomopathogenic bacteria in the Varroa collected from the winter beehive debris. METHODS AND RESULTS: Culture-independent approach was used to analyse the mite-associated bacterial community. Total DNA was extracted from the samples of 10 Varroa female individuals sampled from 27 different sites in Czechia. The 16S rRNA gene was amplified using universal bacterial primers, cloned and sequenced, resulting in a set of 596 sequences representing 29 operational taxonomic units (OTU97). To confirm the presence of bacteria in Varroa, histological sections of the mites were observed. Undetermined bacteria were observed in the mite gut and fat tissue. CONCLUSION: Morganella sp. was the most frequently detected taxon, followed by Enterococcus sp., Pseudomonas sp., Rahnella sp., Erwinia sp., and Arsenophonus sp. The honeybee putative pathogen Spiroplasma sp. was detected at one site and Bartonella-like bacteria were found at four sites. PCR-based analysis using genus-specific primers enabled detection of the following taxa: Enterococcus, Bartonella-like bacteria, Arsenophonus and Spiroplasma. SIGNIFICANCE AND IMPACT OF THE STUDY: We found potentially pathogenic (Spiroplasma) and parasitic bacteria (Arsenophonus) in mites from winter beehive debris. The mites can be reservoirs of the pathogenic bacteria in the apicultures.

[Lung transplantation]. / Lungentransplantation.

Lung transplantation is a therapeutic option for patients with end-stage lung diseases. Selection of candidates requires careful consideration of the disease-specific indications and contraindications for transplantation. Advances have been made in candidate selection via the ability to prognosticate outcomes of various lung diseases and through the implementation of the lung allocation score (LAS) with specific consideration of the degree of urgency and good postoperative survival rate, after neglecting the waiting time. This system has resulted in decreased mortality on the waiting list for lung transplantation. The availability of donor organs can possibly be increased by implementation of ex vivo lung perfusion as an alternative to conventional organ preservation. Risk factors for poor outcomes post-lung transplantation have been identified and understanding of the physiological, cellular and molecular mechanisms responsible for lung and airway damage has been extensively expanded. Primary graft dysfunction, infectious diseases, acute rejection, antibody-mediated rejection, lymphocytic bronchiolitis, obliterative bronchiolitis, restrictive allograft syndrome, and chronic lung allograft dysfunction are well defined complications and continue to be common causes of morbidity and mortality. This article provides a comprehensive update on these topics for the non-transplantation clinician.

Genetic interactions in the ß-adrenoceptor/G-protein signal transduction pathway and survival after coronary artery bypass grafting: a pilot study.

BACKGROUND: In heart failure, ß-adrenergic receptor (ßAR) stimulation desensitizes the receptor, uncouples the downstream Gαs protein, and diminishes signal transduction. We tested the hypotheses that haplotype-tagging single-nucleotide polymorphisms (htSNPs) within the Gαs gene (GNAS) (i) are functionally active and alter Gαs expression, (ii) influence survival after coronary artery bypass grafting (CABG), and (iii) interact with ßAR SNPs. METHODS: Amplification of GNAS intron 1 was followed by cloning, reporter assays, electrophoretic mobility shift assays, and western blots. In a pilot study, 185 patients on ßAR blockade undergoing CABG were studied prospectively. The primary endpoint was cardiac-related mortality at 1 yr. RESULTS: Two htSNPs defined three common haplotypes with altered reporter activity, allele-specific transcription factor binding, and Gαs protein expression (highest in *3 carriers followed by *2 and *1 haplotypes, P=0.013). After CABG, mortality was GNAS diplotype-dependent: *3/*3: 0%; *3/*2: 2.4%; *3/*1: 2.9%; *2/*2: 4.5%; *2/*1: 9.1%; and *1/*1: 20.0% (P=0.004). While ß(1)AR SNPs were not associated with mortality, ß(2)AR Arg16 allele carriers were at higher risk than Gly16 allele carriers (P=0.008). Gene-gene interaction using gene-related risk alleles demonstrated the number of risk alleles to be independently associated with death (hazard ratio 2.3; 95% confidence interval: 1.5-3.5; P=0.0003). Carriers of the no-risk allele had higher maximum isoproterenol-stimulated adenylyl cyclase activities than risk allele carriers (P=0.003). CONCLUSIONS: Interactions in the ßAR/Gαs pathway may be associated with altered mortality after CABG. This could reconcile previously inconclusive data regarding the effects of ßAR SNPs on cardiovascular prognosis.

Complex acute dissection of the aortic arch--a novel hybrid concept for one stage surgical repair.

Acute aortic dissection is a life threatening disease, which is occasionally limited to an ascending aorta only (DeBakey type II). In majority of patients it involves the aortic arch and entire rest of the aorta (DeBakey type I). The standardized cannulation and operation strategy can not be used in cases, when aortic arch branches are involved in dissection (complex aortic arch dissection) or in cases with malperfusion or severely compromised hemodynamics (tamponade or heart failure due to severe aortic valve insufficiency). The aim of this present review is to present the "Essen" treatment concept of complicated acute aortic arch dissection from diagnostics to operation strategy.

Changes in serum KL-6 levels are associated with the development of chronic lung allograft dysfunction in lung transplant recipients.

Chronic lung allograft dysfunction (CLAD) remains a leading cause of death after lung transplantation. KL-6 is a reliable biomarker for various interstitial lung diseases and levels are increased in lung transplant recipients with versus without bronchiolitis obliterans syndrome. This study investigated whether changes in serum KL-6 levels over time were associated with CLAD. Twenty-one lung transplant recipients had serum KL-6 measured (NANOPIA®) at baseline and after 7 years. Changes in serum KL-6 levels from baseline were determined. Receiver operating characteristic curves and Kaplan-Meier analysis were used to test the predictive value of changes in serum KL-6 over time. The average increase in KL-6 in patients with CLAD was 15% versus a 28% decrease in non-CLAD patients (p = .042). An 11% decrease in serum KL-6 level was determined as the best cut-off value to be associated with the development of CLAD (86% sensitivity, 78% specificity). Kaplan-Meier analysis confirmed the association between this cut-off and the development of CLAD (log rank p = .013). In this small cohort, changes in serum KL-6 over time were associated with the development of CLAD after lung transplantation.

Severe left ventricular failure after double lung transplantation: pathophysiology and management.

Patients undergoing bilateral lung transplantation for end-stage pulmonary hypertension may experience various complications. We describe a patient who underwent transplantation for chronic pigeon breeder's disease, who had secondary pulmonary hypertension and deteriorated right heart function, and who developed severe left heart failure during the weaning phase after successful double lung transplantation. The patient was stabilized with catecholamines and an intra-aortic balloon pump. Left heart function increased within 7 days and normalized at Day 18. Otherwise, the post-operative course was uneventful.

Direct visualization of leukocyte/endothelial cell interaction during extracorporeal circulation (ECC) in a new animal model.

OBJECTIVE: The clinical complications of extracorporeal circulation (ECC) have been linked to a systemic activation of cellular and humoral components and to a dysregulation of the microcirculatory compartment. Since to date only in vitro methods exist for evaluation, we developed an animal model to study the effects of ECC on the microcirculation. To establish the model, we assessed whether these effects are dependent on the duration of ECC. METHODS: Intravital fluorescence microscopy was used on the dorsal skinfold chamber preparation in chronically instrumented, awake Syrian golden hamsters. ECC was realized using a micro-rollerpump and a silicon tube shunting blood between the carotid artery and the jugular vein. ECC was performed in three groups for various times (2, 10 and 20 min) after application of heparin at 300 IU/kg body wt. In hamsters, the application of high-dose heparin releases endothelial bound superoxide dismutase (SOD), a natural scavenger of oxygen-derived free radicals. Protocol II assigned two groups receiving heparin at different doses of 50 and 2000 IU/kg body wt. RESULTS: ECC for 2 min served as control to exclude effects from hemodilution and resulted in a minimal induction of leukocyte/endothelial cell interaction. Isovolemic ECC for 20 min resulted in an increase in rolling (from 11 +/- 3 to 38 +/- 20%, mean +/- S.D., P < 0.05) and adherent leukocytes (from 19 +/- 16 to 215 +/- 145 cells/mm2, mean +/- S.D., P < 0.05) in postcapillary venules. Microhemodynamic parameters and functional capillary density were not significantly affected. Arterial blood pressure and heart rate were stable. Heparin at 2000 IU/kg inhibited post-ECC leukocyte adhesion following ECC, whereas 50 IU/kg showed no protective effects. CONCLUSIONS: Leukocyte/endothelial cell interaction, induced by blood contact with synthetic surfaces, was directly visualized in vivo. The number of adherent leukocytes was dependent on the duration of ECC. The application of high-dose heparin followed by release of SOD almost prevented leukocyte activation, suggesting a formation of oxygen free radicals during ECC. The new application of the hamster model may allow to study the underlying pathomechanisms and to develop therapeutic/prophylactic strategies to avert problems associated with ECC.

Deleterious effects of oxygen during extracorporeal circulation for the microcirculation in vivo.

OBJECTIVE: Clinical complications arising from extracorporeal circulation (ECC) have been linked to disturbances in the microcirculation. Hyperoxia, a mainstay of supportive treatment, is clinically used for a variety of pathological states. In previous in vivo animal experiments we found increased leukocyte/endothelial (L/E) cell interaction following ECC due to oxygen derived free radicals. This study was carried out to investigate the link between arterial pO2 during ECC and the potential damage to the microcirculation, supposedly caused by oxygen derived radicals. METHODS: Intravital fluorescence microscopy was used on the dorsal skinfold chamber preparation in syrian golden hamsters. ECC was introduced via a micro-rollerpump (0.7 ml/min) and a 60 cm silicon tube (1 mm inner diameter) shunted between the carotid artery and the jugular vein after application of 300 IE Heparin/kg/bw. Experiments were performed in chronically instrumented, awake animals (age: 10-14 weeks, weight: 65-75 g). Control inspired room air, experimental group 1 inspired 100% oxygen, group 2 received 100% oxygen and 2000 IE of Heparin i.v. (n=7/group), that releases endothelial bound superoxide dismutase, a natural scavenger of oxygen derived free radicals in the hamster. RESULTS: Normobaric inhalation of 100% oxygen increased arterial pO2 from 64+/-8.1 mmHg to 512+/-124 mmHg (P<0.05 vs. baseline). ECC under 100% oxygen reduced functional capillary density (FCD) to 70% of baseline values 8 h after ECC (P<0.05). Adherent leukocytes in postcapillary venules and arterioles increased significantly (P<0.05). 2000 IE Heparin prevented the reduction in FCD and decreased the number of adherent leukocytes. CONCLUSIONS: Reduction in FCD, increased leukocyte adherence to the microvascular endothelium of postcapillary venules and arterioles under hyperoxia compared to ECC under room air conditions, demonstrates harmful effects of oxygen during ECC in vivo. A high dose of Heparin enhances functional capillary density, thus attenuating the microvascular dysfunction/damage in the period after ECC.

Interruption of bronchial circulation leads to a severe decrease in peribronchial oxygen tension in standard lung transplantation technique.

OBJECTIVE: In clinical practice lung transplantation is the only procedure where the transplanted organ is left without its own arterial perfusion. With the interruption of the bronchial arteries the nutritive support is dependent on collateral flow by the pulmonary artery and the oxygen tension of desaturated central venous blood, representing an abnormal physiology. METHODS: To analyze this problem systematically, we used a standard single left lung transplantation model in the pig (n = 12). In accordance with the clinical standard, lung preservation was performed with modified Euro-Collins solution with addition of prostacycline. The duration of ischemia was set to 4 h. Before and after single left lung transplantation tissue oxygen tension in the peribronchial tissue was measured with Licox tissue pO2 microprobes. For validation, the myocardial tissue oxygen tension was recorded simultaneously. The hemodynamic assessment included continuous flow measurement of the left and right pulmonary artery using Transsonic ultrasound flow probes. After transplantation the animals were observed for 4 h. For hypothetic augmentation of collateral blood flow to the peribronchial tissue we administered Nitric oxide (10 ppm) to the ventilation in six pigs (group B). Six pigs (group A) served as a control without the addition of nitric oxide (NO). All pigs were ventilated with a FiO2 of 0.5 resulting in paO2 values between 160 and 200 mmHg. RESULTS: In both groups single lung transplantation led to a significant decrease in peribronchial tissue oxygen tension throughout the observation period. Pre-Tx values of peribronchial tissue oxygen tension (38.31 +/- 6.56 mmHg) decreased to 9.72 +/- 2.55 mmHg in group A and 10.3 +/- 3.61 mmHg in group B after 4 h, which could not be altered by a FiO2 of 1.0 (P < 0.0001). The addition of NO in group B led to a significantly augmented flow in the left pulmonary artery (0.63 +/- 0.31 l/min in group B vs. 0.46 +/- 0.26 l/min group A, P < 0.001) representing 67 vs. 49% of the pre-Tx flow in groups B and A, respectively, but the peribronchial tissue oxygen tension was not influenced (P > 0.05). In both groups A and B, the central venous pO2 did not differ in the postoperative period (41.83 +/- 3.27 mmHg group A vs. 43.26 +/- 2.98 mmHg group B) and was kept in a comparable range to the pretransplantation values (45.23 +/- 3.41 mmHg pre-Tx). CONCLUSIONS: The persistence of a very low peribronchial tissue oxygen tension in the early phase after lung transplantation cannot be influenced by improved pulmonary artery flow and solely relates to the central venous pO2, which cannot be augmented by the addition of NO. This mechanism might be a trigger for anastomotic healing problems, infectious complications and later development of obliterative bronchiolitis (OB).

Impact of ischemia on tissue oxygenation and wound healing: improvement by vasoactive medication.

The influence of vasoactive medication on tissue oxygenation and wound healing was investigated in the ear model of the hairless mouse. Ischemia was induced to the ears by ligating 2 of the 3 main vessel bundels and verified by measurements of tcpO2. Reduced tissue oxygenation was followed by a prolongation of the time required for complete healing of standardized wounds. Treatment with the vasoactive drug Buflomedil (3 mg/kg/day iv.) resulted in enhanced recovery of the tissue from reduced oxygenation and likewise reversed the adverse effects of ischemia on wound healing. These results warrant the use of the drug in patients suffering from delayed wound healing due to peripheral arterial disease.

Visualization of nutritive perfusion following tourniquet ischemia in arterial pattern skin flaps: effect of vasoactive medication.

We present an experimental model that makes it possible to investigate the effects of global ischemia and reperfusion on microvascular perfusion and viability of ill-proportioned (poorly designed) arterial pattern skin flaps in hairless mice. Skin flaps were created on the ears of hairless mice by dissecting two of three nutritional vessel bundles at the ear base. Under these nonischemic conditions, 19 percent of the total flap area went on to necrose (as a result of poor flap design). Global ischemia was induced to the flap tissue for 6 hours with a tourniquet clamp directly after flap incision. The extension of perfused tissue area and flap viability were assessed at the microcirculatory level by intravital video microscopy at 1, 3, 6, and 18 hours and 7 days after reperfusion in animals treated with either normal saline (control) or the vasoactive drug buflomedil hydrochloride (3 mg/kg of body weight per day, i.v., starting 4 hours prior to flap creation and continued at daily intervals until the end of the experiments). In untreated animals (n = 18), 1 hour after clamp release we observed reperfusion of 39.55 percent (38.5/44.9) of total flap area. Reperfusion remained unchanged within the following 5 hours. Within the next 12 hours, reperfused flap area was dramatically reduced to 21.9 percent (15.1/58.4). Seven days thereafter, only 18.8 percent (10.9/42.2) of total flap area remained viable. In contrast, we found in buflomedil-treated animals (n = 18) that 57.3 percent (53.5/62.9) of the total flap tissue was reperfused within the first hour after clamp release (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Long-term occurrence of Hemolivia cf. mauritanica (Apicomplexa: Adeleina: Haemogregarinidae) in captive Testudo marginata (Reptilia: Testudinidae): evidence for cyclic merogony?

Blood smears from wild-caught, long-term captive tortoises, Testudo marginata, revealed the presence of gametocytes of a Hemolivia mauritanica-like hemogregarine in the erythrocytes of 72% tortoises examined. Significant parasitemia was also found in animals living several years in captivity. Experimentally infected tortoises showed no evidence of a decrease in parasitemia level more than 15 mo after infection. Morphologically, stages found in tortoises' erythrocytes were indistinguishable from those referred to by previous workers as H. mauritanica from Testudo graeca. Moreover, successful experimental transmission to Hyalomma aegyptium confirms the conspecificity with H. mauritanica. The occurrence of H. mauritanica gametocytes in tortoise living up to 8 yr in captivity is suggested to result from continuous, long-lasting cyclic merogony in tortoises' parenchymatous organs, which is an unknown phenomenon in the life cycle of Hemolivia spp.

[Update in lung transplantation]. / Aktueller Stand der Lungentransplantation: Pulmonale und nichtpulmonale Komplikationen.

Lung transplantation is an established therapeutic option for selected patients with various end stage pulmonary diseases which prolongs survival and improves quality of life. A multitude of pulmonary and non-pulmonary complications can lead to significant morbidity thus impairing short and long-term survival. Early recognition and fast treatment of these complications are fundamental measures to prevent secondary destructive incidents. This article reviews the most frequent complications arising after lung transplantation.

Enterocytozoon bieneusi in Bovine Viral Diarrhea Virus (BVDV) infected and noninfected cattle herds.

Enterocytozoon bieneusi known as a causative agent of opportunistic infections instigating diarrhoea in AIDS patients was identified also in a number of immunocompetent patients and in a wide range of animals, including cattle. In the present study we tested if the Bovine Viral Diarrhea Virus (BVDV), the most common pathogen underlying immunosuppressive Bovine Viral Diarrhoea (BVD), can enhance the occurrence of opportunistic infections with E. bieneusi in cattle. Six dairy farms were investigated using ELISA to detect antibodies against or antigens arising from BVDV in collected sera. A total of 240 individual faecal samples from four age groups were examined for the presence of E. bieneusi by nested PCR. Sequence analysis of six E. bieneusi positive samples revealed the presence of the genotype I of E. bieneusi, previously described in cattle. The hypothesis expecting higher prevalence of E. bieneusi in BVDV positive cattle herds was not confirmed in this study; however this is the first description about E. bieneusi in cattle in the Czech Republic.

ICU controlled delay for acute type a aortic dissection repair after intervention for total visceral malperfusion: a way out of a dilemma?

Despite immediate surgical repair of the entry site in acute thoracic aortic dissection with visceral malperfusion, the results are poor. Primary restitution of visceral flow by intervention might be one way to cope with this problem, but probably causes ischemia/reperfusion associated problems after prolonged complete visceral ischemia. In this report, we demonstrate a successful attempt of controlled delay of thoracic aortic surgical repair after visceral flow restitution with stable hemodynamics.

Minimally invasive endoscopic port-access intracardiac surgery with one lung ventilation: impact on gas exchange and anaesthesia resources.

Minimally invasive endoscopic intracardiac surgery including one lung ventilation has been proposed to decrease surgical trauma but its impact on oxygenation and resource consumption has not been reported. We compared effects on gas exchange, induction, total anaesthesia time, staffing costs, and complications in 42 consecutive patients to a matched group undergoing similar surgery conventionally. Use of endoscopic compared to conventional surgery evoked a decrease in the P(a)o(2)/F(I)o(2) ratio (mean (SD) 24.1 (14.9) vs 48.9 (14) kPa, p < 0.05) following termination of bypass with one lung ventilation (10 patients showed a P(a)o(2)/F(i)o(2) below 13.3 kPa (100 mmHg)). There was also an increase of anaesthesia induction time (47 (13) vs 31 (9) min, p < 0.05), and an increase by 156 min of total anaesthesia time (474 (89) vs 321 (69) min, p < 0.05). Anaesthetist staffing costs increased by 300%. Thus, minimally invasive endoscopic intracardiac surgery consumes many more anaesthesia resources than conventional surgery and can result in hypoxaemia, but overall can be considered feasible provided that extensive continuous monitoring is employed.

Heart transplantation and consecutive mitral valve replacement.

A 48-year-old man was diagnosed with progressive mitral insufficiency due to fibrosis of papillary muscles and chordae tendineae, necessitating mitral valve replacement (MVR) 8 months after cardiac transplantation. Donor echocardiography and inspection of the heart at procurement were inconspicuous. The patient is alive, free from valve-related complications and functionally improved six years after MVR. The limited yet successful experiences with left-sided valve repair or replacement in the transplanted heart are reviewed.

Immunohistochemical differentiation of eosinophilic heart diseases using antibodies against eosinophil activation markers.

AIMS: Eosinophilic heart syndromes are rare in Western countries and include endocarditis parietalis fibroplastica (EPF) and hypersensitivity myocarditis (HM). There are striking differences in natural history and morphological findings. Since diagnosis can be difficult when analysing small myocardial biopsies lacking the characteristic histological features, we studied a set of immunohistochemical markers in order to characterize the activation status of the infiltrating eosinophils to distinguish between these two entities. METHODS AND RESULTS: This study is based on the investigation of seven explanted hearts and one left ventricular specimen collected during implantation of a left ventricular assist device from a total of seven patients with HM. Also investigated were three right and three left ventricular specimens from five patients with EPF. We used antibodies (Ab) against EG1, and EG2, CD44, and CD69 which have been described as markers to distinguish between resting and activated eosinophils. The EG1 to EG2 ratio of eosinophils and the immunoreactivity against CD44 showed no differences between the two entities. However, eosinophils in the EPF were completely negative for CD69, whereas eosinophils reacted positively within the HM group. CONCLUSION: The immunohistochemical investigation of eosinophilic heart diseases using antibodies against CD69 can be a useful tool to distinguish between hypersensitivity myocarditis and endocarditis parietalis fibroplastica.