Study protocol: a pragmatic, cluster-randomized controlled trial to evaluate the effect of implementation of the Truenat platform/MTB assays at primary health care clinics in Mozambique and Tanzania (TB-CAPT CORE).

BACKGROUND: In 2020, the WHO-approved Molbio Truenat platform and MTB assays to detect Mycobacterium tuberculosis complex (MTB) and resistance to rifampicin directly on sputum specimens. This primary health care center-based trial in Mozambique and Tanzania investigates the effect of Truenat platform/MTB assays (intervention arm) combined with rapid communication of results compared to standard of care on TB diagnosis and treatment initiation for microbiologically confirmed TB at 7 days from enrolment. METHODS: The Tuberculosis Close the Gap, Increase Access, and Provide Adequate Therapy (TB-CAPT) CORE trial employs a pragmatic cluster randomized controlled design to evaluate the impact of a streamlined strategy for delivery of Truenat platform/MTB assays testing at primary health centers. Twenty-nine centers equipped with TB microscopy units were selected to participate in the trial. Among them, fifteen health centers were randomized to the intervention arm (which involves onsite molecular testing using Truenat platform/MTB assays, process process optimization to enable same-day TB diagnosis and treatment initiation, and feedback on Molbio platform performance) or the control arm (which follows routine care, including on-site sputum smear microscopy and the referral of sputum samples to off-site Xpert testing sites). The primary outcome of the study is the absolute number and proportion of participants with TB microbiological confirmation starting TB treatment within 7 days of their first visit. Secondary outcomes include time to bacteriological confirmation, health outcomes up to 60 days from first visit, as well as user preferences, direct cost, and productivity analyses. ETHICS AND DISSEMINATION: TB-CAPT CORE trial has been approved by regulatory and ethical committees in Mozambique and Tanzania, as well as by each partner organization. Consent is informed and voluntary, and confidentiality of participants is maintained throughout. Study findings will be presented at scientific conferences and published in peer-reviewed international journals. TRIAL REGISTRATION: US National Institutes of Health's ClinicalTrials.gov, NCT04568954. Registered 23 September 2020.

[Operative risk of geriatric patients in cardiac surgery]. / Operatives Risiko geriatrischer Patienten in der Herzchirurgie.

BACKGROUND: Despite substantial progress in interventional cardiology, there are still many geriatric patients who require cardiac surgery. Estimation of the operative risk is therefore of great importance. OBJECTIVE: The prognostic value of the geriatric assessment for estimation of the operative risk was evaluated. MATERIAL AND METHODS: Between 2008 and 2009 a geriatric assessment was carried out on 500 patients before an urgent or elective cardiac surgery intervention. The primary endpoints were in-hospital death, death within 30 days after the intervention and stroke. A secondary endpoint was the combination of death, stroke and in-hospital complications. RESULTS: The average age of the patients was 77.1 ± 4.6 years and 44.3% of the particpants were women. Aortic stenosis was the primary reason for surgery in 49.2% of patients and coronary artery disease in 38.8% of patients. Half of the patients (56.5%) showed functional impairments in one or more evaluated domains. Significant limitations in cognitive function were present in 11.8% and in mobility in 2.4% of the patients. The 30-day mortality was 2.9% and stroke occurred in 1.4% of the patients. After multivariate analysis cognitive impairment remained independently associated with the operative mortality (odds ratio OR 3.8, 95% confidence interval CI 1.2-12.7). CONCLUSION: The perioperative mortality of older patients in cardiac surgery is low. A limited functional status detected in the geriatric assessment is associated with an increased mortality. Impaired cognitive function is an independent predictor of postoperative mortality.

Discovery of a novel channel-forming protein in the cell wall of the non-pathogenic Nocardia corynebacteroides.

Detergent extracts of whole cells of the Gram-positive, non-pathogenic, strictly aerobic bacterium Nocardia corynebacteroides contain channel-forming activity. The protein responsible for channel formation was identified using lipid bilayer experiments. It was purified to homogeneity and had an apparent molecular mass of about 134 kDa on SDS-PAGE when it was solubilized at 40 degrees C. When the 134 kDa protein was heated to 100 degrees C for 10 min in sample buffer, it dissociated into subunits with a molecular mass of about 23 kDa and focused at pI of 4.5 during isoelectric focusing. The pure 134 kDa protein was able to increase the specific conductance of artificial lipid bilayer membranes from phosphatidylcholine-phosphatidylserine mixtures by the formation of ion-permeable channels. The channels had an average single-channel conductance of 5.5 nS in 1 M KCl and were found to be cation-selective. Asymmetric addition of the 134 kDa protein to lipid bilayer membranes resulted in an asymmetric voltage-dependence. The analysis of the single-channel conductance as a function of cation radii using the Renkin correction factor and the effect of negative charges on channel conductance suggested that the diameter of the cell wall porin is about 1.0 nm. The channel characteristics of the cell wall channel of N. corynebacteroides were compared with those of other members of the mycolata. They share common features because they are composed of small molecular mass subunits and form large and water-filled channels.

Beating heart operations including hybrid revascularization: initial experiences.

BACKGROUND: The outcome of patients (n = 45) with coronary one- to three-vessel disease undergoing beating heart operations using a recently developed stabilizing device was investigated. METHODS: Left internal mammary artery-to-left anterior descending coronary artery (LIMA-to-LAD) revascularization was carried out alone (n = 31) or as hybrid procedure in combination with a balloon angioplasty (n = 14). RESULTS: All 45 patients underwent a successful LIMA-to-LAD procedure without intraoperative complication during a 21 +/- 8-minute (range, 10 to 53 minutes) LAD occlusion time. In 14 hybrid procedures a total of 19 stenoses including 3 left main stenoses were treated successfully by percutaneous transluminal coronary angioplasty and stenting. The postoperative courses were uneventful with the exception of two surgical reexplorations necessitated by bleeding. No worsening of renal, neurologic, or respiratory functions occurred in any patient. In the group having a single LIMA-to-LAD procedure, early postoperative coronary angiograms (22 of 31) showed a patent LIMA graft and excellent anastomosis; this was also true in 4 patients 12 months after operation as shown in angiograms. All patients undergoing hybrid revascularization demonstrated a patent LIMA-to-LAD anastomosis; in 1 patient there was a dissection in the midlevel of the LIMA, which was stented successfully. The 6-month follow-up angiograms in 7 of 14 patients revealed open LIMA bypass grafts in all patients except 1, who was stented because of dissection. CONCLUSIONS: These data indicate that a beating heart operation including hybrid revascularization is safe and effective in selected patients with coronary one- to three-vessel disease including left main stenosis. This approach may be especially advantageous in comparison with conventional coronary artery bypass grafting in patients with severe concomitant disease.

A new method for coronary occlusion and local stabilization during minimally invasive LIMA-to-LAD-bypass.

The minimally invasive direct coronary occlusion and stabilizing technique (midCOAST)-system, a new device for coronary occlusion and local stabilization during minimally invasive LIMA-to-LAD-bypass is presented. A closed platform with an oval opening in its center provides optimal immobilization together with platform-fixed vessel-loops, which are used for LAD-occlusion. Clinical results in 72 consecutive patients indicate that the midCOAST-device can be safely and effectively used for minimally invasive LIMA-to-LAD-procedure, even in patients with impaired left ventricular function. Due to the optimal immobilization of the target area the quality of the LIMA-to-LAD-anastomosis, documented by post-operative angiography (62/72), was excellent in all cases.

Recombinant hirudin as an anticoagulant during cardiac operations: experiments in a pig model.

OBJECTIVE: The efficacy and safety of recombinant hirudin (r-hirudin) compared with heparin as an anticoagulant during open-heart surgery has been studied in a pig model. METHODS: A total of 18 Göttingen minipigs were randomly divided into three treatment groups and subjected to cardiopulmonary bypass for 1 h. Heparin-treated animals received a bolus of unfractionated heparin of 400 IU/kg body weight. Recombinant hirudin was given by a bolus injection of 1 mg/kg body weight, followed by a 1 h lasting infusion of 1 mg/kg body weight per h. The heparin-anticoagulated animals and one group of the hirudin-treated animals additionally received aprotinin at a dosage of 17500 KIU/kg body weight (KIU, kallikrein inhibitory units). In the second group of r-hirudin-treated animals, the aprotinin was replaced by saline. RESULTS: The extracorporeal circuit remained patent for a 1 h pump period in all of the animals studied. There was no evidence of vascular occlusion or clot formation in the r-hirudin-treated animals. The anticoagulant efficacy of the hirudin protocol used is further demonstrated by the results of electron-microscopical scans of the pump-line filters. Fibrin deposits were visible only in the heparin-treated animals and not in r-hirudin-treated pigs. Despite this strong anticoagulant effect, there was no evidence of an increased bleeding tendency in r-hirudin-treated pigs. Moreover, histological studies showed a statistically significant (P < 0.05) higher incidence of tissue bleeding in the heparin/aprotinin-treated animals compared with the r-hirudin/aprotinin-treated pigs. Studying the platelet function, a statistically significant (P < 0.01) better preserved ADP- and collagen-induced platelet aggregation was seen in the r-hirudin/aprotinin-treated animals when compared with heparin/aprotinin-treated animals. CONCLUSIONS: These data demonstrate that r-hirudin can be used successfully as an alternative anticoagulant to heparin during cardiac operations including cardiopulmonary bypass. The better preservation of platelet function suggests that r-hirudin may reduce the postoperative risk of bleeding.

The unfolded protein response controls induction and activation of ADAM17/TACE by severe hypoxia and ER stress.

The family of ADAM (a disintegrin and metalloproteinase) proteins has been implicated in tumor initiation and progression. ADAM17/tumor necrosis factor-α (TNFα)-converting enzyme (TACE) has been initially recognized to release TNFα as well as its receptors (TNFRs) from the membrane. ADAM17, TNFα and TNFR have been found upregulated in cancer patients, although the underlying mechanisms remain largely unknown. As hypoxia is a hallmark of cancer that can lead to severe stress conditions accumulating in endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), we investigated the role of these stress conditions in the regulation of ADAM17 and release of TNFR1.We found that severe hypoxia induced ADAM17 expression and activity. Although hypoxia-inducible factor 1α (HIF1α) was important to maintain basal ADAM17 mRNA levels during moderate hypoxia, it was not sufficient to induce ADAM17 levels under severe hypoxia. Instead, we found that ADAM17 induction by severe hypoxia can be mimicked by ER stressors such as Thapsigargin and occurs as a consequence of the activation of the PERK/eIF2α/ATF4 and activating transcription factor 6 (ATF6) arms of UPR in several tumor cell lines. ADAM17 expression was also increased in xenografts displaying ER stress because of treatment with the vascular endothelial growth factor (VEGF) inhibitory antibody Bevacizumab. Additionally, severe hypoxia and ER stress activated ADAM17 and ectodomain shedding of TNFR1 involving mitogen-activated protein (MAP) kinases and reactive oxygen species (ROS). Collectively, these results show that ADAM17 is a novel UPR-regulated gene in response to severe hypoxia and ER stress, which is actively involved in the release of TNFR1 under these conditions. These data provide a novel link between severe hypoxic stress conditions and inflammation in the tumor environment.

Preexisting atrial fibrillation as predictor for late-time mortality in patients with end-stage renal disease undergoing cardiac surgery--a multicenter study.

BACKGROUND: Although patients with end-stage renal disease (ESRD) are considered to be high-risk patients in cardiac surgery, the reported studies are rather small, resulting in unsatisfactory analyses of outcome determinants. Therefore, we aimed to identify possible risk factors, with a particular focus on the impact of pre-existing atrial fibrillation (AF) on the postoperative short-term and long-term mortality of ESRD patients undergoing cardiac surgery. METHODS: In a multicenter study 522 patients with ESRD undergoing CABG only (62.9 %), valve surgery only (17.2 %), or both (19.9 %) with comparable demographic and other cardiac risk factor characteristics were investigated retrospectively over a period of 10 years. The outcome was divided into perioperative (within 30 days) and late morbidity and mortality, and multivariate analysis was performed for both. RESULTS: The mean perioperative mortality was 11.5 % and the 5-year survival rate was 42 %. Emergency surgery, insulin-dependent diabetes mellitus, the number of vein grafts and age were identified as risk factors whereas complete revascularization, the use of an internal thoracic artery and the presence of sinus rhythm were identified as beneficial factors for long-term survival. 14.1 % of all patients had pre-existing AF. Although AF was not identified as an independent risk factor for perioperative mortality ( P = 0.59), it was identified as an independent predictor for late mortality ( P < 0.001). Median survival of patients without AF was 1816 days, while for patients with AF it was only 715 days. CONCLUSIONS: AF does represent an independent predictor for long-term but not perioperative mortality in patients with ESRD. However, effective treatment of AF is controversially discussed. Anticoagulation therapy or perioperative ablation of the arrhythmia should be considered in order to improve the survival of these patients.

Recombinant hirudin for cardiopulmonary bypass anticoagulation: a randomized, prospective, and heparin-controlled pilot study.

BACKGROUND: Lepirudin, a recombinant hirudin, is a direct acting thrombin inhibitor that has been used as a heparin alternative in patients with heparin-induced thrombocytopenia requiring on-pump cardiac surgery. To evaluate the efficacy, safety, and clinical utility of lepirudin as a cardiopulmonary bypass (CPB) anticoagulant, we compared lepirudin with heparin in a routine CPB setting. METHODS: Twenty patients were randomly assigned to receive lepirudin (0.25 mg/kg b. w. bolus and 0.2 mg/kg b. w. added to the CPB priming) or heparin (400 U/kg b. w. bolus) with protamine reversal. Lepirudin and heparin anticoagulation during CPB was monitored using the ecarin clotting time or ACT, respectively and additional lepirudin (5 mg) or heparin (5000 U) boluses were administered. RESULTS: The CPB circuit was performed in both groups without thromboembolic complications. Median blood loss during the first 36 hours was statistically higher ( P = 0.007) in the lepirudin group (1.226 +/- 316 ml) compared to the heparin group (869 +/- 189 ml). One patient of the lepirudin group developed pulmonary embolism 24 hours after surgery. This patient was tested homozygous for the FV-Leiden mutation. CONCLUSION: Lepirudin provides effective CPB anticoagulation but induces a higher postoperative blood loss than heparin. Lepirudin should be restricted to patients undergoing CPB who cannot be exposed to heparin.

Lepirudin for cardiopulmonary bypass surgery in a patient with terminal renal insufficiency and acute heparin-induced thrombocytopenia.

A patient with triple heart valve disease, heparin-induced thrombocytopenia, and terminal renal insufficiency was treated successfully using lepirudin for anticoagulation of cardiopulmonary bypass (CPB) and during the postoperative course. Anticoagulatory monitoring was performed with ecarin clotting time during CPB and aPTT postoperatively.

Pulmonary embolism after cesarian section due to heparin-induced thrombocytopenia despite normal platelet count.

Heparin-induced thrombocytopenia (HIT) type II is typically characterized by a decrease in platelet count to values between 20 and 120 x 10 (9)/L or a platelet count fall of greater than 50%. We report on a patient who developed a HIT syndrome, thrombosis of the vena cava, and fulminant pulmonary embolism during heparin treatment after cesarean section, without a significant decrease in platelet count. Lepirudin anticoagulation and ecarin clotting time (ECT) monitoring were used successfully during cardiopulmonary bypass.

Amplification of resistance genes in Haemophilus influenzae plasmids.

Intramolecular amplification produces tandem repeats of tetracycline and combined tetracycline-chloramphenicol resistance determinants in conjugative plasmids of Haemophilus influenzae. This process depends on host recombination pathways. Physical mapping revealed the tetracycline transposon involved in amplification to be almost identical with Tn10, including two IS10 insertion elements. The chloramphenicol resistance determinant of the combined transposon is 1.9 kilobases (kb) in size and is bound by two 1.3-kb inverted repeats. Insertion in the close vicinity of the inside end of the left-hand IS10 generates a deletion of a 1.6-kb Tn10 region. The amplifiable units were resolved to comprise not only the respective resistance transposons, but also an additional 1.6-kb sequence (designated AS) which was demonstrated to be identically present in the different amplification systems studied. AS separates amplified transposons from each other, thereby maintaining the same orientation. Moreover, AS is present at the left flank of the transposons, but is missing at the right one. It was shown that AS represents a general constituent of the H. influenzae plasmids of the 45-kb class. Evaluation of the results suggests that AS is responsible for the recombinational events involved in the gene amplification process.

Study of the properties of a channel-forming protein of the cell wall of the gram-positive bacterium Mycobacterium phlei.

The gram-positive bacterium Mycobacterium phlei was treated with detergents. Reconstitution experiments using lipid bilayers suggested that the detergent extracts contain a channel forming protein. The protein was purified to homogeneity by preparative SDS-PAGE and identified as a protein with an apparent molecular mass of about 135 kDa. The channel-forming unit dissociated into subunits with a molecular mass of about 22 kDa when it was boiled in 80% dimethylsulfoxid (DMSO). The channel has on average a single channel conductance of 4.5 nS in 1 m KCl and is highly voltage-dependent in an asymmetric fashion when the protein is added to only one side of the membrane. Zero-current membrane potential measurements with different salts implied that the channel is highly cation-selective because of negative point charges in or near the channel mouth. Analysis of the single-channel conductance as a function of the hydrated cation radii using the Renkin correction factor and the effect of the negative point charges on the single-channel conductance suggest that the diameter of the cell wall channel is about 1.8 to 2.0 nm. The channel properties were compared with those of other members of the mycolata and suggest that these channels share common features. Southern blots demonstrated that the chromosome of M. phlei and other mycolata tested contain homologous sequences to mspA (gene of the cell wall porin of Mycobacterium smegmatis).

The cell wall porin of the gram-positive bacterium Nocardia asteroides forms cation-selective channels that exhibit asymmetric voltage dependence.

Detergent-solubilized cell wall extracts of the gram-positive, strictly aerobic bacterium Nocardia asteroides contain channel-forming activity as judged from reconstitution experiments using lipid bilayer membranes. The cell wall porin was identified as a protein with an apparent molecular mass of about 84 kDa based on SDS-PAGE. The porin was purified to homogeneity using preparative SDS-PAGE. The 84-kDa protein was no longer observed after heating in SDS buffer. The presumed dissociation products were not observed on SDS-polyacrylamide gels. The cell wall porin increased the specific conductance of artificial lipid bilayer membranes from phosphatidylcholine/phosphatidylserine mixtures by the formation of cation-selective channels, which had an average single-channel conductance of 3.0 nS in 1 M KCl. The single-channel conductance was only moderately dependent on the bulk aqueous KCl concentration, which indicated negative point charge effects on the channel properties. The analysis of the concentration dependence of the single-channel conductance using the effect of negative charges on channel conductance suggested that the diameter of the cell wall channel is about 1.4 nm. Asymmetric addition of the cell wall porin to lipid bilayer membranes resulted in an asymmetric voltage dependence. The cell wall channel switched into substates, when the cis side of the membrane, the side of the addition of the protein, had negative polarity. Positive potentials at the cis side had no influence on the conductance of the cell wall channel.

Origin of Haemophilus influenzae R factors.

The Haemophilus influenzae R plasmids specifying resistance against one, two, or three antibiotics which have emerged in different parts of the world were shown to have closely related but not identical plasmid cores. The gene for ampicillin resistance in the H. influenzae plasmid pKRE5367 is part of a transposon similar to Tn3, which was transposed from pKRE5367 onto RSF1010 in Escherichia coli. An indigenous H. influenzae plasmid (pW266) was isolated. Its properties correspond to those of the H. influenzae R plasmids, except for the presence of a drug resistance transposon. The in vitro-generated H. influenzae R plasmids carrying an ampicillin resistance transposon, a tetracycline resistance transposon, and a transposon for combined tetracycline-chloramphenicol resistance resembled the natural isolates. The findings support the hypothesis that the R plasmids of H. influenzae are of multiclonal evolutionary origin.