Systemic coagulation parameters in mice after treatment with vascular targeting agents

BACKGROUND: Vascular targeting of malignant tumors has become a clinically validated new treatment approach with clear patient benefit. However clinical studies have also revealed that some types of vascular targeting agents (VTAs) are prone to coagulation system side effects. It is therefore essential to predetermine coagulation parameters in preclinical studies. As of to date, this has rarely been done, predominantly due to technical issues. The goal of this study was to establish and apply a standardized process, whereby systemic coagulation activation can be routinely measured in mice. RESULTS: We have evaluated a number of sampling techniques and coagulation tests regarding their suitability for this purpose. We were able to adapt two assays measuring soluble fibrin, a marker for a prethrombotic status. Thus, soluble fibrin could be measured for the first time in mice. All assays were validated in a positive control model for systemic coagulation activation, i.e. lipopolysaccharide-induced endotoxemia. Based on our results, we selected a panel of coagulation tests, which are both feasable and informative for preclinical testing of VTAs: soluble fibrin, thrombin-antithrombin complexes, free antithrombin III, white blood cell counts and platelet counts. The effect of tumor transplants on coagulation parameters was evaluated using this panel. We then applied this set of assays in treatment studies with a VTA developed in our laboratory to investigate a potential systemic coagulation activation. CONCLUSION: We have established a standardized panel of assays that can be used to test murine blood samples for coagulation activation in preclinical studies. All tests are feasible to perform in any research laboratory without specialized equipment. In addition, this is the first report to measure soluble fibrin, an early marker of systemic coagulation activation, in mice. The panel was applied on tumor bearing mice and mice treated with a VTA. We suggest its general application for coagulation activation analyses in mice

Development of Antibodies against Anthrose Tetrasaccharide for Specific Detection of Bacillus anthracis Spores

Methods for the immunological detection of Bacillus anthracis in various environmental samples and the discrimination of B. anthracis from other members of the B. cereus group are not yet well established. To generate specific discriminating antibodies, we immunized rabbits, mice, and chickens with inactivated B. anthracis spores and, additionally, immunized rabbits and mice with the tetrasaccharide beta-Ant-(1-->3)-alpha-L-Rhap-(1-->3)-alpha-L-Rhap-(1-->2)-L-Rhap. It is a constituent of the exosporium glycoprotein BclA and contains the newly discovered sugar anthrose 2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-beta-D-glucose. The BclA protein is a major component of the exosporium of B. anthracis spores and is decorated by the tetrasaccharide indicated above. The anthrose-containing tetrasaccharide chain seems to be highly specific for B. anthracis, which makes it a key biomarker for the detection of these spores. The different immunizations led to anthrose-reactive polyclonal and monoclonal antibodies which were analyzed by various methods to characterize their ability to discriminate between B. anthracis and other Bacillus spp. Multiple applications, such as enzyme-linked immunosorbent assay, indirect immunofluorescence assay, and electron microscopy, revealed the specificities of the polyclonal and monoclonal antibodies generated for B. anthracis spore detection. All polyclonal antibodies were able to correctly identify the B. anthracis strains tested and showed only minimal cross-reactivities with other Bacillus strains. Moreover, the antibodies generated proved functional in a new capture assay for B. anthracis spores and could therefore be useful for the detection of spores in complex samples

Harmonizing methods for wildlife abundance estimation and pathogen detection in Europe-a questionnaire survey on three selected host-pathogen combinations

__Background:__ The need for wildlife health surveillance as part of disease control in wildlife, domestic animals and humans on the global level is widely recognized. However, the objectives, methods and intensity of existing wildlife health surveillance programs vary greatly among European countries, resulting in a patchwork of data that are difficult to merge and compare. This survey aimed at evaluating the need and potential for data harmonization in wildlife health in Europe. The specific objective was to collect information on methods currently used to estimate host abundance and pathogen prevalence. Questionnaires were designed t

Early mobilisation in critically ill COVID-19 patients: a subanalysis of the ESICM-initiated UNITE-COVID observational study

Background Early mobilisation (EM) is an intervention that may improve the outcome of critically ill patients. There is limited data on EM in COVID-19 patients and its use during the first pandemic wave. Methods This is a pre-planned subanalysis of the ESICM UNITE-COVID, an international multicenter observational study involving critically ill COVID-19 patients in the ICU between February 15th and May 15th, 2020. We analysed variables associated with the initiation of EM (within 72 h of ICU admission) and explored the impact of EM on mortality, ICU and hospital length of stay, as well as discharge location. Statistical analyses were done using (generalised) linear mixed-effect models and ANOVAs. Results Mobilisation data from 4190 patients from 280 ICUs in 45 countries were analysed. 1114 (26.6%) of these patients received mobilisation within 72 h after ICU admission; 3076 (73.4%) did not. In our analysis of factors associated with EM, mechanical ventilation at admission (OR 0.29; 95% CI 0.25, 0.35; p = 0.001), higher age (OR 0.99; 95% CI 0.98, 1.00; p ≤ 0.001), pre-existing asthma (OR 0.84; 95% CI 0.73, 0.98; p = 0.028), and pre-existing kidney disease (OR 0.84; 95% CI 0.71, 0.99; p = 0.036) were negatively associated with the initiation of EM. EM was associated with a higher chance of being discharged home (OR 1.31; 95% CI 1.08, 1.58; p = 0.007) but was not associated with length of stay in ICU (adj. difference 0.91 days; 95% CI − 0.47, 1.37, p = 0.34) and hospital (adj. difference 1.4 days; 95% CI − 0.62, 2.35, p = 0.24) or mortality (OR 0.88; 95% CI 0.7, 1.09, p = 0.24) when adjusted for covariates. Conclusions Our findings demonstrate that a quarter of COVID-19 patients received EM. There was no association found between EM in COVID-19 patients' ICU and hospital length of stay or mortality. However, EM in COVID-19 patients was associated with increased odds of being discharged home rather than to a care facility. Trial registration ClinicalTrials.gov: NCT04836065 (retrospectively registered April 8th 2021)

Surveillance of tularaemia in Kosovo*, 2001 to 2010

Tularaemia, caused by Francisella tularensis, had not been registered in Kosovo* before an outbreak in 1999 and 2000. A national surveillance system has been implemented in Kosovo* since 2000 to monitor a number of diseases, including tularaemia. Antibody detection in human sera was used for laboratory diagnosis of tularaemia and F. tularensis lipopolysaccharide antigen was used as a marker of infection. The purpose of this study is to describe the incidence of tularaemia in Kosovo* after the 1999–00 outbreak. In 2001 and 2002, a second outbreak occurred, with 327 serologically confirmed cases. From 2001 to 2010, 25–327 cases were registered per year, giving a mean annual incidence of 5.2 per 100,000 population. The most likely sources of infection were contaminated drinking water and food. The dominant clinical manifestations were the glandular (79%) and ulcero-glandular (21%) forms. By 2010, the disease had spread throughout Kosovo*. Presumably as a result of war and subsequent environmental disruption, mass population displacement and breakdown of sanitation and hygiene, the two major outbreaks of tularaemia resulted in the establishment of an active endemic area of tularaemia in Kosovo*

Tularaemia seroprevalence of captured and wild animals in Germany: the fox (Vulpes vulpes) as a biological indicator

A total of 2475 animals from Germany, both captive and wild, were tested for antibodies against Francisella tularensis to obtain more knowledge about the presence of this pathogen in Germany. An indirect and a competitive ELISA served as screening methods, positive and inconclusive samples were confirmed by Western blot. Of the zoo animals sampled between 1992 and 2007 (n = 1122), three (0·3%) were seropositive. The seroconversion of a hippopotamus in Berlin Zoo was documented. From 1353 serum samples of wild foxes (Vulpes vulpes), raccoon dogs (Nyctereutes procyonoides) and wild boars (Sus scrofa), collected between 2005 and 2009 in the federal state of Brandenburg (surrounding Berlin), a total of 101 (7·5%) tested positive for antibodies to F. tularensis lipopolysaccharide. Our results indicate a higher seroprevalence of F. tularensis in wildlife in eastern Germany than commonly assumed. Furthermore, we found foxes and raccoon dogs to be biological indicators for tularaemia

Academic investigator-initiated trials and the challenge of sponsor responsibility: the Cologne Sponsor Model

Background With the amendment to the German Drug Law in 2004, the conduct of clinical trials changed by at least two main aspects: (1) The principles of Good Clinical Practice (GCP) were implemented in the national legislation, and (2) for the first time, the function of the sponsor of a clinical trial and the clinical trial itself have become legally binding definitions. By that, legal differences between industrial and academic clinical trials no longer exist. Clinical trials initiated by investigators have to fulfil the same requirements while the entire sponsor responsibility has to be carried out by the Coordinating Investigator or his institution including implementation of a quality management system according to the GCP. Purpose The Cologne Sponsor Model is an effective approach with settings, structures, basic features, action, and reporting lines, as well as funding for clinical trials initiated in an academic environment. Results The University of Cologne assumes the sponsor responsibility for clinical trials organised by the university researchers according to law. Sponsor's duties are delegated to a central operational unit of the sponsor - the Clinical Trials Center Cologne - which further delegates duties to the Coordinating Investigator. Clinical Trials Center Cologne was established in 2002 to support the performance of clinical trials at the university by offering comprehensive advisory and practical services covering all aspects of study planning and conduct. Furthermore, a specialised division of its quality management department acts as an independent sponsor's Quality Assurance Unit. The Clinical Trials Center Cologne has established a quality management system consisting of different components (1) to enable a reasoned decision to subsequent delegation, (2) for risk-based surveillance of trial conduct (audits, monitoring-checks, and reports), and (3) support and training of the Coordinating Investigator. Limitations Double functions of persons and departments in the university environment sometimes make it difficult to define roles in such a model. Therefore, it is necessary to establish clear reporting lines and moreover to monitor regularly and carefully the roles and responsibilities. Conclusions With the combination of central management and support, control and independence of the researchers, our model represents a 'risk-based' system that offers a sensible option that fulfils the requirements of legal regulations and GCP for trials organised within the university environment. Clinical Trials 2012; 9: 781-787. http://ctj.sagepub.co

Surveillance of tularaemia in Kosovo*, 2001 to 2010

Tularaemia, caused by Francisella tularensis, had not been registered in Kosovo* before an outbreak in 1999 and 2000. A national surveillance system has been implemented in Kosovo* since 2000 to monitor a number of diseases, including tularaemia. Antibody detection in human sera was used for laboratory diagnosis of tularaemia and F. tularensis lipopolysaccharide antigen was used as a marker of infection. The purpose of this study is to describe the incidence of tularaemia in Kosovo* after the 1999–00 outbreak. In 2001 and 2002, a second outbreak occurred, with 327 serologically confirmed cases. From 2001 to 2010, 25–327 cases were registered per year, giving a mean annual incidence of 5.2 per 100,000 population. The most likely sources of infection were contaminated drinking water and food. The dominant clinical manifestations were the glandular (79%) and ulcero-glandular (21%) forms. By 2010, the disease had spread throughout Kosovo*. Presumably as a result of war and subsequent environmental disruption, mass population displacement and breakdown of sanitation and hygiene, the two major outbreaks of tularaemia resulted in the establishment of an active endemic area of tularaemia in Kosovo*