Low-Molecular-Weight Heparin Resistance and Its Viscoelastic Assessment in Critically Ill COVID-19 Patients.

Critically ill COVID-19 patients present an inflammatory and procoagulant status with a high rate of relevant macro- and microvascular thrombosis. Furthermore, high rates of heparin resistance have been described; yet, individualized anticoagulation by drug monitoring has not been sufficiently researched. We analyzed data from critically ill COVID-19 patients treated at Innsbruck Medical University Hospital with routinely adapted low-molecular-weight heparin (LMWH) doses according to anti-Xa peak levels, and regularly performed ClotPro analyses (a viscoelastic hemostatic whole blood test). A total of 509 anti-Xa peak measurements in 91 patients were categorized as below (<0.008 IU/mL/mg), within (0.008-0-012 IU/mL/mg) or above (> 0.012 IU/mL/mg) expected ranges with respect to the administered LMWH doses. Besides intergroup comparisons, correlations between anti-Xa levels and ClotPro clotting times (CTs) were performed (226 time points in 84 patients). Anti-Xa peak levels remained below the expected range in the majority of performed measurements (63.7%). Corresponding patients presented with higher C-reactive protein and D-dimer but lower antithrombin levels when compared with patients achieving or exceeding the expected range. Consequently, higher enoxaparin doses were applied in the sub-expected anti-Xa range group. Importantly, 47 (51.6%) patients switched between groups during their intensive care unit (ICU) stay. Anti-Xa levels correlated weakly with IN test CT and moderately with Russell's viper venom (RVV) test CT. Critically ill COVID-19 patients present with a high rate of LMWH resistance but with a variable LMWH response during their ICU stay. Therefore, LMWH-anti-Xa monitoring seems inevitable to achieve adequate target ranges. Furthermore, we propose the use of ClotPro's RVV test to assess the coagulation status during LMWH administration, as it correlates well with anti-Xa levels but more holistically reflects the coagulation cascade than anti-Xa activity alone.

[Effects of automated external defibrillators on hands-off intervals in lay rescuers]. / Einfluss verschiedener automatisierter externer Defibrillatoren auf die Hands-off-Intervalle von Ersthelfern.

Background: Survival chances after out-of-hospital cardiac arrests caused by hyperdynamic electric cardiac rhythms can be significantly improved by early defibrillation with automated external defibrillators (AEDs). As postulated in international guidelines, the resulting hands-off intervals should not exceed 10 s. Objectives: We investigated delay in onset of chest compressions and the length of hands-off intervals during defibrillation associated with the application of AEDs. Materials and methods: In a prospective, randomized, single-blinded observational study, the resuscitation efforts by first year medical students were analyzed in different emergency scenarios on manikins. Delay in onset of chest compressions and the length of hands-off intervals between voice prompts from four conventional devices were compared during shockable and nonshockable rhythms. Satisfaction with the device, difficulties with the application, and suggested improvements were assessed by questionnaire. Results: In a total of 70 applications, the start with thoracic compressions was delayed by a mean of 115 s. On average, the first shock was administered after 125 s in shockable heart rhythms. Perishock pauses of less than 10 s were achieved with none of the tested devices. Hands-off intervals during defibrillation differed significantly between the devices (p < 0.001). Improvements were suggested regarding marking, voice prompts, and electrodes. Conclusions: Perishock pause of less than 10 s was not achieved with any of the tested devices. Shortened and more precise voice prompts as well as more clearly arranged labeling and layout of pads are needed to simplify application, reduce delayed onset of chest compressions and shorten hands-off intervals.

Impaired fibrinolysis in critically ill COVID-19 patients.

BACKGROUND: Critically ill coronavirus disease 2019 (COVID-19) patients present with a hypercoagulable state with high rates of macrovascular and microvascular thrombosis, for which hypofibrinolysis might be an important contributing factor. METHODS: We retrospectively analysed 20 critically ill COVID-19 patients at Innsbruck Medical University Hospital whose coagulation function was tested with ClotPro® and compared with that of 60 healthy individuals at Augsburg University Clinic. ClotPro is a viscoelastic whole blood coagulation testing device. It includes the TPA test, which uses tissue factor (TF)-activated whole blood with added recombinant tissue-derived plasminogen activator (r-tPA) to induce fibrinolysis. For this purpose, the lysis time (LT) is measured as the time from when maximum clot firmness (MCF) is reached until MCF falls by 50%. We compared COVID-19 patients with prolonged LT in the TPA test and those with normal LT. RESULTS: Critically ill COVID-19 patients showed hypercoagulability in ClotPro assays. MCF was higher in the EX test (TF-activated assay), IN test (ellagic acid-activated assay), and FIB test (functional fibrinogen assay) with decreased maximum lysis (ML) in the EX test (hypofibrinolysis) and highly prolonged TPA test LT (decreased fibrinolytic response), as compared with healthy persons. COVID-19 patients with decreased fibrinolytic response showed higher fibrinogen levels, higher thrombocyte count, higher C-reactive protein levels, and decreased ML in the EX test and IN test. CONCLUSION: Critically ill COVID-19 patients have impaired fibrinolysis. This hypofibrinolytic state could be at least partially dependent on a decreased fibrinolytic response.

Response patterns of routinely measured inflammatory and coagulatory parameters in sepsis.

BACKGROUND: Sepsis is characterized by a pro-inflammatory and pro-coagulatory shift which can induce life-threatening complications. Close monitoring and risk stratification of sepsis patients is crucial for proper treatment and consequently patient outcome. Therefore, this study focuses on the response patterns of inflammatory and coagulatory parameters used in clinical routines to estimate the course of sepsis. METHODS: A total of 1,110 patients diagnosed with sepsis were retrospectively analyzed to identify response patterns for risk stratification of routine parameters measured at the peak level of C-reactive protein. Cluster analysis was used and the differences in the patient characteristics and 28-day survival were assessed. Cox proportional hazards regression model for survival stratified by the clusters was performed. RESULTS: The analyses revealed the parameters to have five distinct response patterns. These clusters reflect the etiology as well as the course of sepsis associated with different mortalities. Here, impairment of the liver plays a crucial role in the ability to appropriately respond to sepsis. Of the routinely measured parameters, C-reactive protein and antithrombin seem to be unspecific for stratification of septic patients. Adjusted for the individual clusters, survival was associated with an increase in fibrinogen (p = 0.0042), platelets (p = 0.0003) and PT (p = 0.001) as well as a decrease in leukocytes (p = 0.034). CONCLUSIONS: This study reveals that patients have distinct response patterns of inflammatory and coagulatory parameters depending on disease etiology. These patterns are associated with different mortalities although the patients have similar levels of C-reactive protein. Independently of the type of response, good coagulatory capacity seems to be crucial for patient survival.

Impact of wine manufacturing practice on the occurrence of fining agents with allergenic potential.

Proteinogenic wine fining agents are hidden allergens and could present a risk for consumers with allergies. Therefore, the European Parliament adopted Directive 2003/89/EC amending Directive 2000/13/EC to declare ingredients, contaminations and processing aids that are known to trigger allergic reactions. The Amendment Regulation (EU) 1266/2010 excluded the labelling of wines which are processed with hen's egg and products thereof until 30 June 2012 to get more scientific findings. After 1 July 2012 wine fining agents have to be declared if above 0.25 mg l(-1) (Regulation (EU) 579/2012 in conjunction with article 120 g of Regulation (EU) 1234/2007). The Organisation International de la Vigne et du Vin (OIV) advises this limit of detection (LOD) for potential allergenic residues of proteins. Wine fining agents are processing aids and according to the wine producer's knowledge will be removed after coagulation by filtration or other production steps. Due to lack of scientific data, residues of fining agents in the final product could not be excluded. In this risk assessment, highly sensitive ELISA methods for ovalbumin of known origin for wine have been developed. The objective was to investigate the presence of allergen residues in wine after certain technological treatments were applied to remove the wine fining agents. For all developed ELISA methods the LODs are in the low µg l(-1) range between 5 and 10 µg l(-1) fining agent, whereas the LOQ varies between 5 and 80 µg l(-1) fining agent. The results of the investigation of well-known wines and fining agents demonstrate that white wines fined with white or ovalbumin from hen's egg could retain allergens. The use of certain technological procedures during wine processing leads to different results. In white wine, bentonite or sheet filtration followed by sterile filtration lead to wines containing no detectable amounts of ovalbumin. In red wine, especially the final sterile filtration removes the fining agents.

Evaluation of the efficiency of enological procedures on lysozyme depletion in wine by an indirect ELISA method.

Potential residues of the potent allergen lysozyme used as a microbial stabilizing agent in wine production might pose a serious health thread to susceptible individuals. Therefore, EU legislation requires the labeling of the allergenic agent, if it is present in the final product. To allow for product testing, an indirect ELISA method to be specifically used in wine analysis was developed and validated. Furthermore, trial wines treated with defined amounts of lysozyme were subjected to an array of different filtration and other enological processing regimes in order to evaluate their potential to deplete the allergen content of the wines. By these means, processing methods ought to be identified that can be integrated in a good manufacturing practice guideline to enable wine producers to utilize lysozyme in their cellars and still provide wines free of allergenic residues. However, among the enological procedures under scrutiny, only bentonite fining proved to be capable of significantly reducing the allergenic residues.

Development of a sensitive ELISA for the detection of casein-containing fining agents in red and white wines.

Fining of wine with proteinogenic fining agents such as casein from cow's milk is a traditional and commonly used technique all over the world. Casein and other proteins from cow's milk are well-known food allergens, which pose a risk for allergic consumers. Temporary regulations exempting the labeling of milk and products thereof in wine expired. Since July 1, 2012, these fining agents have to be declared on the wine label under Regulation (EU) No. 579/2012 in conjunction to article 120g of Regulation (EU) No. 1234/2007 if exceeding the threshold of 0.25 mg/L allergenic protein. The aim of the presented study was to develop sensitive ELISA methods for the detection of casein in white and red wines and to investigate the risk of allergenic residues in fined wines. In this context it was shown that the used substance for calibration is highly relevant. Casein wine fining agents of different commercial producers were investigated by LDS-PAGE and immunoblot. In addition to casein, they contain other milk proteins, which are potentially allergic and therefore have to be incorporated in the development of antibodies for an ELISA method to be set up. An indirect ELISA for the investigation of white wine was developed. The LOD is 0.1 mg/L. For red wine the LOD is 0.2 mg/L in an indirect sandwich ELISA setup. The LOD of the indirect sandwich ELISA for white wine depends on the calibration standard. It is 0.1 mg/L for the fining agent casein and 0.01 mg/L for casein from a chemical trader. It is also shown that the use of different technological procedures during winemaking leads to no detectable amounts of casein in various wine samples.

Prospective multicenter study evaluating fecal calprotectin in adult acute bacterial diarrhea.

BACKGROUND: Every year, about 2.2 million deaths occur worldwide due to diarrhea. Reliable diagnosis of patients with acute infectious diarrhea remains a formidable challenge to the clinicians. This is the first study reporting use of fecal calprotectin in diagnosing acute diarrhea. The aim was to compare the diagnostic accuracy of fecal calprotectin, fecal lactoferrin, and guaiac-based fecal occult blood test in a diverse group of consecutive patients with acute diarrhea in which routine bacterial stool cultures and cytotoxins for Clostridium difficile were performed. METHODS: This was a prospective case-control multicenter study from January 2004 until October 2007 in 2383 consecutive patients with acute diarrhea. They provided stool samples for performing cultures. Patients with positive cultures and an equal number of matched controls with negative cultures underwent fecal occult blood test and calprotectin and lactoferrin assays. RESULTS: Calprotectin, lactoferrin, and fecal occult blood tests demonstrated sensitivity and specificity of 83% and 87%, 78% and 54%, and 38% and 85%, respectively, for diagnosing acute bacterial diarrhea. CONCLUSIONS: Calprotectin showed high correlation with bacteriologically positive infectious diarrhea compared with lactoferrin and fecal occult blood test. It may potentially revolutionize management algorithm for patients with acute diarrhea. As a screening test, calprotectin can generate results within hours to support presumptive diagnosis of infectious diarrhea, which can decide suitability of stool samples for culture.

Necrotizing enterocolitis of the neonate with Clostridium perfringens: diagnosis, clinical course, and role of alpha toxin.

The severity of the clinical course in necrotizing enterocolitis (NEC) associated with Clostridium perfringens (Cp) may support the hypothesis of a specific disease. We conducted a case control study of infants diagnosed with NEC, who underwent surgical treatment over a 7-year period. Patient histories examined characteristics of the infants, bacterial infection as well as NEC's severity, antibiotic treatment, and clinical course. Infants infected with NEC associated with Cp were compared with NEC patients without Cp. The alpha toxin from Cp type A was detected in most of the isolated strains. Cp was identified as a causative agent of NEC in nine cases. As compared with the control group (n = 32), the onset of disease was earlier in life, the clinical course more severe, and patients had a larger extent of gangrene. Portal venous gas was evident in 77% of all Cp cases, as compared to 25% in the control group. The mortality rate was 44% in the Cp group, and only 18.7% in the control group. Type A Clostridium perfringens was identified in six cases. In each isolate alpha toxin production was proven, but without any correlation to the severity of the clinical course, the extent of intestinal gangrene or mortality. In premature infants NEC in conjunction with Cp seems to be more severe than other NEC cases; it also entails higher mortality and morbidity. Alpha toxin concentrations do not correlate with the severity of the disease. Portal venous gas is highly suggestive for the diagnosis of Cp infection.

In vitro activities of faropenem, ertapenem, imipenem and meropenem against Borrelia burgdorferi s.l.

Little is known about the in vitro activity of penems and carbapenems against the spirochete Borrelia burgdorferi. Here, faropenem, ertapenem, imipenem and meropenem as well as the third-generation cephalosporin ceftriaxone and tobramycin were tested in vitro against 11 isolates of the B. burgdorferi sensu lato complex. On a microg/mL basis, ertapenem was the most potent carbapenem (minimal inhibitory concentration (MIC) range: 0.015-0.125 microg/mL), with in vitro activity comparable with that of ceftriaxone against Borrelia. These findings are supported by the results of time-kill experiments in a Borrelia afzelii skin isolate, demonstrating a >3 log10 unit (99.9%) reduction of the inoculum after 96 h of exposure to either drug at a concentration of three log2 unit dilutions above the respective MIC.

Bacterial colonization of patients undergoing international air transport: a prospective epidemiologic study.

OBJECTIVE: To investigate the incidence of multiresistant bacteria in patients treated in foreign hospitals and repatriated by international interhospital air transport. METHODS: This was a prospective epidemiologic study on patients who were hospitalized in a foreign country and repatriated to a hospital in their home country by international aeromedical transport on scheduled airlines or ambulance jets. The study was carried out by the Department of Pediatrics, Department of Medical Microbiology, University Hospital Frankfurt, Germany, and MedCall Germany, the organizing company for repatriation. One hundred and three patients, who were hospitalized abroad, required repatriation by international interhospital air transport and met the entry criteria. RESULTS: Four hundred and eighty-three swabs from brow, nose, ear, throat, groin or axilla were taken from 103 patients, mean age 62 years. They were transported from southern and eastern European countries, Morocco, Egypt, Ghana, Tunisia, Pakistan, the United States and the Bahamas to destinations in Germany, the UK, Belgium, Switzerland, the United States and Japan. Forty-four patients were in an intensive care unit (ICU) for 6.5 days (median) prior to transport. Forty patients received antibiotics at the time of repatriation. Acinetobacter, Enterobacter, Pseudomonas, Proteus, Staphylococcus aureus and Candida were found. Methicillin-resistant Staphylococcus aureus (MRSA) was found in three patients (3%), ventilated for 17 days, and treated in the ICU for a median of 23 days. In 2 patients, multiresistant Acinetobacter baumanii and multiresistant Klebsiella pneumoniae were found. CONCLUSIONS: Bacterial colonization of patients undergoing international interhospital air transport does not differ from that of patients in a European hospital. Multiresistant bacteria, especially MRSA, were found only in ICU patients. There is no elevated risk of transmission of multiresistant bacteria in this patient group compared to other patients treated in hospital, especially in ICUs.

Biological cost of rifampin resistance from the perspective of Staphylococcus aureus.

Resistance determinants that interfere with normal physiological processes in the bacterial cell usually cause a reduction in biological fitness. Fitness assays revealed that 17 of 18 in vitro-selected chromosomal mutations within the rpoB gene accounting for rifampin resistance in Staphylococcus aureus were associated with a reduction in the level of fitness. There was no obvious correlation between the level of resistance to rifampin and the level of fitness loss caused by rpoB mutations. Among 23 clinical rifampin-resistant S. aureus isolates from six countries, only seven different rpoB genotypes could be identified, whereby the mutation 481His-->Asn was present in 21 (91%) of these 23 isolates. The mutation 481His-->Asn, in turn, which confers low-level rifampin resistance on its own, was not shown to be associated with a cost of resistance in vitro. The restriction to distinct mutations that confer rifampin resistance in vivo, as demonstrated here, appears to be determined by the Darwinian fitness of the organisms.

Standardised in vitro susceptibility testing of Borrelia burgdorferi against well-known and newly developed antimicrobial agents--possible implications for new therapeutic approaches to Lyme disease.

Lyme disease represents a disorder of potentially chronic proportions, and relatively little is known about the in vivo pharmacodynamic interactions of antimicrobial agents with borreliae. So far, evidence-based drug regimens for the effective treatment of Lyme disease have not been definitively established. Moreover, therapeutic failures have been reported for almost every suitable antimicrobial agent currently available. Resistance to treatment and a protracted course of the disease, therefore, continue to pose problems for clinicians in the management of patients suffering from chronic Lyme disease. Further characterisation of the antibiotic susceptibility pattern and a better understanding of the interactions of B. burgdorferi with antimicrobial agents are urgently needed and continue to be crucial owing to considerable differences in the experimental conditions and test methods applied. The development of easily performed, new techniques for the sensitivity testing of B. burgdorferi provides the opportunity to study factors affecting the bacteriostatic and bactericidal action of recently introduced chemotherapeutic agents under more standardised conditions. For the first time, these studies provide direct evidence that, in addition to beta-lactams, macrolides, and tetracyclines which are recommended for stage-dependent treatment of Lyme borreliosis, other recently introduced substances, such as fluoroquinolones, everninomycins, and the ketolide family of antimicrobial agents, also show enhanced in vitro activity against borreliae. Some of these compounds, if effective in vivo as well, may prove to be useful agents in the treatment of certain manifestations of Lyme disease. As such, their potential role should be evaluated further by in vivo experiments and clinical trials. Finally, these antimicrobial agents may turn out to be very effective therapeutic alternatives on account of their oral availability, favourable pharmacodynamic profiles, and high tissue levels in cases where beta-lactames or tetracyclines cannot be administered without detrimental side-effects.