Sensitive and rapid detection of ganciclovir resistance by PCR based MALDI-TOF analysis

Cytomegalovirus (CMV) infection is associated with significant morbidity and mortality in transplant recipients. Resistance against ganciclovir is increasingly observed. According to current guidelines, direct drug resistance testing is not always performed due to high costs and work effort, even when resistance is suspected.

Rapid assessment of cerebral autoregulation by near-infrared spectroscopy and a single dose of phenylephrine

Rapid bedside determination of cerebral blood pressure autoregulation (AR) may improve clinical utility. We tested the hypothesis that cerebral Hb oxygenation (HbDiff) and cerebral Hb volume (HbTotal) measured by near-infrared spectroscopy (NIRS) would correlate with cerebral blood flow (CBF) after single dose phenylephrine (PE). Critically ill patients requiring artificial ventilation and arterial lines were eligible. During rapid blood pressure rise induced by i.v. PE bolus, ΔHbDiff and ΔHbTotal were calculated by subtracting values at baseline (normotension) from values at peak blood pressure elevation (hypertension). With the aid of NIRS and bolus injection of indocyanine green, relative measures of CBF, called blood flow index (BFI), were determined during normotension and during hypertension. BFI during hypertension was expressed as percentage from BFI during normotension (BFI%). Autoregulation indices (ARIs) were calculated by dividing BFI%, ΔHbDiff, and ΔHbTotal by the concomitant change in blood pressure. In 24 patients (11 newborns and 13 children), significant correlations between BFI% and ΔHbDiff (or ΔHbTotal) were found. In addition, the associations between Hb-based ARI and BFI%-based ARI were significant with correlation coefficients of 0.73 (or 0.72). Rapid determination of dynamic AR with the aid of cerebral Hb signals and PE bolus seems to be reliable.

Diagnostic value of animal-side antibody assays for rapid detection of Mycobacterium bovis or Mycobacterium microti infection in South American camelids

Tuberculosis (TB) in South American camelids (SAC) is caused by Mycobacterium bovis or Mycobacterium microti. Two serological methods, rapid testing (RT) and the dual-path platform (DPP) assay, were evaluated using naturally infected SAC. The study population included 156 alpacas and 175 llamas in Great Britain, Switzerland, and the United States. TB due to M. bovis (n = 44) or M. microti (n = 8) in 35 alpacas and 17 llamas was diagnosed by gross pathology examination and culture. Control animals were from herds with no TB history. The RT and the DPP assay showed sensitivities of 71% and 74%, respectively, for alpacas, while the sensitivity for llamas was 77% for both assays. The specificity of the DPP assay (98%) was higher than that of RT (94%) for llamas; the specificities of the two assays were identical (98%) for alpacas. When the two antibody tests were combined, the parallel-testing interpretation (applied when either assay produced a positive result) enhanced the sensitivities of antibody detection to 89% for alpacas and 88% for llamas but at the cost of lower specificities (97% and 93%, respectively), whereas the serial-testing interpretation (applied when both assays produced a positive result) maximized the specificity to 100% for both SAC species, although the sensitivities were 57% for alpacas and 65% for llamas. Over 95% of the animals with evidence of TB failed to produce skin test reactions, thus confirming concerns about the validity of this method for testing SAC. The findings suggest that serological assays may offer a more accurate and practical alternative for antemortem detection of camelid TB.

Rapid downregulation of innate immune cells, interleukin-12 and interleukin-23 in generalized pustular psoriasis with infliximab in combination with acitretin

Generalized pustular psoriasis (GPP) is a severe inflammatory disease characterized by recurrent eruptions of sterile pustules on erythematous skin. Although tumor necrosis factor (TNF) antagonists may lead to a rapid resolution of GPP, the mechanism of action of these agents remains to be investigated. Here, we sought to evaluate markers of immune response in the skin of a patient who experienced a rapid amelioration of GPP after treatment with infliximab and acitretin.

A new microbiopsy system enables rapid preparation of tissue for high-pressure freezing

A microbiopsy system was developed to overcome long sampling times for tissues before they are cryo-fixed by high-pressure freezing. A commercially available biopsy gun was adapted to the needs of small-organ excisions, and biopsy needles were modified to allow small samples (0.6 mm x 1.2 mm x 0.3 mm) to be taken. Specimen platelets with a central slot of the same dimensions as the biopsy are used. A self-made transfer device (in the meantime optimized by Leica-Microsystems [Vienna, Austria]) coordinates the transfer of the excised sample from the biopsy needle into the platelet slot and the subsequent loading in a specimen holder, which is then introduced into a high-pressure freezer (Leica EM PACT; Leica Microsystems, Vienna, Austria). Thirty seconds preparation time is needed from excision until high-pressure freezing. Brain, liver, kidney and muscle excisions of anesthetised rats are shown to be well frozen.

Rapid typing of Moraxella catarrhalis subpopulations based on outer membrane proteins using mass spectrometry

Moraxella catarrhalis is a major mucosal pathogen of the human respiratory tract both in children and in adults. Two subpopulations of this organism have been described that differ in 16S rRNA gene sequence and virulence traits. Three 16S rRNA types have been defined. 2-DE followed by protein identification by MS revealed significant differences in the outer membrane protein (OMP) patterns of each M. catarrhalis 16S rRNA type. Approximately 130 features were detected on the 2-DE map of each M. catarrhalis 16S rRNA type. However, only 50 features were expressed by all strains. Furthermore, direct profiling of isolated OMP using MALDI-TOF MS resulted in a characteristic spectral fingerprint for each 16S rRNA type. Fingerprints remained identical when intact cells instead of isolated OMP were analyzed. This finding suggests that the source of desorbed ions is the outer membrane. Based on the fingerprint we were able to assign 18 well-characterized clinical M. catarrhalis isolates to the correct subpopulation. Therefore, MALDI-TOF of intact M. catarrhalis provides a rapid and robust tool for M. catarrhalis strain typing that could be applied in epidemiological studies.

Rapid endocytosis of copper-zinc superoxide dismutase into human endothelial cells: role for its vascular activity

Cytosolic CuZn-SOD (SOD1) is a dimeric, carbohydrate-free enzyme with a molecular weight of about 32 kDa and also circulates in human blood plasma. Due to its molecular mass it has been believed that the enzyme cannot penetrate the cell membrane. Here we report that rapid endocytosis of FITC-CuZn-SOD into human endothelial cells occurs within 5 min. Moreover, relaxation of rat aortic rings in response to CuZn-SOD is associated with a lag time of 45-60 s and only observed in the presence of intact endothelial cells. The results indicate acute and rapid endothelial cell endocytosis of CuZn-SOD, possibly via activation of a receptor-mediated pathway. Intracellular uptake via endocytosis may contribute to the vascular effects of CuZn-SOD, including vasodilation, and is likely to play a role in regulation of vascular tone and diseases such as atherosclerosis.

Rapid measurement of S-100B serum protein levels by Elecsys S100 immunoassay in patients undergoing carotid artery stenting or endarterectomy

OBJECTIVES:: This study was designed to apply the rapid Elecsys(R) S100 immunoassay for real-time measurement of S100 protein serum levels indicating acute brain damage in patients undergoing carotid artery stenting (CAS) or endarterectomy (CEA). DESIGN AND METHODS:: Data of 14 CAS patients were compared to those of 43 CEA and 14 control patients undergoing coronary angiography (CA). S100 serum levels were measured by the full-automatic Elecsys(R) S100 immunoassay and compared to those obtained by the well-established LIA-mat(R) S100 system. RESULTS:: In contrast to CAS and CA patients, median S100 serum levels of CEA patients significantly increased to 0.24 ng/mL before declamping, but subsequently returned to baseline. Three CEA patients with neurological deficits showed sustained elevated S100 levels 6 h after extubation. Absolute S100 values were not significantly different between the two methods. Bland-Altman plot analyses displayed a good agreement, mostly indicating slightly smaller values applying the Elecsys(R) S100 system. CONCLUSIONS:: The Elecsys(R) S100 system appears to be suitable for rapid real-time detection of neurological deficits in patients undergoing CAS and CEA. Persistent elevations of Elecsys(R) S100 levels during CEA were associated with prolonged neurological disorders, whereas transient increases seem to represent impaired blood-brain barrier integrity without neurological deficits.