In vitro tests in drug hypersensitivity diagnosis

The diagnosis of a drug hypersensitivity reaction (DHR) is a challenging task because multiple and complex mechanisms are involved. Better understanding of immunologic pathomechanisms in DHRs and rapid progress in cellular-based in-vitro tests can help to adjust the correct diagnostic strategy to individual patients with different clinical manifestations of drug allergy. Thus, drug hypersensitivity diagnosis needs to rely on a combination of medical history and different in vivo and in vitro tests. In this article, the authors discuss current in vitro techniques, most recent findings, and new promising tools in the diagnosis of T-cell-mediated drug hypersensitivity.

Sleepiness and vigilance tests

Objective assessments of subjective complaints such as sleepiness, tiredness or fatigue using sleepiness and vigilance tests aim to identify its causes and to judge the fitness to drive or to work of the affected person. "Vigilance" comprises wakefulness, alertness and attention and is therefore not merely reciprocal to sleepiness. Since it is a complex phenomenon with several dimensions it is unlikely to be appropriately assessed by one single "vigilance test". One important dimension of vigilance discussed here is wakefulness with its counterpart of overt sleep and the whole spectrum of various levels in between. The transit zone between full wakefulness and overt sleep is mainly characterised by the subjective complaint of sleepiness, which cannot be measured directly. Only the consequences of reduced wakefulness such as a shortened sleep latency, slowed cognitive function and prolonged reaction time can be measured objectively. It is, therefore, more promising to combine a battery of subjective and objective tests to answer a specific question in order to achieve the most appropriate description for a given clinical or medicolegal situation. However even then we must keep in mind that many other important aspects of fitness to drive / fitness to work such as neurological, psychiatric and neuropsychological functions including risk taking behaviour are not covered by vigilance tests. A comprehensive, multidisciplinary approach is essential in such situations.

Test Blueprints - Exposing Side Effects in Execution Traces to Support Writing Unit Tests

Writing unit tests for legacy systems is a key maintenance task. When writing tests for object-oriented programs, objects need to be set up and the expected effects of executing the unit under test need to be verified. If developers lack internal knowledge of a system, the task of writing tests is non-trivial. To address this problem, we propose an approach that exposes side effects detected in example runs of the system and uses these side effects to guide the developer when writing tests. We introduce a visualization called Test Blueprint, through which we identify what the required fixture is and what assertions are needed to verify the correct behavior of a unit under test. The dynamic analysis technique that underlies our approach is based on both tracing method executions and on tracking the flow of objects at runtime. To demonstrate the usefulness of our approach we present results from two case studies.

JExample: Exploiting Dependencies Between Tests to Improve Defect Localization

To quickly localize defects, we want our attention to be focussed on relevant failing tests. We propose to improve defect localization by exploiting dependencies between tests, using a JUnit extension called JExample. In a case study, a monolithic white-box test suite for a complex algorithm is refactored into two traditional JUnit style tests and to JExample. Of the three refactorings, JExample reports five times fewer defect locations and slightly better performance (-8-12\%), while having similar maintenance characteristics. Compared to the original implementation, JExample greatly improves maintainability due the improved factorization following the accepted test quality guidelines. As such, JExample combines the benefits of test chains with test quality aspects of JUnit style testing.

In vitro allergy tests compared to intradermal testing in horses with recurrent airway obstruction

Recurrent airway obstruction (RAO) is a common condition in stabled horses characterised by small airway inflammation, airway neutrophilia and obstruction following exposure of susceptible horses to mouldy hay and straw and is thus regarded as a hypersensitivity reaction to mould spores. However, the role of IgE-mediated reactions in RAO remains unclear. The aim of the study was to investigate with a serological IgE ELISA test (Allercept), an in vitro sulfidoleukotriene (sLT) release assay (CAST) and with intradermal testing (IDT) whether serum IgE and IgE-mediated reactions against various mould, mite and pollen extracts are associated with RAO. IDT reactions were evaluated at different times in order to detect IgE-mediated immediate type reactions (type I hypersensitivity reactions, 0.5-1 h), immune complex-mediated late type reactions (type III reactions, 4-10 h) and cell-mediated delayed type reactions (type IV hypersensitivity reactions 24-48 h). In the serological test, overall the control horses displayed more positive reactions than the RAO-affected horses but the difference was not significant. Comparison of the measured IgE levels showed that the RAO-affected horses had slightly higher IgE levels against Aspergillus fumigatus than controls (35 and 16 AU, respectively, p<0.05), but all values were below the cut off (150 AU) of the test. In the sLT release assay, seven positive reactions were observed in the RAO-affected horses and four in the controls but this difference was not significant. A significantly higher proportion of late type IDT reactions was observed in RAO-affected horses compared to controls (25 of 238 possible reactions versus 12 of 238 possible reactions, respectively, p<0.05). Interestingly, four RAO-affected but none of the control horses reacted with the recombinant mould allergen A. fumigatus 8 (rAsp f 8, p<0.05), but only late phase and delayed type reactions were observed. In all three tests the majority of the positive reactions was observed with the mite extracts (64%, 74% and 88% of all positive reactions, respectively) but none of the tests showed a significant difference between RAO-affected and control animals. Our findings do not support that IgE-mediated reactions are important in the pathogenesis of RAO. Further studies are needed to investigate whether sensitisation to mite allergens is of clinical relevance in the horse and to understand the role of immune reactions against rAsp f 8.

Field performance of two rapid screening tests in active surveillance of transmissible spongiform encephalopathies in small ruminants

Recently, screening tests for monitoring the prevalence of transmissible spongiform encephalopathies specifically in sheep and goats became available. Although most countries require comprehensive test validation prior to approval, little is known about their performance under normal operating conditions. Switzerland was one of the first countries to implement 2 of these tests, an enzyme-linked immunosorbent assay (ELISA) and a Western blot, in a 1-year active surveillance program. Slaughtered animals (n = 32,777) were analyzed in either of the 2 tests with immunohistochemistry for confirmation of initial reactive results, and fallen stock samples (n = 3,193) were subjected to both screening tests and immunohistochemistry in parallel. Initial reactive and false-positive rates were recorded over time. Both tests revealed an excellent diagnostic specificity (>99.5%). However, initial reactive rates were elevated at the beginning of the program but dropped to levels below 1% with routine and enhanced staff training. Only those in the ELISA increased again in the second half of the program and correlated with the degree of tissue autolysis in the fallen stock samples. It is noteworthy that the Western blot missed 1 of the 3 atypical scrapie cases in the fallen stock, indicating potential differences in the diagnostic sensitivities between the 2 screening tests. However, an estimation of the diagnostic sensitivity for both tests on field samples remained difficult due to the low disease prevalence. Taken together, these results highlight the importance of staff training, sample quality, and interlaboratory comparison trials when such screening tests are implemented in the field.

The Costs and Predictive Factors of Bovine Respiratory Disease in Standardized Steer Tests

A retrospective study of 2,146 feedlot cattle in 17 feedlot tests from 1988 to 1997 was conducted to determine the impact of bovine respiratory disease (BRD) on veterinary treatment costs, average daily gain, carcass traits, mortality, and net profit. Morbidity caused by BRD was 20.6%. The average cost to treat each case of BRD was $12.39. Mortality rate of calves diagnosed and treated for BRD was 5.9% vs. .35% for those not diagnosed with BRD. Average daily gain differed between treated and non-treated steers during the first 28 days on feed but did not differ from 28 days to harvest. Net profit was $57.48 lower for treated steers. Eighty-two percent of this difference was due to a combination of mortality and treatment costs. Eighteen percent of the net profit difference was due to improved performance and carcass value of the non-treated steers. Data from 496 steers and heifers in nine feedlot tests were used to determine the effects of age, weaning, and use of modified live virus or killed vaccines prior to the test to predict BRD. Younger calves, non-weaned calves, and calves vaccinated with killed vaccines prior to the test had higher BRD morbidity than those that were older, weaned, or vaccinated with modified live virus vaccines, respectively. Treatment regimes that precluded relapse resulting in re-treatment prevented reduced performance and loss of carcass value. Using modified live virus vaccines and weaning calves 30 days prior to shipment reduced the incidence of BRD.

Ranking of tests for pain hypersensitivity according to their discriminative ability in chronic neck pain

BACKGROUND AND OBJECTIVES Quantitative sensory testing (QST) is widely used to investigate peripheral and central sensitization. However, the comparative performance of different QST for diagnostic or prognostic purposes is unclear. We explored the discriminative ability of different quantitative sensory tests in distinguishing between patients with chronic neck pain and pain-free control subjects and ranked these tests according to the extent of their association with pain hypersensitivity. METHODS We performed a case-control study in 40 patients and 300 control subjects. Twenty-six tests, including different modalities of pressure, heat, cold, and electrical stimulation, were used. As measures of discrimination, we estimated receiver operating characteristic curves and likelihood ratios. RESULTS The following quantitative sensory tests displayed the best discriminative value: (1) pressure pain threshold at the site of the most severe neck pain (fitted area under the receiver operating characteristic curve, 0.92), (2) reflex threshold to single electrical stimulation (0.90), (3) pain threshold to single electrical stimulation (0.89), (4) pain threshold to repeated electrical stimulation (0.87), and (5) pressure pain tolerance threshold at the site of the most severe neck pain (0.86). Only the first 3 could be used for both ruling in and out pain hypersensitivity. CONCLUSIONS Pressure stimulation at the site of the most severe pain and parameters of electrical stimulation were the most appropriate QST to distinguish between patients with chronic neck pain and asymptomatic control subjects. These findings may be used to select the tests in future diagnostic and longitudinal prognostic studies on patients with neck pain and to optimize the assessment of localized and spreading sensitization in chronic pain patients.

A connection between two nonparametric tests for perfect ranking in balanced ranked set sampling

In this note, we show that an extension of a test for perfect ranking in a balanced ranked set sample given by Li and Balakrishnan (2008) to the multi-cycle case turns out to be equivalent to the test statistic proposed by Frey et al. (2007). This provides an alternative interpretation and motivation for their test statistic.

Through the European Looking Glass: Citizenship Tests in the USA, Australia, and Canada

Discussing new or recently reformed citizenship tests in the USA, Australia, and Canada, this article asks whether they amount to a restrictive turn of new world citizenship, similar to recent developments in Europe. I argue that elements of a restrictive turn are noticeable in Australia and Canada, but only at the level of political rhetoric, not of law and policy, which remain liberal and inclusive. Much like in Europe, the restrictive turn is tantamount to Muslims and Islam moving to the center of the integration debate.

Non-phenotypic tests to detect and characterize antibiotic resistance mechanisms in Enterobacteriaceae

In the past 2 decades, we have observed a rapid increase of infections due to multidrug-resistant Enterobacteriaceae. Regrettably, these isolates possess genes encoding for extended-spectrum β-lactamases (e.g., blaCTX-M, blaTEM, blaSHV) or plasmid-mediated AmpCs (e.g., blaCMY) that confer resistance to last-generation cephalosporins. Furthermore, other resistance traits against quinolones (e.g., mutations in gyrA and parC, qnr elements) and aminoglycosides (e.g., aminoglycosides modifying enzymes and 16S rRNA methylases) are also frequently co-associated. Even more concerning is the rapid increase of Enterobacteriaceae carrying genes conferring resistance to carbapenems (e.g., blaKPC, blaNDM). Therefore, the spread of these pathogens puts in peril our antibiotic options. Unfortunately, standard microbiological procedures require several days to isolate the responsible pathogen and to provide correct antimicrobial susceptibility test results. This delay impacts the rapid implementation of adequate antimicrobial treatment and infection control countermeasures. Thus, there is emerging interest in the early and more sensitive detection of resistance mechanisms. Modern non-phenotypic tests are promising in this respect, and hence, can influence both clinical outcome and healthcare costs. In this review, we present a summary of the most advanced methods (e.g., next-generation DNA sequencing, multiplex PCRs, real-time PCRs, microarrays, MALDI-TOF MS, and PCR/ESI MS) presently available for the rapid detection of antibiotic resistance genes in Enterobacteriaceae. Taking into account speed, manageability, accuracy, versatility, and costs, the possible settings of application (research, clinic, and epidemiology) of these methods and their superiority against standard phenotypic methods are discussed.