Profiling of Volatile Organic Compounds in Exhaled Breath As a Strategy to Find Early Predictive Signatures of Asthma in Children

Wheezing is one of the most common respiratory symptoms in preschool children under six years old. Currently, no tests are available that predict at early stage who will develop asthma and who will be a transient wheezer. Diagnostic tests of asthma are reliable in adults but the same tests are difficult to use in children, because they are invasive and require active cooperation of the patient. A non-invasive alternative is needed for children. Volatile Organic Compounds (VOCs) excreted in breath could yield such non-invasive and patient-friendly diagnostic. The aim of this study was to identify VOCs in the breath of preschool children (inclusion at age 2–4 years) that indicate preclinical asthma. For that purpose we analyzed the total array of exhaled VOCs with Gas Chromatography time of flight Mass Spectrometry of 252 children between 2 and 6 years of age. Breath samples were collected at multiple time points of each child. Each breath-o-gram contained between 300 and 500 VOCs; in total 3256 different compounds were identified across all samples. Using two multivariate methods, Random Forests and dissimilarity Partial Least Squares Discriminant Analysis, we were able to select a set of 17 VOCs which discriminated preschool asthmatic children from transient wheezing children. The correct prediction rate was equal to 80% in an independent test set. These VOCs are related to oxidative stress caused by inflammation in the lungs and consequently lipid peroxidation. In conclusion, we showed that VOCs in the exhaled breath predict the subsequent development of asthma which might guide early treatment.     

Freezing of tissue-limits for the autoradiographic localization of diffusible substances

Frozen thin sections and sections from freeze-dried and embedded tissue are used for the autoradiographic localization of diffusible substances at the electron microscope level. The presence of ice crystals in such sections may limit the autoradiographic resolution. Ice crystals are formed during freezing and may grow during subsequent processing of tissue. The contribution of ice crystal growth to the final image was estimated by measuring the distribution of the ice crystal sizes in freeze-etch replicas and in sections from freeze-dried and embedded tissues. A surface layer (10-15 mu) without visible ice crystals was present in both preparations. Beneath this surface layer the diameter of ice crystals increased towards the interior with the same relationship between crystal size and distance from the surface in the freeze-etch preparation as in the freeze-dry preparation. Ice crystal growth occurring during a much longer time during freeze-drying compared to freeze-etching does not significantly contribute to the final image in the electron microscope. The formation of ice crystals during freezing determines to a large extent the image (and therefore the autoradiographic resolution) of freeze-dry preparations and this probably holds also for thin cryosections of which examples are given.     

Complex phylogenetic distribution of a non-canonical genetic code in green algae

Background  

A non-canonical nuclear genetic code, in which TAG and TAA have been reassigned from stop codons to glutamine, has evolved independently in several eukaryotic lineages, including the ulvophycean green algal orders Dasycladales and Cladophorales. To study the phylogenetic distribution of the standard and non-canonical genetic codes, we generated sequence data of a representative set of ulvophycean green algae and used a robust green algal phylogeny to evaluate different evolutionary scenarios that may account for the origin of the non-canonical code.     

Effect of etanercept in polymyalgia rheumatica: a randomized controlled trial

Introduction  

To elucidate in polymyalgia rheumatica (PMR) the role of tumor necrosis factor (TNF) α and the therapeutic potential of blockade with soluble TNF-α receptor, we carried out the first randomized controlled trial with etanercept in PMR.     

Catch and measure–mass spectrometry‐based immunoassays in biomarker research

Mass spectrometry-based (MS) methods are effective tools for discovering protein biomarker candidates that can differentiate between physiological and pathophysiological states. Promising candidates are validated in studies comprising large patient cohorts. Here, targeted protein analytics are used to increase sample throughput. Methods involving antibodies, such as sandwich immunoassays or Western blots, are commonly applied at this stage. Highly-specific and sensitive mass spectrometry-based immunoassays that have been established in recent years offer a suitable alternative to sandwich immunoassays for quantifying proteins. Mass Spectrometric ImmunoAssays (MSIA) and Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA/iMALDI) are two prominent types of MS-based immunoassays in which the capture is done either at the protein or the peptide level. We present an overview of these emerging types of immunoassays and discuss their suitability for the discovery and validation of protein biomarkers. This article is part of a Special Issue entitled: Biomarkers: A Proteomic Challenge.     

Biological Instability in a Chlorinated Drinking Water Distribution Network

The purpose of a drinking water distribution system is to deliver drinking water to the consumer, preferably with the same quality as when it left the treatment plant. In this context, the maintenance of good microbiological quality is often referred to as biological stability, and the addition of sufficient chlorine residuals is regarded as one way to achieve this. The full-scale drinking water distribution system of Riga (Latvia) was investigated with respect to biological stability in chlorinated drinking water. Flow cytometric (FCM) intact cell concentrations, intracellular adenosine tri-phosphate (ATP), heterotrophic plate counts and residual chlorine measurements were performed to evaluate the drinking water quality and stability at 49 sampling points throughout the distribution network. Cell viability methods were compared and the importance of extracellular ATP measurements was examined as well. FCM intact cell concentrations varied from 5×10³ cells mL⁻¹ to 4.66×10⁵ cells mL⁻¹ in the network. While this parameter did not exceed 2.1×10⁴ cells mL⁻¹ in the effluent from any water treatment plant, 50% of all the network samples contained more than 1.06×10⁵ cells mL⁻¹. This indisputably demonstrates biological instability in this particular drinking water distribution system, which was ascribed to a loss of disinfectant residuals and concomitant bacterial growth. The study highlights the potential of using cultivation-independent methods for the assessment of chlorinated water samples. In addition, it underlines the complexity of full-scale drinking water distribution systems, and the resulting challenges to establish the causes of biological instability.     

Embryotoxicity of sex steroidal hormone combinations in nonhuman primates: I. Norethisterone acetate + ethinylestradiol and progesterone + estradiol benzoate (Macaca mulatta, Macaca fascicularis, and Papio cynocephalus)

Two sex steroid hormone combinations which have been used clinically as tests for detection of early pregnancy were examined for embryotoxic effects in macaques and baboons. Norethisterone acetate and ethinyl estradiol (NEA + EE) were orally administered to rhesus and cynomolgus monkeys and baboons at dosages ranging from one to 1,000 times the human dose equivalent (HDE) during days 20-50 of pregnancy. Progesterone and estradiol benzoate (P + EB) were delivered by two to six intramuscular injections to rhesus and cynomolgus monkeys between gestational days 20 and 35 at 0.1-25 X HDE. Fetuses were examined following cesarean section at 100 +/- 2 days (NEA + EE) or at term (P + EB). The results showed increased embryolethality over controls at 100-1,000 X HDE (NEA + EE) and at 10 and 25 X HDE (P + EB). Besides growth retardation, isolated cases of minor nongenital malformations were observed only in cynomolgus monkeys following treatment with both hormone combinations mainly at embryolethal dose levels and were considered spontaneous in nature. Virilization of female cynomolgus fetuses following NEA + EE treatment was manifested as two cases of clitoral enlargement in the 300 X HDE group and two cases of increased anogenital distance with reduced vaginal opening in the 1,000 X HDE group. The highest dose of NEA + EE was also maternally toxic, as two maternal deaths occurred at the end of the treatment period. One dead female cynomolgus fetus exposed to P + EB (10 X HDE) also exhibited masculinized external genitalia.(ABSTRACT TRUNCATED AT 250 WORDS)     

Species Diversity,Phylogeny and Large Scale Biogeographic Patterns of the Genus Padina (Phaeophyceae,Dictyotales)

The brown algal genus Padina (Dictyotales, Phaeophyceae) is distributed worldwide in tropical and temperate seas. Global species diversity and distribution ranges, however, remain largely unknown. Species‐level diversity was reassessed using DNA‐based, algorithmic species delineation techniques based on cox3 and rbcL sequence data from 221 specimens collected worldwide. This resulted in estimates ranging from 39 to 61 putative species (ESUs), depending on the technique as well as the locus. We discuss the merits, potential pitfalls, and evolutionary and biogeographic significance of algorithmic species delineation. We unveil patterns whereby ESUs are in all but one case restricted to either the Atlantic or Indo‐Pacific Ocean. Within ocean basins we find evidence for the vast majority of ESUs to be confined to a single marine realm. Exceptions, whereby ESUs span up to three realms, are located in the Indo‐Pacific Ocean. Patterns of range‐restricted species likely arise by repeated founder events and subsequent peripatric speciation, hypothesized to dominate speciation mechanisms for coastal marine organisms in the Indo‐Pacific. Using a three‐gene (cox3, psaA and rbcL), relaxed molecular clock phylogenetic analysis we estimated divergence times, providing a historical framework to interpret biogeographic patterns.