Detection of Human Bocavirus mRNA in Respiratory Secretions Correlates with High Viral Load and Concurrent Diarrhea

Human bocavirus (HBoV) is a parvovirus recently identified in association with acute respiratory infections (ARI). Despite its worldwide occurrence, little is known on the pathogenesis of HBoV infections. In addition, few systematic studies of HBoV in ARI have been conducted in Latin America. Therefore, in order to test whether active viral replication of human bocavirus is associated with respiratory diseases and to understand the clinical impact of this virus in patients with these diseases, we performed a 3-year retrospective hospital-based study of HBoV in outpatients and inpatients with symptoms of Acute Respiratory Infections (ARI) in Brazil. Nasopharyngeal aspirates (NPAs) from 1015 patients with respiratory symptoms were tested for HBoV DNA by PCR. All samples positive for HBoV were tested by PCR for all other respiratory viruses, had HBoV viral loads determined by quantitative real time PCR and, when possible, were tested by RT-PCR for HBoV VP1 mRNA, as evidence of active viral replication. HBoV was detected in 4.8% of patients, with annual rates of 10.0%, 3.0% and 3.0% in 2005, 2006 and 2007, respectively. The range of respiratory symptoms was similar between HBoV-positive and HBoV-negative ARI patients. However, a higher rate of diarrhea was observed in HBoV-positive patients. High HBoV viral loads (> 10(8) copies/mL) and diarrhea were significantly more frequent in patients with exclusive infection by HBoV and in patients with detection of HBoV VP1 mRNA than in patients with viral co-infection, detected in 72.9% of patients with HBoV. In summary, our data demonstrated that active HBoV replication was detected in a small percentage of patients with ARI and was correlated with concurrent diarrhea and lack of other viral co-infections.

Respiratory allergy to Blomia tropicalis: Immune response in four syngeneic mouse strains and assessment of a low allergen-dose, short-term experimental model

Background: The dust mite Blomia tropicalis is an important source of aeroallergens in tropical areas. Although a mouse model for B. tropicalis extract (BtE)-induced asthma has been described, no study comparing different mouse strains in this asthma model has been reported. The relevance and reproducibility of experimental animal models of allergy depends on the genetic background of the animal, the molecular composition of the allergen and the experimental protocol. Objectives: This work had two objectives. The first was to study the anti-B. tropicalis allergic responses in different mouse strains using a short-term model of respiratory allergy to BtE. This study included the comparison of the allergic responses elicited by BtE with those elicited by ovalbumin in mice of the strain that responded better to BtE sensitization. The second objective was to investigate whether the best responder mouse strain could be used in an experimental model of allergy employing relatively low BtE doses. Methods: Groups of mice of four different syngeneic strains were sensitized subcutaneously with 100 mu g of BtE on days 0 and 7 and challenged four times intranasally, at days 8, 10, 12, and 14, with 10 mu g of BtE. A/J mice, that were the best responders to BtE sensitization, were used to compare the B. tropicalis-specific asthma experimental model with the conventional experimental model of ovalbumin (OVA)-specific asthma. A/J mice were also sensitized with a lower dose of BtE. Results: Mice of all strains had lung inflammatory-cell infiltration and increased levels of anti-BtE IgE antibodies, but these responses were significantly more intense in A/J mice than in CBA/J, BALB/c or C57BL/6J mice. Immunization of A/J mice with BtE induced a more intense airway eosinophil influx, higher levels of total IgE, similar airway hyperreactivity to methacholine but less intense mucous production, and lower levels of specific IgE, IgG1 and IgG2 antibodies than sensitization with OVA. Finally, immunization with a relatively low BtE dose (10 mu g per subcutaneous injection per mouse) was able to sensitize A/J mice, which were the best responders to high-dose BtE immunization, for the development of allergy-associated immune and lung inflammatory responses. Conclusions: The described short-term model of BtE-induced allergic lung disease is reproducible in different syngeneic mouse strains, and mice of the A/J strain was the most responsive to it. In addition, it was shown that OVA and BtE induce quantitatively different immune responses in A/J mice and that the experimental model can be set up with low amounts of BtE.

Monitoring stress tolerance and occurrences of upper respiratory illness in basketball players by means of psychometric tools and salivary biomarkers

The aim of the study was to evaluate the possible relationships between stress tolerance, training load, banal infections and salivary parameters during 4 weeks of regular training in fifteen basketball players. The Daily Analysis of Life Demands for Athletes` questionnaire (sources and symptoms of stress) and the Wisconsin Upper Respiratory Symptom Survey were used on a weekly basis. Salivary cortisol and salivary immunoglobulin A (SIgA) were collected at the beginning (before) and after the study, and measured by enzyme-linked immunosorbent assay (ELISA). Ratings of perceived exertion (training load) were also obtained. The results from ANOVA with repeated measures showed greater training loads, number of upper respiratory tract infection episodes and negative sensation to both symptoms and sources of stress, at week 2 (p < 0.05). Significant increases in cortisol levels and decreases in SIgA secretion rate were noted (before to after). Negative sensations to symptoms of stress at week 4 were inversely and significantly correlated with SIgA secretion rate. A positive and significant relationship between sources and symptoms of stress at week 4 and cortisol levels were verified. In summary, an approach incorporating in conjunction psychometric tools and salivary biomarkers could be an efficient means of monitoring reaction to stress in sport. Copyright (C) 2010 John Wiley & Sons, Ltd.

Registration of temporal sequences of coronal and sagittal MR images through respiratory patterns

This work discusses the determination of the breathing patterns in time sequence of images obtained from magnetic resonance (MR) and their use in the temporal registration of coronal and sagittal images. The registration is made without the use of any triggering information and any special gas to enhance the contrast. The temporal sequences of images are acquired in free breathing. The real movement of the lung has never been seen directly, as it is totally dependent on its surrounding muscles and collapses without them. The visualization of the lung in motion is an actual topic of research in medicine. The lung movement is not periodic and it is susceptible to variations in the degree of respiration. Compared to computerized tomography (CT), MR imaging involves longer acquisition times and it is preferable because it does not involve radiation. As coronal and sagittal sequences of images are orthogonal to each other, their intersection corresponds to a segment in the three-dimensional space. The registration is based on the analysis of this intersection segment. A time sequence of this intersection segment can be stacked, defining a two-dimension spatio-temporal (2DST) image. The algorithm proposed in this work can detect asynchronous movements of the internal lung structures and lung surrounding organs. It is assumed that the diaphragmatic movement is the principal movement and all the lung structures move almost synchronously. The synchronization is performed through a pattern named respiratory function. This pattern is obtained by processing a 2DST image. An interval Hough transform algorithm searches for synchronized movements with the respiratory function. A greedy active contour algorithm adjusts small discrepancies originated by asynchronous movements in the respiratory patterns. The output is a set of respiratory patterns. Finally, the composition of coronal and sagittal image pairs that are in the same breathing phase is realized by comparing of respiratory patterns originated from diaphragmatic and upper boundary surfaces. When available, the respiratory patterns associated to lung internal structures are also used. The results of the proposed method are compared with the pixel-by-pixel comparison method. The proposed method increases the number of registered pairs representing composed images and allows an easy check of the breathing phase. (C) 2010 Elsevier Ltd. All rights reserved.

Classical and alternative components of the mitochondrial respiratory chain in pathogenic fungi as potential therapeutic targets

The frequency of opportunistic fungal infection has increased drastically, mainly in patients who are immunocompromised due to organ transplant, leukemia or HIV infection. In spite of this, only a few classes of drugs with a limited array of targets, are available for antifungal therapy. Therefore, more specific and less toxic drugs with new molecular targets is desirable for the treatment of fungal infections. In this context, searching for differences between mitochondrial mammalian hosts and fungi in the classical and alternative components of the mitochondrial respiratory chain may provide new potential therapeutic targets for this purpose.

Changes in neutrophil surface receptor expression, degranulation, and respiratory burst activity following moderate and high intensity exercise

Intense exercise stimulates the systemic release of a variety of factors that alter neutrophil surface receptor expression and functional activity. These alterations may influence resistance to infection after intense exercise. The aim of this study was to examine the influence of exercise intensity on neutrophil receptor expression, degranulation (measured by plasma and intracellular myeloperoxidase concentrations), and respiratory burst activity. Ten well-trained male runners ran on a treadmill for 60 min at 60% [moderate-intensity exercise (MI)] and 85% maximal oxygen consumption [high-intensity exercise (HI)]. Blood was drawn immediately before and after exercise and at 1 h postexercise. Immediately after HI, the expression of the neutrophil receptor CD16 was significantly below preexercise values (P < 0.01), whereas MI significantly reduced CD35 expression below preexercise values (P < 0.05). One hour after exercise at both intensities, there was a significant decline in CD11b expression (P < 0.05) and a further decrease in CD16 expression compared with preexercise values (P < 0.01). CD16 expression was lower 1 h after HI than 1 h after MI (P < 0.01). Immediately after HI, intracellular myeloperoxidase concentration was less than preexercise values (P < 0.01), whereas plasma myeloperoxidase concentration was greater (P < 0.01), indicating that HI stimulated neutrophil degranulation. Plasma myeloperoxidase concentration was higher immediately after HI than after MI (P < 0.01). Neutrophil respiratory burst activity increased after HI (P < 0.01). In summary, both MI and HI reduced neutrophil surface receptor expression. Although CD16 expression was reduced to a greater extent after HI, this reduction did not impair neutrophil degranulation and respiratory burst activity.

Children’s respiratory health and daily particulate levels in ten non-urban communities

We conducted a study to assess the association between the acute respiratory health of children and the levels of particulates in communities near and away from active opencast coal mines. The study enrolled children aged 1–11 years from the general population of five socioeconomically matched pairs of nonurban communities in northern England. Diaries of respiratory events were collected for 1405 children, and information was collected on the consultations of 2442 children with family/general practitioners over the 6-week study periods during 1996–1997, with concurrent monitoring of particulate levels. The associations found between daily PM10 levels and respiratory symptoms were frequently small and positive and sometimes varied between communities. The magnitude of these associations were in line with those from previous studies, even though daily particulate levels were low, and the children were drawn from the general population, rather than from the population with respiratory problems. The associations among asthma reliever use, consultations with general practitioners, and daily particulate levels were of a similar strength but estimated less precisely. The strength of association between all respiratory health measures and particulate levels was similar in communities near and away from opencast coal mining sites.

Nest temperature and respiratory gases during natural incubation in the broad-shelled river turtle, Chelodina expansa (Testudinata : Chelidae)

Temperature was monitored in three natural nests, and oxygen and carbon dioxide partial pressure monitored in one natural nest of the broad-shelled river turtle, Chelodina expansa, throughout incubation. Nest temperature decreased after nest construction in autumn, remained low during winter and gradually increased in spring to a maximum in summer. In a nest where temperature was recorded every hour, temperature typically fluctuated through a 2 degrees C cycle on a daily basis throughout the entire incubation period, and the nest always heated faster than it cooled. Oxygen and carbon dioxide partial pressures in this nest were similar to soil oxygen and carbon dioxide partial pressures for the first 5 months of incubation, but nest respiratory gas tensions deviated from the surrounding soil over the last three months of incubation. Nest respiratory gas tensions were not greatly different from those in the atmosphere above the ground except after periods of rain. After heavy rain during the last 3 months of incubation the nest became moderately hypoxic (P-O2 similar to 100 Torr) and hypercapnic (P-CO2 similar to 50 Torr) for several successive days. These short periods of hypoxia and hypercapnia were not lethal.

New techniques for biopsy and culture of human olfactory epithelial neurons

Objective: To improve the success of culturing olfactory neurons from human nasal mucosa by investigating the intranasal distribution of the olfactory epithelium and devising new techniques for growing human olfactory epithelium in vitro. Design: Ninety-seven biopsy specimens were obtained from 33 individuals, aged 21 to 74 years, collected from 6 regions of the nasal cavity. Each biopsy specimen was bisected, and 1 piece was processed for immunohistochemistry or electron microscopy while the other piece was dissected further for explant culture. Four culture techniques were performed, including whole explants and explanted biopsy slices. Five days after plating, neuronal differentiation was induced by means of a medium that contained basic fibroblast growth factor. After another 5 days, cultures were processed for immunocytochemical analysis. Results: The probability of finding olfactory epithelium in a biopsy specimen ranged from 30% to 76%, depending on its location. The dorsoposterior regions of the nasal septum and the superior turbinate provided the highest probability, but, surprisingly, olfactory epithelium was also found anteriorly and ventrally on both septum and turbinates. A new method of culturing the olfactory epithelium was devised. This slice culture technique improved the success rate for generating olfactory neurons from 10% to 90%. Conclusions: This study explains and overcomes most of the variability in the success in observing neurogenesis in cultures of adult human olfactory epithelium. The techniques presented here make the human olfactory epithelium a useful model for clinical research into certain olfactory dysfunctions and a model for the causes of neurodevelopmental and neurodegenerative diseases.

Errors in lamina growth of primary olfactory axons in the rat and mouse olfactory bulb

In the adult olfactory nerve pathway of rodents, each primary olfactory axon forms a terminal arbor in a single glomerulus in the olfactory bulb. During development, axons are believed to project directly to and terminate precisely within a glomerulus without any exuberant growth or mistargeting. To gain insight into mechanisms underlying this process, the trajectories of primary olfactory axons during glomerular formation were studied in the neonatal period. Histochemical staining of mouse olfactory bulb sections with the lectin Dolichos biflorus-agglutinin revealed that many olfactory axons overshoot the glomerular layer and course into the deeper laminae of the bulb in the early postnatal period. Single primary olfactory axons were anterogradely labelled either with the lipophilic carbocyanine dye, 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), or with horseradish peroxidase (HRP) by localized microinjections into the nerve fiber layer of the rat olfactory bulb. Five distinct trajectories of primary olfactory axons were observed in DLI-labelled preparations at postnatal day 1.5 (P1.5). Axons either coursed directly to and terminated specifically within a glomerulus, branched before terminating in a glomerulus, bypassed glomeruli and entered the underlying external plexiform layer, passed through the glomerular layer with side branches into glomeruli, or branched into more than one glomerulus. HRP-labelled axon arbors from eight postnatal ages were reconstructed by camera lucida and were used to determine arbor length, arbor area, and arbor branch number. Whereas primary olfactory axons display errors in laminar targeting in the mammalian olfactory bulb, axon arbors typically achieve their adult morphology without exuberant growth. Many olfactory axons appear not to recognize appropriate cues to terminate within the glomerular layer during the early postnatal period. However, primary olfactory axons exhibit precise targeting in the glomerular layer after P5.5, indicating temporal differences in either the presence of guidance cues or the ability of axons to respond to these cues. (C) 1999 Wiley-Liss, Inc.

Dynamic spatiotemporal expression patterns of neurocan and phosphacan indicate diverse roles in the developing and adult mouse olfactory system

The chondroitin sulfate proteoglycans neurocan and phosphacan are believed to modulate neurite outgrowth by binding to cell adhesion molecules, tenascin, and the differentiation factors heparin-binding growth-associated molecule and amphoterin. To assess the role of these chondroitin sulfate proteoglycans in the olfactory system, we describe here their expression patterns during both embryonic and postnatal development in the mouse. Immunoreactivity for neurocan was first detected in primary olfactory neurons at embryonic day 11.5 (E11.5). Neurocan was expressed by primary olfactory axons as they extended toward the rostral pole of the telencephalon as well as by their arbors in glomeruli after they contacted the olfactory bulb. The role of neurocan was examined by growing olfactory neurons on an extracellular matrix substrate containing neurocan or on extracellular matrix in the presence of soluble neurocan. In both cases, neurocan strongly promoted neurite outgrowth. These results suggest that neurocan supports the growth of primary olfactory axons through the extracellular matrix as they project to the olfactory bulb during development. Phosphacan, unlike neurocan, was present within the mesenchyme surrounding the E11.5 and E12.5 nasal cavity. This expression decreased at E13.5, concomitant with a transient appearance of phosphacan in nerve fascicles. Within the embryonic olfactory bulb, phosphacan was localised to the external and internal plexiform layers. However, during early postnatal development phosphacan was concentrated in the glomerular layer. These results suggest that phosphacan may play a role in delineating the pathway of growing olfactory axons as well as defining the laminar organization of the bulb. Together, the spatiotemporal expression patterns of neurocan and phosphacan indicate that these chondroitin sulfate proteoglycans have diverse in situ roles, which are dependent on context-specific interactions with extracellular and cell adhesion molecules within the developing olfactory nerve pathway. (C) 2000 Wiley-Liss, Inc.

Changes in intra-abdominal pressure during postural and respiratory activation of the human diaphragm

In humans, when the stability of the trunk is challenged in a controlled manner by repetitive movement of a limb, activity of the diaphragm becomes tonic but is also modulated at the frequency of limb movement. In addition, the tonic activity is modulated by respiration. This study investigated the mechanical output of these components of diaphragm activity. Recordings were made of costal diaphragm, abdominal, and erector spinae muscle electromyographic activity; intra-abdominal, intrathoracic, and transdiaphragmatic pressures; and motion of the rib cage, abdomen, and arm. During limb movement the diaphragm and transversus abdominis were tonically active with added phasic modulation at the frequencies of both respiration and limb movement. Activity of the other trunk muscles was not modulated by respiration. Intra-abdominal pressure was increased during the period of limb movement in proportion to the reactive forces from the movement. These results show that coactivation of the diaphragm and abdominal muscles causes a sustained increase in intra-abdominal pressure, whereas inspiration and expiration are controlled by opposing activity of the diaphragm and abdominal muscles to vary the shape of the pressurized abdominal cavity.

Sonification supports eyes-free respiratory monitoring and task time-sharing

Three experiments explored the effectiveness of continuous auditory displays, or sonifications, for conveying information about a simulated anesthetized patient's respiration. Experiment 1 established an effective respiratory sonification. Experiment 2 showed an effect of expertise in the use of respiratory sonification and revealed that some apparent differences in sonification effectiveness could be accounted for by response bias. Experiment 3 showed that sonification helps anesthesiologists to maintain high levels of awareness of the simulated patient's state while performing other tasks more effectively than when relying upon visual monitoring of the simulated patient state. Overall, sonification of patient physiology beyond traditional pulse oximetry appears to be a viable and useful adjunct to visual monitors. Actual and potential applications of this research include monitoring in a wide variety of busy critical care contexts.