Serum cytokine levels in neonates with congenital diaphragmatic hernia.

Cytokines play an important role in immune regulation and fetal lung development. The systemic inflammatory response in newborns with congenital diaphragmatic hernia (CDH) has not been characterized so far. We compared various concentrations of cytokines in serum from newborns with CDH and in healthy term neonates. We analyzed cytokine patterns of CDH newborns under extracorporeal membrane oxygenation (ECMO) and mechanical ventilation (MV).38 newborns with CDH were included: ECMO group (n=13) and non-ECMO group (n=25). Healthy term neonates served as controls (n=13). Serum samples were obtained prospectively after birth and during therapy.Concentrations of IFN-α, IL-3,-6,-7,-8,-10, MIP-1α,-1ß and TNF-α in serum of newborns with CDH were higher than in umbilical cord blood of term neonates. Infants with severe CDH requiring ECMO therapy had higher postnatal IL-8,-10, and MIP-1α levels than newborns with milder disease in the non-ECMO treated group. IL-10 progressively decreased during the first 3 days following birth under ECMO. In contrast, the chemokine MIP-1α remained elevated during ECMO therapy compared to mechanically ventilated CDH newborns.The pattern of cytokines in the serum of newborns with CDH showed significant elevations compared to term neonates. Our findings indicate that CDH is associated with systemic inflammatory response immediately after birth. ECMO and MV show a similar increase of IL-1α and IP-10 in CDH newborns assuming a persistent pulmonary inflammatory reaction irrespective of the conducted treatment.

Implication of pulse oxymetry screening for detection of congenital heart defects.

BACKGROUND: In newborns congenital heart defects can take an asymptomatic course, causing a diagnostic gap in the routine examination. Therefore pulsoxymetric screening is under discussion, as it could close this diagnostic gap. PATIENTS AND METHODS: Non-invasive postductal peripheral oxygen saturation assessment was carried out in 3 364 term neonates, 6-36 h of age, in 2008. In asymptomatic neonates with values > or = 95%, no further steps were applied. In those with values between 90% and 94% and no clinical abnormalities, a check-up was carried out 4-6 h later. Echocardiography was performed when the initial value was below 90% or persisted < 95 %. RESULTS: A total of 18 (0.5%) abnormal pulse oximetry values requiring echocardiographic investigation were found in the 3 364 neonates examined. 9 congenital heart defects that had not been recognized prenatally were diagnosed. 4 of these children were also found to have anomalies at the clinical examination. Persistent fetal circulation was noted in 2 of the neonates.In addition neonatal infections has been detected in 7 newborns. 1 neonate with stenosis of the aortic isthmus and 1 with pulmonary stenosis were missed in the screening program, with pulse oximetry saturation levels >95%. These data represent a sensitivity of 82% and a specificity of 99.9%, with a positive predictive value of 50% and a negative predictive value of 99.9%. CONCLUSIONS: Together with the clinical examination, pulse oximetry in neonates is a screening method that has high levels of sensitivity and specificity for early diagnosis of congenital heart defects. The risk-benefit profile may favour pulse oximetry to be standardized and universally used.

Individual versus social pathway to honeybee worker reproduction (Apis mellifera): pollen or jelly as protein source for oogenesis?

Honeybee workers, Apis mellifera, can reproduce in queenless colonies. The production of queen-like pheromones may be associated with their reproductive activity and induce nestmates to respond by feeding them. Such frequent trophallaxis could supply their protein needs for oogenesis, constituting a social pathway to worker reproduction. However, some individuals can develop ovaries without producing queen pheromones. The consumption of protein-rich pollen could be an alternative solitary pathway for them to satisfy this dietary requirement. In order to investigate the way in which workers obtain proteins for oogenesis, we created orphaned worker groups and determined ovarian and pheromonal development in relation to pollen consumption of selected workers. Individuals that did not consume pollen had significantly more developed ovaries and produced significantly more queen mandibular pheromone than workers that fed directly on pollen. Our results suggest that workers producing queen-like secretions are fed trophallactically. However, reproductive workers that lacked queen pheromones had consumed little or no pollen, suggesting that they also obtained trophallaxis. Although pollen consumption might contribute to sustaining oogenesis, it does not appear to be sufficient. Trophallaxis as a means of obtaining proteins seems to be necessary to attain reproductive status in queenless honeybee colonies.

[New guidelines for the assessment of bioavailability and bioequivalence]. / Neue Richtlinien für die Beurteilung der Bioverfügbarkeit/Bioäquivalenz.

Bioavailability and bioequivalence studies are essential in the clinical development of medicinal products and the optimization of pharmaceutical forms. Bioavailability means the rate and extent to which the active substance or active moiety is absorbed from a pharmaceutical form and becomes available at the site of action. In practice, drug concentration-time courses are measured in the systemic circulation, and the area under the curve (AUC) as well as the observed maximum concentration (C(max)) are determined. Products are considered bioequivalent if their bioavailabilities after administration of the same molar doses are similar to such a degree that their effects, with respect to both efficacy and safety, will be essentially the same and thus, there are no relevant differences in terms of AUC and C(max). In 2002 a revised version of the 'Note for Guidance on the Investigation of Bioavailability and Bioequivalence' came into effect (CPMP/EWP/QWP/1401/98). Relevant changes in comparison to the previous version are: request for GLP-compliant bioanalytical measurements; for long half-life drugs a truncated AUC is acceptable; acceptance criteria for bioequivalence assessment and requirements for a waiver of bioequivalence studies were further specified. In this context the Biopharmaceutics Classification System (BCS) seems appropriate to decide whether in special cases of rapidly dissolving solid oral dosage forms a biowaiver may be granted or not. Products not considered critical in this matter are medicinal products for which the formulation does not affect the rate and extent of absorption, i. e. bioavailability, of the active moiety. Highly soluble (and highly permeable) drugs (BCS class I) are such candidates. Comprehensive state-of-the-art guidance on the design, conduct and analysis of bioavailability and bioequivalence studies is given in the current European guideline.

Honeybee combs: construction through a liquid equilibrium process?

Geometrical investigations of honeycombs and speculations on how honeybees measure and construct the hexagons and rhombi of their cells are centuries old. Here we show that honeybees neither have to measure nor construct the highly regular structures of a honeycomb, and that the observed pattern of combs can be parsimoniously explained by wax flowing in liquid equilibrium. The structure of the combs of honeybees results from wax as a thermoplastic building medium, which softens and hardens as a result of increasing and decreasing temperatures. It flows among an array of transient, close-packed cylinders which are actually the self-heated honeybees themselves. The three apparent rhomboids forming the base of each cell do not exist but arise as optical artefacts from looking through semi-transparent combs.

A comparison of the dance language in Apis mellifera carnica and Apis florea reveals striking similarities.

Honeybees have a dance language by which successful foragers inform nestmates about attractive food patches. The classical concept of dialects in the dance language of honeybees points to two differences in the dances by different species and races, firstly in the flight distance at which the dancers start performing waggle dances instead of round dances, and secondly in the circuit duration of the waggle dance performed for a given flight distance. However, recent findings have indicated that the dance language is influenced and affected by a number of parameters, both genetic and environmental. The current study was carried out to see whether the distance at which dancers change from round dances to waggle dances is statistically different in two different species, Apis mellifera carnica and A. florea and to develop a set of definitions for such comparative studies. Results show that the two species do not differ in the relative proportion of waggle dances and round dances performed at a given distance. Thus, this study points to the need of addressing the dialect question again.

Raman spectroscopic study of spatial distribution of propolis in comb of Apis mellifera carnica (Pollm.).

Micro-Raman spectroscopy and Raman mapping are applied to investigate the spatial distribution and chemical composition of wax and propolis in the comb of Apis mellifera carnica (Pollm). A thick layer of propolis at the rim of some cells is identified by Raman spectroscopy. Raman mapping is applied to resolve the distribution of propolis and wax on a micron scale. Both components are connected at the rim of the cell with a mixture of wax and propolis. A layer of almost pure propolis is found on top of the mixture. It appears that even in the mixture, where both components come into close contact, the propolis and the wax remain separated and keep their chemical identity.

Recruitment of honeybees to non-scented food sources.

Small groups of honeybees (five to nine individuals) were trained to forage at feeders 150 m, 300 m and 800 m from an observation hive. Their behaviour in the hive and at the feeder was recorded by observers that maintained continuous radio contact with one another. At low concentrations of sugar in the feeder (0.5 mol x l(-1)) foragers do not dance in the hives, their flights to the feeder are often undertaken alone, they land immediately after arrival at the site and no recruits from the hive landed on the feeder during 30 h of observation. Raising the concentration of sugar in the feeder to 2 mol x l(-1) leads to vigorous dancing by the foragers and the gradual (over 10-15 min) synchronisation of their flights so that they arrive in groups of up to five bees at the feeder and undertake circular "buzzing" flights before landing. Such behaviour of the foragers is associated with the appearance of recruits which were never seen to fly around the feeder and land alone or before the foragers. Recruits typically circle the feeder together with foragers and land with them or continue their circling flights to land about 10 s later. While circling the feeder recruits, but not foragers, will fly after a moving lure if the presentation of the lure is accompanied by the release of geraniol scent. We propose that recruits that have witnessed a waggle dance are unlikely to find a non-scented feeder unless the foragers continue their flights to that feeder and provide supplementary visual and/or olfactory cues, at least in the vicinity of the feeder. We propose that the synchronisation of the flights of foragers and their behaviour at the feeding site is a strategy designed to overcome a navigational gap in the recruiting process in which the dance can indicate the general area of a food source but not the precise position of a highly localised site.

Phase reversal of vibratory signals in honeycomb may assist dancing honeybees to attract their audience.

Forager honeybees dancing on the comb are able to attract dance-followers from distances across the comb that are too remote for tactile or visual signals to play a role. An alternative signal could be the vibrations of the comb at 200-300 Hz generated by dancing bees but which, without amplification, may not be large enough to alert remote dance-followers. We describe here, however, an unexpected property of honeycomb when it is subjected to vibration at around 200 Hz that would represent an effective amplification of the vibratory signals for remote dance-followers. We find that, at a specific distance from the origin of an imposed vibration, the walls across a single comb cell abruptly reverse the phase of their displacement and move in opposite directions to one another. Behavioural measurements show that the distance from which the majority of remote dance-followers are recruited coincides with the location of this phase-reversal phenomenon relative to the signal source. We propose that effective signal amplification by the phase-reversal phenomenon occurs when bees straddle a cell across which the phase reversal is expressed. Such a bee would be subjected to a situation in which the legs were moving towards and away from one another instead of in the same direction. In this manner, remote dance-followers could be alerted to a dancer performing in their vicinity.

Honeybee dances communicate distances measured by optic flow.

In honeybees, employed foragers recruit unemployed hive mates to food sources by dances from which a human observer can read the distance and direction of the food source. When foragers collect food in a short, narrow tunnel, they dance as if the food source were much farther away. Dancers gauge distance by retinal image flow on the way to their destination. Their visually driven odometer misreads distance because the close tunnel walls increase optic flow. We examined how hive mates interpret these dances. Here we show that recruited bees search outside in the direction of the tunnel at exaggerated distances and not inside the tunnel where the foragers come from. Thus, dances must convey information about the direction of the food source and the total amount of image motion en route to the food source, but they do not convey information about absolute distances. We also found that perceived distances on various outdoor routes from the same hive could be considerably different. Navigational errors are avoided as recruits and dancers tend to fly in the same direction. Reported racial differences in honeybee dances could have arisen merely from differences in the environments in which these bees flew.

Visual constraints in foraging bumblebees: flower size and color affect search time and flight behavior.

In optimal foraging theory, search time is a key variable defining the value of a prey type. But the sensory-perceptual processes that constrain the search for food have rarely been considered. Here we evaluate the flight behavior of bumblebees (Bombus terrestris) searching for artificial flowers of various sizes and colors. When flowers were large, search times correlated well with the color contrast of the targets with their green foliage-type background, as predicted by a model of color opponent coding using inputs from the bees' UV, blue, and green receptors. Targets that made poor color contrast with their backdrop, such as white, UV-reflecting ones, or red flowers, took longest to detect, even though brightness contrast with the background was pronounced. When searching for small targets, bees changed their strategy in several ways. They flew significantly slower and closer to the ground, so increasing the minimum detectable area subtended by an object on the ground. In addition, they used a different neuronal channel for flower detection. Instead of color contrast, they used only the green receptor signal for detection. We relate these findings to temporal and spatial limitations of different neuronal channels involved in stimulus detection and recognition. Thus, foraging speed may not be limited only by factors such as prey density, flight energetics, and scramble competition. Our results show that understanding the behavioral ecology of foraging can substantially gain from knowledge about mechanisms of visual information processing.

Worker piping in honey bee swarms and its role in preparing for liftoff.

Worker piping, previously reported only in hives, was observed in swarms as they prepared to liftoff to fly to a new home. Pipers are excited bees which scramble through the swarm cluster, pausing every second or so to emit a pipe. Each pipe consists of a sound pulse which lasts 0.82 +/- 0.43 s and rises in fundamental frequency from 100-200 Hz to 200-250 Hz. Many. if not all, of the pipers are nest-site scouts. The scouts pipe when it is time to stimulate the non-scouts to warm themselves to a flight-ready temperature (35 degrees C) in preparation for liftoff. The time-course of worker piping matches that of swarm warming, both start at a low level, about an hour before liftoff, and both build to a climax at liftoff. When we excluded pipers from bees hanging in the cool, outermost layer of a swarm cluster, we found that these bees did not warm up. The form of worker piping that we have studied in swarms differs from the form of worker piping that others have studied in hives. We call the two forms "wings-together piping" (in swarms) and "wings-apart piping" (in hives).

The role of leaf structure in vibration propagation.

The leaf and its structural components play a key role in the propagation of short transient signals produced by insects. In this paper, it is shown how the complex structure of an apple leaf could be modeled by a much simpler one for the analysis of vibratory signal propagation. Waves were produced by impacts of small spheres and the propagation studied using two laser vibrometers, followed by a wavelets analysis. Three components of the leaf were investigated: the midvein, minor veins, and the interspaced homogeneous regions making up the leaf lamina. The loss of signal energy over the leaf lamina and across minor veins and midvein was studied. For the midvein, the loss of energy decreased from 80% at the leaf base to 40% at the apex. For minor veins, the loss of energy decreased from 70% at the leaf base to 31% at the apex. The loss in homogeneous regions was 40%. A signal decomposition into two frequency ranges, above and below 1.7 kHz, showed that the midvein acted as a low-pass filter. As energy loss was mainly a function of vein diameter and not vein type, veins smaller or equal to 0.2 mm were considered as equivalent to homogeneous regions. Hence, a model leaf reduced to the leaf lamina and veins with a diameter >0.2 mm is retained for the study of signal propagation in a leaf.

Debris removal by head-pushing in A. florea Fabr. honeybees.

The nest of the dwarf honeybee A. florea Fabr. consists of a single comb attached to a tree branch. Recruitment dances take place on the upper surface of the comb that must therefore be kept clear of debris. We report here, for the first time, a behaviour that serves for removing leaves and other foreign objects from the surface of the comb. Individual workers crawl under the object and lift it with their heads, pushing it towards the rim where it eventually slides off the comb. Objects that are heavier or fixed at one end such as leaves are nevertheless lifted and kept away from the surface for up to several minutes. This "head-pushing" is frequently performed without the aid of mandibles, and individuals performing it maintain a distinctive posture, holding the forelegs at an angle without touching the object. Repeated involvement of particular individuals indicate that head-pushers might form a distinct task group.

Comb-wax discrimination by honeybees tested with the proboscis extension reflex.

We used the proboscis extension reflex of honeybees to test their ability to discriminate between comb waxes of different ages (wax scales, 1-week-old wax, 2- to 3-year-old wax, 8- to 10-year-old wax). Such waxes differ in their chemical composition, and an ability to discriminate between them may aid the orientation of the bees in the nest. To train the bees, we used whole extracts of waxes and four different fractions of the whole extract based on different elutions of solid-phase extractions (extract I, fraction A eluted with hexane and fraction B with diethylether; extract II, fraction B further subdivided into fraction C by elution with isopropylchloride and fraction D by elution with diethylether). In a differential training regime (six learning and six test trials) with whole extracts or with the different fractions, we paired one type of wax with a reward and another with no reward. The bees learned to discriminate between all tested pairs of whole extracts. The two subfractions (fractions A and B) gave different results: the bees could discriminate between waxes of different ages when fraction B was used but not when fraction A was used. A further subdivision of fraction B into fractions C and D showed that only fraction D contained the elements that enabled bees to discriminate between old and new wax. Fraction D makes up only 5?8 % of the total wax mass and contains hydroxy alkyl esters (5?6 % of the total wax mass), primary alcohols (0.3?0.5 % of the total wax mass) and acids (0.06?1. 0 % of the total wax mass). Fractions A and C (together forming 62?64 % of the total wax mass), which consist of unbranched and branched aliphatic hydrocarbons and alkyl esters, could not be discriminated by the bees. The remaining wax mass (25?29 %) was eluted with a mixture of chloroform, methanol and water (13:5:1) as fraction E.

Behaviour-locked signal analysis reveals weak 200-300 Hz comb vibrations during the honeybee waggle dance.

Waggle-dancing honeybees produce vibratory movements that may facilitate communication by indicating the location of the waggle dancer. However, an important component of these vibrations has never been previously detected in the comb. We developed a method of fine-scale behavioural analysis that allowed us to analyze separately comb vibrations near a honeybee waggle dancer during the waggle and return phases of her dance. We simultaneously recorded honeybee waggle dances using digital video and laser-Doppler vibrometry, and performed a behaviour-locked Fast Fourier Transform analysis on the substratum vibrations. This analysis revealed significantly higher-amplitude 200-300 Hz vibrations during the waggle phase than during the return phase (P=0.012). We found no significant differences in the flanking frequency regions between 100-200 Hz (P=0.227) and 300-400 Hz (P=0.065). We recorded peak waggle phase vibrations from 206 to 292 Hz (244+/-28 Hz; mean +/- s. d., N=11). The maximum measured signal - noise level was +12.4 dB during the waggle phase (mean +5.8+/-2.7 dB). The maximum vibrational velocity, calculated from a filtered signal, was 128 microm s(-)(1) peak-to-peak, corresponding to a displacement of 0.09 microm peak-to-peak at 223 Hz. On average, we measured a vibrational velocity of 79+/-28 microm s(-)(1) peak-to-peak from filtered signals. These signal amplitudes overlap with the detection threshold of the honeybee subgenual organ.

Honeybee navigation: nature and calibration of the "odometer".

There are two theories about how honeybees estimate the distance to food sources. One theory proposes that distance flown is estimated in terms of energy consumption. The other suggests that the cue is visual, and is derived from the extent to which the image of the world has moved on the eye during the trip. Here the two theories are tested by observing dances of bees that have flown through a short, narrow tunnel to collect a food reward. The results show that the honeybee's "odometer" is visually driven. They also provide a calibration of the dance and the odometer in visual terms.

Ultrastructure and physiology of the CO2 sensitive sensillum ampullaceum in the leaf-cutting ant Atta sexdens.

The sensilla ampullacea on the apical antennomere of the leaf-cutting ant Atta sexdens were investigated regarding both their responses to CO2 and their ultrastructure. By staining the sensillum during recording, we confirmed that the sensilla ampullacea are responsible for CO2 perception. We showed that the sensory neurons of the sensilla ampullacea are continuously active without adaptation during stimulation with CO2 (test duration: 1 h). This feature should enable ants to assess the absolute CO2 concentration inside their nests. Sensilla ampullacea have been found grouped mainly on the dorso-lateral side of the distal antennal segment. Scanning and transmission electron microscopic investigations revealed that the external pore opens into a chamber which connects to the ampulla via a cuticular duct. We propose protection against evaporation as a possible function of the duct. The ampulla houses a peg which is almost as long as the ampulla and shows cuticular ridges on the external wall. The ridges are separated by furrows with cuticular pores. The peg is innervated by only one sensory neuron with a large soma. Its outer dendritic segment is enveloped by a dendritic sheath up to the middle of the peg. From the middle to the tip numerous dendritic branches (up to 100) completely fill the distal half of the peg. This is the first report of a receptor cell with highly branched dendrites and which probably is tuned to CO2 exclusively.

Honeybee waggle dance: recruitment success depends on the dance floor

The waggle dance of the honeybee Apis mellifera, used to recruit nestmates to a food source, takes place on the surface of the combs in the dark hive. The mechanism of information transfer between dancer and follower bees is not entirely understood. The results presented here reveal a novel factor that must be brought into any consideration of this mechanism, namely that the nature of the floor on which the bees dance has a considerable influence on the recruitment of nestmates to a food source. Dancers on combs with open empty cells recruit three times as many nestmates to a food source as dancers on capped brood cells.