Chemical Deception and Structural Adaptation in Microdon (Diptera, Syrphidae, Microdontinae), a Genus of Hoverflies Parasitic on Social Insects.

Various organisms, especially arthropods, are able to live as parasites in ant nests and to prey upon ant broods without eliciting any aggressive behaviour in the hosts. Understanding how these intruders are able to break the ants' communication codes in their favour represents a challenging and intriguing evolutionary question. We studied the chemical strategies of three European hoverfly species, Microdon mutabilis (parasitic on Formica cunicularia), M. analis (parasitic on Lasius emarginatus) and M. devius (parasitic on L. distinguendus). The peculiar slug-like larvae of these three species live inside ant nests feeding upon their broods. Gas chromatography-mass spectrometry analyses show that: 1) these parasites mimic the host brood rather than the ant workers, although each differs distinctly in the extent of chemical mimicry; 2) isolation experiments indicate that after 14 days the responsible cuticular hydrocarbons (CHCs) are not passively acquired but synthesized by the fly larvae. Additionally, Microdon larvae show an array of protective structural features, such as a thick and multi-layered cuticle, retractable head, dome-shaped tergum and a flat and strongly adhesive "foot" (sternum). This combination of protective chemical and structural features represents a successful key innovation by Microdon species, and one that may facilitate host switching. The results of a preliminary adoption analysis confirm that Microdon larvae of at least some species can readily be accepted by different species of ants.

Genetic diversity of Mycobacterium avium complex strains isolated in Argentina by MIRU-VNTR.

Mycobacterium avium sp. avium (MAA), M. avium sp. hominissuis (MAH), and M. avium sp. paratuberculosis (MAP) are the main members of the M. avium complex (MAC) causing diseases in several hosts. The aim of this study was to describe the genetic diversity of MAC isolated from different hosts. Twenty-six MAH and 61 MAP isolates were recovered from humans and cattle, respectively. GenoType CM® and IS1311-PCR were used to identify Mycobacterium species. The IS901-PCR was used to differentiate between MAH and MAA, while IS900-PCR was used to identify MAP. Genotyping was performed using a mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) scheme (loci: 292, X3, 25, 47, 3, 7, 10, 32) and patterns (INMV) were assigned according to the MAC-INMV database (http://mac-inmv.tours.inra.fr/). Twenty-two (22/26, 84·6%) MAH isolates were genotyped and 16 were grouped into the following, INMV 92, INMV 121, INMV 97, INMV 103, INMV 50, and INMV 40. The loci X3 and 25 showed the largest diversity (D: 0·5844), and the global discriminatory index (Hunter and Gaston discriminatory index, HGDI) was 0·9300. MAP (100%) isolates were grouped into INMV 1, INMV 2, INMV 11, INMV 8, and INMV 5. The HGDI was 0·6984 and loci 292 and 7 had the largest D (0·6980 and 0·5050). MAH presented a higher D when compared with MAP. The MIRU-VNTR was a useful tool to describe the genetic diversity of both MAH and MAP as well as to identify six new MAH patterns that were conveniently reported to the MAC-INMV database. It was also demonstrated that, in the geographical region studied, human MAC cases were produced by MAH as there was no MAA found among the human clinical samples.

Unexpected silicon localization in calcium carbonate exoskeleton of cultured and fossil coccolithophores.

Coccolithophores, marine calcifying phytoplankton, are important primary producers impacting the global carbon cycle at different timescales. Their biomineral structures, the calcite containing coccoliths, are among the most elaborate hard parts of any organism. Understanding the morphogenesis of coccoliths is not only relevant in the context of coccolithophore eco-physiology but will also inform biomineralization and crystal design research more generally. The recent discovery of a silicon (Si) requirement for crystal shaping in some coccolithophores has opened up a new avenue of biomineralization research. In order to develop a mechanistic understanding of the role of Si, the presence and localization of this chemical element in coccoliths needs to be known. Here, we document for the first time the uneven Si distribution in Helicosphaera carteri coccoliths through three synchrotron-based techniques employing X-ray Fluorescence and Infrared Spectromicroscopy. The enrichment of Si in specific areas of the coccoliths point to a targeted role of this element in the coccolith formation. Our findings mark a key step in biomineralization research because it opens the door for a detailed mechanistic understanding of the role Si plays in shaping coccolith crystals.

Static magnetic field effect on the Fremy's salt-ascorbic acid chemical reaction studied by continuous-wave electron paramagnetic resonance.

Static magnetic field effect in the framework of the radial pair mechanism (RPM) theory was studied on the biologically significant chemical reaction between ascorbic acid and Fremy's salt. The data indicate that the reaction rate depends on the applied magnetic field strength. The time scale of the studied reaction and the improved continuous-wave electron paramagnetic resonance system allowed for the first time the direct comparison of the amplitude differences between exposed and control samples in the strictly same boundary conditions. Until now the RPM was studied in a different time scale, focusing only on faster reactions by time-resolved techniques or by spectrophotometer measurement. The magnetic field effects presently measured can not be extended tout court to living systems; however the understanding of magnetic field sensitivity in basic chemical reaction in vitro could help clarifying the underlying basic step of interaction between magnetic fields and biological systems.

Assessment of biodegradation of the anionic surfactant sodium lauryl ether sulphate used in two foaming agents for mechanized tunnelling excavation.

The anionic surfactant sodium lauryl ether sulphate (SLES) is the main component in most foaming agents used for mechanized tunneling excavation. The process produces huge amounts of soil debris that can have a potential impact on ecosystems. The lack of accurate information about SLES persistence in excavated soil has aroused increasing concern about how it is recycled. The objective of this study was to assess SLES biodegradability in two commercial foaming agents (P1 and P2). Microcosm experiments were performed with two different soils collected from a tunnel construction site and conditioned with P1 or P2 (85.0 or 83.0 mg kg -1 of SLES, respectively). At selected times soil samples were collected for assessing the SLES residual concentration using Pressured Liquid Extraction followed by methylene blue active substance analysis (MBAS). Simultaneously, soil microbial abundance (DAPI counts), viability (Live/Dead method), activity (dehydrogenase analysis) and phylogenetic structure (Fluorescent In Situ Hybridization) were evaluated. SLES halved faster in the silty-clay soil (6 d) than in the gravel in a clay-silty-sand matrix (8-9 days). At day 28 it was degraded in both soils. Its biodegradation was ascribed to the significant increase in Gamma-Proteobacteria. At this time, the spoil material can be considered as a by-product.

EPO-releasing neural precursor cells promote axonal regeneration and recovery of function in spinal cord traumatic injury.

BACKGROUND: Spinal cord injury (SCI) is a debilitating condition characterized by a complex of neurological dysfunctions ranging from loss of sensation to partial or complete limb paralysis. Recently, we reported that intravenous administration of neural precursors physiologically releasing erythropoietin (namely Er-NPCs) enhances functional recovery in animals following contusive spinal cord injury through the counteraction of secondary degeneration. Er-NPCs reached and accumulated at the lesion edges, where they survived throughout the prolonged period of observation and differentiated mostly into cholinergic neuron-like cells. OBJECTIVE: The aim of this study was to investigate the potential reparative and regenerative properties of Er-NPCs in a mouse experimental model of traumatic spinal cord injury. METHODS AND RESULTS: We report that Er-NPCs favoured the preservation of axonal myelin and strongly promoted the regrowth across the lesion site of monoaminergic and chatecolaminergic fibers that reached the distal portions of the injured cord. The use of an anterograde tracer transported by the regenerating axons allowed us to assess the extent of such a process. We show that axonal fluoro-ruby labelling was practically absent in saline-treated mice, while it resulted very significant in Er-NPCs transplanted animals. CONCLUSION: Our study shows that Er-NPCs promoted recovery of function after spinal cord injury, and that this is accompanied by preservation of myelination and strong re-innervation of the distal cord. Thus, regenerated axons may have contributed to the enhanced recovery of function after SCI.

Size and stability of dipalmitoylphosphatidylcholine/cholesterol unilamellar vesicles are affected by interaction with proteins.

The effect of entrapping the enzyme ascorbate oxidase into dipalmitoylphosphatidylcholine/cholesterol vesicles, was studied by conventional transmission electron microscopy and freeze-fracture. The freeze-fracture technique has definitely demonstrated the unilamellar nature of empty and enzyme-loaded vesicles. Images of freeze-fractured and label-fractured liposomes also indicate that the observed reduction of vesicles volume could be related to the localization of ascorbate oxidase across the membrane. The membrane localization of ascorbate oxidase may explain the oxidation of externally added ascorbate by intact enzyme-loaded liposomes. Finally, the ageing of liposomes appears to be accelerated in the presence of proteins.

Room temperature electron spin resonance of superoxide dismutase-loaded liposomes and erythrocytes. A direct approach to the interaction of O2- with cells.

Human erythrocytes were enriched with bovine superoxide dismutase by fusion with liposomes containing the entrapped enzyme. Liquid solution ESR of intact cells at room temperature was used to measure directly the increase in the superoxide dismutase content. From the spectral characteristics (g-value and hyperfine splitting tensor), the structural integrity of the Cu site of the enzyme was found to be unaffected by the liposome preparation procedure or the incubation with cells. Changes in the ESR signal size were used to test directly the interaction of superoxide with the enzyme entrapped in liposomes or delivered to erythrocytes. It was found that the liposome-entrapped enzyme does not react with externally generated O2-, but once delivered to red blood cells this reaction can take place. This is the first demonstration of O2- -scavenging activity by superoxide dismutase delivered into an intact cell structure and is therefore to be considered as strong evidence for activity of this enzyme under in vivo conditions.

Binding and iron delivering of ovotransferrin to cholesterol-depleted chick-embryo red blood cells.

Binding and iron delivering of ovotransferrin (OTf) were evaluated using 14-day old chick-embryo red blood cells (CERBC) and cholesterol-depleted by treatment with chicken egg phosphatidyl choline (E-PC) liposomes. Liposome-treated CERBC assayed for their cholesterol content showed a cholesterol depletion depending on the incubation time, being 25% (w/w) of the maximum cellular removal of cholesterol seen after 22 h incubation at 37 degrees C. Total phosphorus content did not change either for the various samples or during the different incubation times, indicating that specific cholesterol removal occurred, as confirmed also by the increased membrane fluidity revealed through fluorescence anisotropy measurements. The apparent dissociation constant (Kd) of control and treated CERBC was almost of the same value at the same incubation time, ranging from 0.30 microM after 0.25 h incubation to 0.19 microM after 14 or 22 h incubation. In all experiments, the maximum value of bound OTf molecules per cell (Bmax) notably decreased as incubation time increased. But, in cholesterol partly depleted CERBC, the decrease of the Bmax values was less pronounced as the incubation time increased. As far as binding experiments were concerned, iron uptake studies showed that uptaking capacities decreased as incubation time increased. Considering both binding and iron uptake, at the same incubation time, liposome-treated CERBC were slightly more efficient with respect to untreated samples. In any case a passive iron delivering could be evidenced after 22 h incubation. It is suggested that cholesterol may tune binding and iron uptake by either regulating or affecting the expression or mobility of the OTf receptor.

The binding of human serum transferrin to its specific receptor reconstituted into liposomes.

Human placental transferrin receptor (HPTR), purified following a procedure based on affinity chromatography step, was reconstituted by the detergent dialysis method into various kinds of phosphatidylcholine vesicles and the receptor ability to bind 125I-labelled human serum transferrin (HST) was then evaluated. In our experimental conditions, the binding of the labelled protein to its specific receptor showed several features, in particular: (1) in cholesterol/1-alpha-dipalmitoylphosphatidyl choline (CHO/DPPC) liposomes, a positive cooperatively of the transferrin binding resulted at the lowest cholesterol/phospholipids (C/P) ratio; 1-alpha-dioleylphosphatidyl choline (DOPC) and phosphatidic acid (PA) containing liposomes showed an opposite binding curve trend; (2) the apparent dissociation constant (K'd) did not change significantly as a function of the lipid composition, being always around 1.00 x 10(-6) M; (3) the encapsulation capacity of liposomes decreased from 27% to about 13% with increasing amounts of cholesterol and was around 20% in the presence of DOPC or PA; about 8-13% of this receptor was found to be functional; (4) receptor-loaded liposomes treated with polyclonal anti-HPTR rabbit antibodies showed a remarkable binding decrease for transferin. All these results seem to point out the crucial role played by the environment in the binding behaviour of the transferrin receptor.

Regional differences in signalling transduction pathways among smooth muscle cells from rabbit colon.

Smooth muscle cells (SMC) from the circular muscle layer of rabbit colon, taken from the proximal and distal regions that are known to have different physiological and motor activities, were used to highlight distinct regional intrinsic myogenic properties and to investigate the correlations between receptor and signalling transduction pathways. Contractile agonists were shown to be more potent on proximal than on distal SMC in inducing contraction and intracellular Ca(2+) increase. Concentration-response curves of agonists-induced Ca(2+) increase were constantly shifted to the right, though remaining parallel, with respect to contraction curves, independently of the region analysed. Using agents activating different steps of cAMP-or cGMP-mediated intracellular cascades, main regional differences were revealed as far as relaxation was concerned. Relaxation of proximal SMC was found to be essentially cGMP mediated, while that of distal SMC was cAMP mediated. In conclusion, the motor patterns of the two regions appear to be influenced by distinct regional biochemical characteristics that are intrinsic to colonic SMC.

Amelioration of neutrophil membrane function underlies granulocyte-colony stimulating factor action in glycogen storage disease 1b.

Glycogen storage disease (GSD) 1b is a metabolic disorder characterized by a deficiency of glucose 6-phosphate transporter and neutrophil alterations, which are reduced in number and functionally impaired. The present study aimed at investigating neutrophil dysfunction correlating submembrane and cytoskeletal changes at different ages with or without granulocyte-colony stimulating factor (G-CSF) treatment. GSD1b neutrophils showed reduced expression and diffused localization of focal adhesion kinase (FAK) and actin. No abnormalities were observed in GSD1a patient neutrophils. Gelsolin was also slightly reduced in neutrophils of GSD1b patients. When patients were treated for at least 3 months with G-CSF, the neutrophil number and the expression of FAK and actin were significantly increased. Granulocyte colony-stimulating factor treatment was similarly effective when performed in 1 year old patients. FAK auto- and IL-8-mediated phosphorylations were already affected as early as 1 year of age. G-CSF treatment also improved this alteration. Our data suggest that neutrophil dysfunction in GSD1b patients might be related to functional impairment and disorganization of proteins of the sub-membrane apparatus, and that G-CSF treatment counteracts neutropenia and prevents the progressive alterations of neutrophil sub-membrane proteins.

Opiates act centrally on GH and PRL release.

This study was conducted in order to further understand whether the effect of opiates on growth hormone and prolactin release was exerted centrally or at least in part peripherally. Male rats were treated with morphine-HCl after pretreatment with saline with either the opiate antagonist naloxone-HCl or the quaternary derivative naloxone methyl bromide (Naloxone-Br), the latter of which does not cross the blood brain barrier. Morphine-HCl elicited a clear cut increase in prolactin and growth hormone release after pretreatment with naloxone-Br, but not after pretreatment with naloxone-HCl. Naloxone-Br, however, was able to inhibit the effect of morphine when administered directly in the brain ventricles. To further confirm these results, we administered the quaternary derivative morphine-methyl-iodide (morphine-I), which unlike morphine-HCl, does not cross the blood brain barrier. Morphine-I was ineffective in eliciting growth hormone and prolactin release when administered peripherally, but was effective when administered intraventricularly.

Plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations during pregnancy in normal and drug-addicted women and their newborn.

Most studies of plasma beta-endorphin concentrations in pregnant women show that these are highly elevated. This might indicate a role for opiate peptides during pregnancy and in the fetus-mother relationship. We measured plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations in normal and drug-addicted women during pregnancy, labor, and delivery, and in their newborn infants. Peptides were measured by RIA after extraction and concentration on silica columns and separation by high pressure liquid chromatography. In both normal and drug-addicted mothers we found an increase in plasma beta-endorphin during pregnancy, without a concomitant increase in plasma beta-lipotropin or metenkephalin. Only beta-lipotropin increased dramatically in both groups at delivery, whereas beta-endorphin and met-enkephalin remained unchanged. Peptide concentrations in umbilical plasma were similar to those in peripheral plasma of the mothers. On day 1 of life plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations in the newborn from normal mothers were higher than in nonpregnant adult subjects and gradually decreased toward normal adult values by day 5 of life. Plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations of newborn infants of drug-addicted mothers increased dramatically on day 2 and 3 of life, up to 1000-fold the concentrations of normal adults, and remained elevated up to 40 days after birth. In conclusion, beta-endorphin, beta-lipotropin, and met-enkephalin concentrations during pregnancy are not affected by drug addiction, whereas in the newborn of drug addicted mothers concentrations of these compounds are markedly increased.

Methionine-enkephalin, substance P, and homovanillic acid in the CSF of parkinsonian patients.

Methionine-enkephalin, substance P, and homovanillic acid concentrations were measured in the CSF of subjects not affected by neurologic disorders (group 1), and in parkinsonian patients who had a slight or moderate (group 2) or severe (group 3) disability. Homovanillic acid and substance P concentrations in the CSF of groups 2 and 3 were respectively lower and higher than in group 1. On the contrary, an increase in CSF methionine-enkephalin content was found only in group 2. Our results confirm in humans the close relation between the dopaminergic and peptidergic transmissions in the nigrostriatal system that has been observed in experimental animals.

Glycosaminoglycans boost insulin-like growth factor-I-promoted neuroprotection: blockade of motor neuron death in the wobbler mouse.

Wobbler mice display forelimb weakness, altered paw positioning, reduced running speed, muscle atrophy and motor neuron loss; co-treatment with glycosaminoglycans and insulin-like growth factor-I counteracts the progression of the disease. Reportedly, treatment with glycosaminoglycans or insulin-like growth factor-I slows the early stages of progressive forelimb dysfunction in wobbler mice. Our aim was to study whether the combination of these two drugs would result in greater neuroprotective effects. In a group of wobbler mice, combined treatment with daily s.c. administration of 20 microg/kg insulin-like growth factor-I and 1 mg/kg glycosaminoglycans was begun upon diagnosis at three weeks of age and continued for the next six weeks. This treatment halted motor neuron loss and markedly reduced the decay of forelimb muscle morphometry and function. Moreover, the mouse phenotype itself was strikingly improved. The effect of the combination treatment was significantly higher than that of the single drugs, even at a dosage as high as 1 mg/kg insulin-like growth factor-I. The ability of the insulin-like growth factor-I/glycosaminoglycans pharmacological cocktail to arrest the progression of motor neuron disease in wobbler mice and the safety of the low dose of insulin-like growth factor-I used hold promise that this combination might represent a novel approach for the treatment of motor neuron disease and peripheral neuropathies.

Fate of autologous dermal stem cells transplanted into the spinal cord after traumatic injury (TSCI).

Rat dermis is a source of cells capable of growing in vitro and, in appropriate conditions, forming floating spheres constituted by nestin-positive cells. We have clonally grown these spheres up to the 15th generation. These spheres can be dissociated into cells that differentiate in vitro under appropriate conditions, these cells are labeled by antibodies to immature neuron markers such as nestin and beta-tubulin III and, later, to mature neuron markers such as microtubule-associated protein 2 and neurofilaments. However, most cells are positive to the astroglial marker glia fibrillary acidic protein (GFAP). When sphere-derived cells are transplanted into the spinal cord after traumatic injury, their migration into the lesion cavity is optimal but their differentiation is dependent upon the time interval between lesioning and cell transplantation. Injection of skin-derived stem cell within 30 min from injury yields mainly membrane activated complex-1 (MAC-1), cluster of differentiation-4 (CD-4) and CD-8 positive cells, that 60-90 days later undergo apoptosis. However, when transplantation is performed 7 days after injury, most cells (65% of total) are positive to staining with antibodies to GFAP, others (16%) to neurofilaments, and a smaller amount (2%) to the endothelial marker, platelet endothelial cell adhesion molecule. Thus our study shows that delayed transplantations of dermis-derived stem cells yield healthy cells that do not die, migrate to the lesion site, and there differentiate mainly in cells expressing glia and neuronal markers. On the other hand there is the possibility of dye transfer from labeled cells to endogenous cells, and this might influence the data.

Muscle reinnervation following neonatal nerve crush. Interactive effects of glycosaminoglycans and insulin-like growth factor-I.

This study shows that glycosaminoglycans promote muscle reinnervation following neonatal sciatic nerve injury. Such an effect appears to be mediated by insulin-like growth factor-1. The glycosaminoglycan moiety of proteoglycans is a constituent of the basal lamina active on nerve regeneration by means of the interaction with laminin and with several growth factors. We have previously shown that supplementation of glycosaminoglycans affects neuronal degeneration and regeneration. In this study we report that following neonatal lesion of the rat sciatic nerve glycosaminoglycan treatment promoted extensor digitorum longus muscle reinnervation with consequent improvement of muscle morphology. In saline-treated rats, reinnervation was only partial and there was a marked muscle fibre atrophy. In addition glycosaminoglycan treatment of lesioned rats increased insulin-like growth factor-I messenger RNA and protein in the reinnervated muscle, and insulin-like growth factor-I and insulin-like growth factor binding protein-3 plasma levels. Similarly, treatment of nerve lesioned rats with insulin-like growth factor-I promoted muscle reinnervation and prevention of muscle fibre atrophy, higher levels of insulin-like growth factor-I in the reinnervated muscle and of insulin-like growth factor-I and insulin-like growth factor binding proteins in plasma. These data suggest that glycosaminoglycans are potent stimulants of muscle reinnervation and that their effects may be mediated by increased levels of insulin-like growth factor-I.

On the distribution of [met5]- and [leu5]-enkephalins in the brain of the rat, guinea-pig and calf.

1 By means of a highly sensitive radioimmunoassay, the content of [met5]-enkephalin (ME) and [leu5]-enkephalin (LE) was measured in various regions of the rat, guinea-pig and calf brain. Provisions were made to differentiate ME from LE by the use of cyanogen bromide (CNBr) to destroy methionine, the carboxy terminal amino acid in the ME sequence. This allows for correction of possible errors due to the cross-reactivity of the ME to the LE antiserum. Evidence is presented to demonstrate that the specific radioimmunoassay combined with the CNBr technique is a valid method to measure LE and ME content in crude tissue extracts. 2 In all the species studied, enkephalins appeared to be highly concentrated in the striatum and hypothalamus while very low amounts were found in the cerebe-lum and hippocampus. 3 Although the ratio between ME and LE content varied from area to area, the ME content in every region of the rat, guinea-pig and calf brain was more than 4 fold greater than that of LE.

Structural and biochemical alterations in the dorsal horn of the spinal cord caused by peripheral nerve lesions.

Substance P and Met-enkephalin were detected by radioimmunoassay and immunocytochemistry in the rat lumbar spinal cord. The sciatic nerve was lesioned by resecting a piece and the proximal stump was either ligated, for limiting neurite outgrowth, or intraperitoneally sutured, allowing the formation of a large neuroma. Ten days post lesioning both peptide levels dropped approximately 50% and the punctate immunoreactivity decreased in the dorsal horn. Lesioning both sciatic nerves did not accelerate nor increase the extent of peptide loss compared to monolateral lesions. Immunocytochemistry showed that after bilateral lesioning also the punctate immunoreactivity in the dorsal horn decreased less drastically. However, FRAP staining of the dorsal horn decreased according to the lesion paradigms, mono- and bilaterally with the same intensity. Therefore nerve lesions trigger the process, but the peptidergic loss seems intraspinally regulated. In addition, both kinds of abnormal neurite outgrowth similarly altered peptide levels and distribution in the spinal cord. Our data suggest that pain states related to peripheral nerve lesions may be due to opiate peptide loss rather than to neuroma.