Rapid and Precise Semi-Automatic Axon Quantification in Human Peripheral Nerves

We developed a time-efficient semi-automated axon quantification method using freeware in human cranial nerve sections stained with paraphenylenediamine (PPD). It was used to analyze a total of 1238 facial and masseteric nerve biopsies. The technique was validated by comparing manual and semi-automated quantification of 129 (10.4%) randomly selected biopsies. The software-based method demonstrated a sensitivity of 94% and a specificity of 87%. Semi-automatic axon counting was significantly faster (p<0.001) than manual counting. It took 1hour and 47minutes for all 129 biopsies (averaging 50sec per biopsy, 0.04seconds per axon). The counting process is automatic and does not need to be supervised. Manual counting took 21hours and 6minutes in total (average 9minutes and 49seconds per biopsy, 0.52seconds per axon). Our method showed a linear correlation to the manual counts (R=0.944 Spearman rho). Attempts have been made by several research groups to automate axonal load quantification. These methods often require specific hard- and software and are therefore only accessible to a few specialized laboratories. Our semi-automated axon quantification is precise, reliable and time-sparing using publicly available software and should be useful for an effective axon quantification in various human peripheral nerves

On Scottish sawflies, with results of the 14th International Sawfly Workshop, in the southern Highlands, 2010 (Hymenoptera, Symphyta).

Die Ergebnisse der Feldarbeit aus dem 14. Internationalen Sawfly Workshop werden vorgestellt. Dies fand im südlichen schottischen Hochland, überwiegend in Perthshire, statt. Einige Nachweise von Pflanzenwespen aus anderen Teilen von Schottland sind enthalten. Erstnachweise für die Britischen Inseln werden für acht tenthredinide Arten erbracht: Amauronematus mimus, Phyllocolpa alienata, P. erythropyga, P. plicalapponum, P. plicaphylicifolia, P. prussica, Pristiphora thalictri und Tenthredo ignobilis. A. mimus ist auch aus Irland nachgewiesen. Amauronematus stenogaster ist aus der Liste der Symphyta der Britischen Inseln zu entfernen. Erstmalig sind in Schottland eine Cephiden-Art (Hartigia xanthostoma) und zehn Tenthrediniden gefunden worden (Claremontia uncta, Dolerus brevicornis, Empria basalis, Empria parvula, Parna apicalis, Pristiphora decipiens, Pristiphora leucopus, Pristiphora testacea, Tenthredo mandibularis und Tenthredopsis ornata). Merkmale für die Unterscheidung einzelner Arten werden dargestellt. Der Locus typicus von Nematus herbaceae Cameron, 1876 ist geklärt. Geranium pratense ist als Wirtspflanze von Macrophya albipuncta bestätigt. Salix reticulata ist eine Wirtspflanze von Amauronematus mcluckieae. Sedum rosea kann eine Wirtspflanze von Tenthredo ignobilis sein. Allgemeine Schlussfolgerungen: hoher Artenreichtum der Pflanzenwespenfauna von Perthshire im Vergleich zu anderen Regionen in Schottland, darunter mehrere Arten, die auf den Britischen Inseln nur von dort bekannt sind; die Wichtigkeit der Erhaltung der Lebensräume von Felssimsen; große Defizite in unseren Kenntnissen bei vielen Arten, insbesondere zu Wirtspflanzen, Phänologie und Verbreitung; hoher Wert der Malaise-Falle in faunistischen Untersuchungen von Symphyta, weil das Spektrum von erfassten Taxa sich unterscheidet von dem was mit Streifnetz erfasst wird; Zusammensetzung des 'fall-out’ auf Schneeresten deutet darauf hin, dass Pflanzenwespen in einem größeren Ausmaß als bisher angenommen, aktiver Dispersion innerhalb von Landmassen unterliegen.StichwörterBritish Isles, Cephidae, Pamphiliidae, Tenthredinoidea, hosts, phenology, distribution.Results of fieldwork are presented from the 14th International Sawfly Workshop, held in the southern Highlands of Scotland, mostly in Perthshire. Some records of Symphyta from other parts of Scotland are included. First records in the British Isles are given for eight tenthredinid species: Amauronematus mimus, Phyllocolpa alienata, P. erythropyga, P. plicalapponum, P. plicaphylicifolia, P. prussica, Pristiphora thalictri and Tenthredo ignobilis. A. mimus is also recorded from Ireland. Amauronematus stenogaster is removed from the list of British Isles Symphyta. Recorded for the first time in Scotland are one cephid (Hartigia xanthostoma) and ten tenthredinids (Claremontia uncta, Dolerus brevicornis, Empria basalis, Empria parvula, Parna apicalis, Pristiphora decipiens, Pristiphora leucopus, Pristiphora testacea, Tenthredo mandibularis and Tenthredopsis ornata). Characters useful for the distinction of some species are illustrated. The type locality of Nematus herbaceae Cameron, 1876 is clarified. Geranium pratense is confirmed as a host of Macrophya albipuncta. Salix reticulata is a host of Amauronematus mcluckieae [mandatory correction of spelling of species name]. Sedum rosea may be a host of Tenthredo ignobilis. General conclusions: high species richness of Perthshire sawfly fauna compared to other regions of Scotland, including several species only known in British Isles from there; importance to conservation of rock-ledge habitats; large data deficits for many species, particularly on hosts, phenology and distribution; high value of Malaise traps in faunal survey of Symphyta, because spectrum of taxa captured differs from that recorded by hand-netting; as indicated by species and sex composition of ‘fall-out’ on snow patches, adult sawflies undertake active dispersal, within landmasses, to a greater extent than is often assumed.KeywordsBritish Isles, Cephidae, Pamphiliidae, Tenthredinoidea, hosts, phenology, distribution

Tissue Viability of Free Flaps after Extracorporeal Perfusion Using a Modified Hydroxyethyl Starch Solution

Background: In free flap surgery, tissue is stored under hypothermic ischemia. Extracorporeal perfusion (EP) has the potential to extend storage time and the tissue's perspective of survival. In the present study, the aim is to improve a recently established, simplified extracorporeal perfusion system. Methods: Porcine musculus rectus abdominis were stored under different conditions. One group was perfused continuously with a simplified one-way perfusion system for six hours, while the other received only a single flush but no further treatment. A modified hydroxyethyl starch solution was used as a perfusion and flushing solution. Vitality, functionality, and metabolic activity of both groups were analyzed. Results: Perfused muscles, in contrast to the ischemically stored ones, showed no loss of vitality and significantly less functionality loss, confirming the superiority of storage under continuous perfusion over ischemic storage. Furthermore, in comparison to a previous study, the results were improved even further by using a modified hydroxyethyl starch solution. Conclusion: The use of EP has major benefits compared to the clinical standard static storage at room temperature. Continuous perfusion not only maintains the oxygen and nutrient supply but also removes toxic metabolites formed due to inadequate storage conditions

Morphological and Physiological Responses to Drought Stress of European Provenances of Scots Pine

Increased frequency and intensity of drought episodes as a consequence of current and predicted climatic changes require an understanding of the intra-specific variability in structural and physiological characteristics of forest trees. Adaptive plasticity and genotypic variability are considered two of the main processes by which trees can either be selected or can acclimate to changing conditions. We tested for the relative importance of genotypic variability, phenotypic plasticity and their interaction by comparing the growth and physiological performance of 15 provenances of Scots pine (Pinus sylvestris L.), under two contrasting irrigation regimes. Selected provenances representing the distribution range of the species in Anatolia, Turkey, were contrasted with seed sources spanning the range from Spain to the UK, in Europe. We found a strong latitudinal differentiation among the 15 provenances for survival after drought, largely the result of the higher mortality of some western and central European provenances. Differentiation in diameter and height growth was also clear with the worst provenance coming from Western Europe (UK). Among the Turkish provenances, the more extreme southern high-elevation populations showed greater survival and lower growth rates overall. Differences in growth and survival were related to differences in photosynthetic pigment and nutrient contents and in the photosynthetic efficiency of photosystem II. Plasticity was strongest for growth characters and pigment contents.WoSScopu

Host Shifts from Lamiales to Brassicaceae in the Sawfly Genus Athalia

Plant chemistry can be a key driver of host shifts in herbivores. Several species in the sawfly genus Athalia are important economic pests on Brassicaceae, whereas other Athalia species are specialized on Lamiales. These host plants have glucosides in common, which are sequestered by larvae. To disentangle the possible direction of host shifts in this genus, we examined the sequestration specificity and feeding deterrence of iridoid glucosides (IGs) and glucosinolates (GSs) in larvae of five species which either naturally sequester IGs from their hosts within the Plantaginaceae (Lamiales) or GSs from Brassicaceae, respectively. Furthermore, adults were tested for feeding stimulation by a neo-clerodane diterpenoid which occurs in Lamiales. Larvae of the Plantaginaceae-feeders did not sequester artificially administered p-hydroxybenzylGS and were more deterred by GSs than Brassicaceae-feeders were by IGs. In contrast, larvae of Brassicaceae-feeders were able to sequester artificially administered catalpol (IG), which points to an ancestral association with Lamiales. In line with this finding, adults of all tested species were stimulated by the neo-clerodane diterpenoid. Finally, in a phylogenetic tree inferred from genetic marker sequences of 21 Athalia species, the sister species of all remaining 20 Athalia species also turned out to be a Lamiales-feeder. Fundamental physiological pre-adaptations, such as the establishment of a glucoside transporter, and mechanisms to circumvent activation of glucosides by glucosidases are therefore necessary prerequisites for successful host shifts between Lamiales and Brassicaceae

The benzene metabolite para-benzoquinone is genotoxic in human, phorbol-12-acetate-13-myristate induced, peripheral blood mononuclear cells at low concentrations

Benzene is one of the most prominent occupational and environmental pollutants. The substance is a proven human carcinogen that induces hematologic malignancies in humans, probably at even low doses. Yet knowledge of the mechanisms leading to benzene-induced carcinogenesis is still incomplete. Benzene itself is not genotoxic. The generation of carcinogenic metabolites involves the production of oxidized intermediates such as catechol, hydroquinone and para-benzoquinone (p-BQ) in the liver. Further activation to the ultimate carcinogenic intermediates is most probably catalyzed by myeloperoxidase (MPO). Yet the products of the MPO pathway have not been identified. If an oxidized benzene metabolite such as p-BQ was actually the precursor for the ultimate carcinogenic benzene metabolite and further activation proceeds via MPO mediated reactions, it should be possible to activate p-BQ to a genotoxic compound in vitro. We tested this hypothesis with phorbol-12-acetate-13-myristate (PMA) activated peripheral blood cells exposed to p-BQ, using the cytokinesis-block micronucleus test. Addition of 20–28 ng/ml PMA caused a significant increase of micronuclei at low and non-cytotoxic p-BQ concentrations between 0.04 and 0.2 μg/ml (0.37–1.85 μM). Thus with PMA or p-BQ alone no reproducible elevation of micronuclei was seen up to toxic concentrations. PMA and p-BQ induce micronuclei when administered jointly. Our results add further support to the hypothesis that MPO is a key enzyme in the activation of benzene

Translational toxicology in setting occupational exposure limits for dusts and hazard classification – a critical evaluation of a recent approach to translate dust overload findings from rats to humans

Background We analyze the scientific basis and methodology used by the German MAK Commission in their recommendations for exposure limits and carcinogen classification of “granular biopersistent particles without known specific toxicity” (GBS). These recommendations are under review at the European Union level. We examine the scientific assumptions in an attempt to reproduce the results. MAK’s human equivalent concentrations (HECs) are based on a particle mass and on a volumetric model in which results from rat inhalation studies are translated to derive occupational exposure limits (OELs) and a carcinogen classification. Methods We followed the methods as proposed by the MAK Commission and Pauluhn 2011. We also examined key assumptions in the metrics, such as surface area of the human lung, deposition fractions of inhaled dusts, human clearance rates; and risk of lung cancer among workers, presumed to have some potential for lung overload, the physiological condition in rats associated with an increase in lung cancer risk. Results The MAK recommendations on exposure limits for GBS have numerous incorrect assumptions that adversely affect the final results. The procedures to derive the respirable occupational exposure limit (OEL) could not be reproduced, a finding raising considerable scientific uncertainty about the reliability of the recommendations. Moreover, the scientific basis of using the rat model is confounded by the fact that rats and humans show different cellular responses to inhaled particles as demonstrated by bronchoalveolar lavage (BAL) studies in both species. Conclusion Classifying all GBS as carcinogenic to humans based on rat inhalation studies in which lung overload leads to chronic inflammation and cancer is inappropriate. Studies of workers, who have been exposed to relevant levels of dust, have not indicated an increase in lung cancer risk. Using the methods proposed by the MAK, we were unable to reproduce the OEL for GBS recommended by the Commission, but identified substantial errors in the models. Considerable shortcomings in the use of lung surface area, clearance rates, deposition fractions; as well as using the mass and volumetric metrics as opposed to the particle surface area metric limit the scientific reliability of the proposed GBS OEL and carcinogen classification.International Carbon Black Associatio