An inter-laboratory validation of a real time PCR assay to measure host excretion of bacterial pathogens, particularly of Mycobacterium bovis

Advances in the diagnosis of Mycobacterium bovis infection in wildlife hosts may benefit the development of sustainable approaches to the management of bovine tuberculosis in cattle. In the present study, three laboratories from two different countries participated in a validation trial to evaluate the reliability and reproducibility of a real time PCR assay in the detection and quantification of M. bovis from environmental samples. The sample panels consisted of negative badger faeces spiked with a dilution series of M. bovis BCG Pasteur and of field samples of faeces from badgers of unknown infection status taken from badger latrines in areas with high and low incidence of bovine TB (bTB) in cattle. Samples were tested with a previously optimised methodology. The experimental design involved rigorous testing which highlighted a number of potential pitfalls in the analysis of environmental samples using real time PCR. Despite minor variation between operators and laboratories, the validation study demonstrated good concordance between the three laboratories: on the spiked panels, the test showed high levels of agreement in terms of positive/negative detection, with high specificity (100%) and high sensitivity (97%) at levels of 10(5) cells g(-1) and above. Quantitative analysis of the data revealed low variability in recovery of BCG cells between laboratories and operators. On the field samples, the test showed high reproducibility both in terms of positive/negative detection and in the number of cells detected, despite low numbers of samples identified as positive by any laboratory. Use of a parallel PCR inhibition control assay revealed negligible PCR-interfering chemicals co-extracted with the DNA. This is the first example of a multi-laboratory validation of a real time PCR assay for the detection of mycobacteria in environmental samples. Field studies are now required to determine how best to apply the assay for population-level bTB surveillance in wildlife.     

Effects of snout dimensions on the hydrodynamics of suction feeding in juvenile and adult seahorses

Seahorses give birth to juveniles having a fully functional feeding apparatus, and juvenile feeding behaviour shows striking similarities to that of adults. However, a significant allometric growth of the snout is observed during which the snout shape changes from relatively short and broad in juveniles to relatively long and slender in adults. Since the shape of the buccal cavity is a critical determinant of the suction performance, this snout allometry will inevitably affect the suction feeding ability. To test whether the snout is optimised for suction feeding throughout an ontogeny, we simulated the expansion of different snout shapes varying from extremely long and slender to short and broad for juvenile and adult snout sizes, using computational fluid dynamic models. Our results showed that the snout diameter at the start of the simulations is involved in a trade-off between the realizable suction volume and expansion time on the one hand (improving with larger initial diameters), and maximal flow velocity on the other hand (improving with smaller initial diameters). Moreover suction performance (suction volume as well as maximal attainable flow velocity) increased with decreasing snout length. However, an increase in snout length decreases the time to reach the prey by the cranial rotation, which may explain the prevalence of long snouts among syngnathid fishes despite the reduced suction performance. Thus, the design of the seahorse snout revolves around a trade-off between the ability to generate high-volume suction versus minimisation of the time needed to reach the prey by the cranial rotation.     

Chronic Obstructive Pulmonary Emphysema—Is Exercise Beneficial?

Following a six-week program of training in a series of exercises, a significant number of patients with chronic obstructive pulmonary emphysema showed decided improvement in functional activity. Subjective improvement also was noted and kept the patient motivation high.Preliminary observations indicated that the improvement could be maintained long after the end of the training period.     

Investigation of the diversity of effector genes in the banana pathogen,Fusarium oxysporum f. sp. cubense,reveals evidence of horizontal gene transfer

It is hypothesized that the virulence of phytopathogenic fungi is mediated through the secretion of small effector proteins that interfere with the defence responses of the host plant. In Fusarium oxysporum, one family of effectors, the Secreted In Xylem (SIX) genes, has been identified. We sought to characterize the diversity and evolution of the SIX genes in the banana‐infecting lineages of F. oxysporum f. sp. cubense (Foc). Whole‐genome sequencing data were generated for the 23 genetic lineages of Foc, which were subsequently queried for the 14 known SIX genes (SIX1–SIX14). The sequences of the identified SIX genes were confirmed in a larger collection of Foc isolates. Genealogies were generated for each of the SIX genes identified in Foc to further investigate the evolution of the SIX genes in Foc. Within Foc, variation of the SIX gene profile, including the presence of specific SIX homologues, correlated with the pathogenic race structure of Foc. Furthermore, the topologies of the SIX gene trees were discordant with the topology of an infraspecies phylogeny inferred from EF‐1α/RPB1/RPB2 (translation elongation factor‐1α/RNA polymerase II subunit I/RNA polymerase II subunit II). A series of topological constraint models provided strong evidence for the horizontal transmission of SIX genes in Foc. The horizontal inheritance of pathogenicity genes in Foc counters previous assumptions that convergent evolution has driven the polyphyletic phylogeny of Foc. This work has significant implications for the management of Foc, including the improvement of diagnostics and breeding programmes.     

Characterization and Molecular Profiling of PSEN1 Familial Alzheimer's Disease iPSC-Derived Neural Progenitors

Presenilin 1 (PSEN1) encodes the catalytic subunit of γ-secretase, and PSEN1 mutations are the most common cause of early onset familial Alzheimer''s disease (FAD). In order to elucidate pathways downstream of PSEN1, we characterized neural progenitor cells (NPCs) derived from FAD mutant PSEN1 subjects. Thus, we generated induced pluripotent stem cells (iPSCs) from affected and unaffected individuals from two families carrying PSEN1 mutations. PSEN1 mutant fibroblasts, and NPCs produced greater ratios of Aβ42 to Aβ40 relative to their control counterparts, with the elevated ratio even more apparent in PSEN1 NPCs than in fibroblasts. Molecular profiling identified 14 genes differentially-regulated in PSEN1 NPCs relative to control NPCs. Five of these targets showed differential expression in late onset AD/Intermediate AD pathology brains. Therefore, in our PSEN1 iPSC model, we have reconstituted an essential feature in the molecular pathogenesis of FAD, increased generation of Aβ42/40, and have characterized novel expression changes.     

The Trajectory of Dispersal Research in Conservation Biology. Systematic Review

Dispersal knowledge is essential for conservation management, and demand is growing. But are we accumulating dispersal knowledge at a pace that can meet the demand? To answer this question we tested for changes in dispersal data collection and use over time. Our systematic review of 655 conservation-related publications compared five topics: climate change, habitat restoration, population viability analysis, land planning (systematic conservation planning) and invasive species. We analysed temporal changes in the: (i) questions asked by dispersal-related research; (ii) methods used to study dispersal; (iii) the quality of dispersal data; (iv) extent that dispersal knowledge is lacking, and; (v) likely consequences of limited dispersal knowledge. Research questions have changed little over time; the same problems examined in the 1990s are still being addressed. The most common methods used to study dispersal were occupancy data, expert opinion and modelling, which often provided indirect, low quality information about dispersal. Although use of genetics for estimating dispersal has increased, new ecological and genetic methods for measuring dispersal are not yet widely adopted. Almost half of the papers identified knowledge gaps related to dispersal. Limited dispersal knowledge often made it impossible to discover ecological processes or compromised conservation outcomes. The quality of dispersal data used in climate change research has increased since the 1990s. In comparison, restoration ecology inadequately addresses large-scale process, whilst the gap between knowledge accumulation and growth in applications may be increasing in land planning. To overcome apparent stagnation in collection and use of dispersal knowledge, researchers need to: (i) improve the quality of available data using new approaches; (ii) understand the complementarities of different methods and; (iii) define the value of different kinds of dispersal information for supporting management decisions. Ambitious, multi-disciplinary research programs studying many species are critical for advancing dispersal research.     

New Insights into Muscle Function during Pivot Feeding in Seahorses

Seahorses, pipefish and their syngnathiform relatives are considered unique amongst fishes in using elastic recoil of post-cranial tendons to pivot the head extremely quickly towards small crustacean prey. It is known that pipefish activate the epaxial muscles for a considerable time before striking, at which rotations of the head and the hyoid are temporarily prevented to allow energy storage in the epaxial tendons. Here, we studied the motor control of this system in seahorses using electromyographic recordings of the epaxial muscles and the sternohyoideus-hypaxial muscles with simultaneous high-speed video recordings of prey capture. In addition we present the results from a stimulation experiment including the muscle hypothesised to be responsible for the locking and triggering of pivot feeding in seahorses (m. adductor arcus palatini). Our data confirmed that the epaxial pre-activation pattern observed previously for pipefish also occurs in seahorses. Similar to the epaxials, the sternohyoideus-hypaxial muscle complex shows prolonged anticipatory activity. Although a considerable variation in displacements of the mouth via head rotation could be observed, it could not be demonstrated that seahorses have control over strike distance. In addition, we could not identify the source of the kinematic variability in the activation patterns of the associated muscles. Finally, the stimulation experiment supported the previously hypothesized role of the m. adductor arcus palatini as the trigger in this elastic recoil system. Our results show that pre-stressing of both the head elevators and the hyoid retractors is taking place. As pre-activation of the main muscles involved in pivot feeding has now been demonstrated for both seahorses and pipefish, this is probably a generalized trait of Syngnathidae.     

Induced feeding preferences in dicofol-resistant two-spotted spider mites (Acari: Tetranychidae)

Preadult rearing conditions affected the behavior of dicofol-resistant two-spotted spider mites (Tetranychus urticae). Resistant spider mites reared on dicofol-treated leaves initiated a significantly greater number of feeding bouts on dicofol-treated leaves than did genetically identical spider mites reared on residue-free leaves. Therefore the prior exposure of resistant spider mites resulted in induced feeding preferences that could exacerbate the potential outcome of the resistance by resulting in greater amounts of feeding by resistant individuals on dicofol-treated areas. Since resistant individuals that had not experienced dicofol in their lifetime did not display this feeding preference, avoidance of this phenomenon of induced feeding preference may be an undescribed value of rotations of pesticides.