Studies on the comparative biology of Aphanomyces invadans

Aphanomyces invadans Willoughby et al, 1995 (as A. invaderis) is the recently-named Oomycete fungus that has been shown to be involved in EUS (epizootic ulcerative syndrome), a highly damaging disease of wild and cultured, Asian freshwater and estuarine fishes. The present study shows that A. invadans is the only species, out of a number of isolates from EUS-affected areas in Thailand, that is capable of sustained growth in snakehead fish muscle tissue and reproducing EUS lesions, and is therefore pathognomic to the disease. A. invadans is characterised, and distinguished from the saprophytic isolates, by means of: growth at various temperatures; growth on different media; level of extracellular enzymes produced; susceptibility to various chemicals; aspects of zoospore and germling behaviour; ultrastructure; immunocytochemistry; protein and carbohydrate electrophoresis banding patterns; lectin and polyclonal antibody binding characteristics by means of Western blot analysis; biochemical fingerprinting using pyrolysis mass spectra (PyMS); and molecular studies involving random amplification of polymorphic DNA (RAPD). A. invadans is shown to be indistinguishable from pathogenic Aphanomyces isolates from two other fish diseases, namely Japanese mycotic granulomatosis (MG) and Australian red spot disease (RSD) using the techniques described above. RAPD analyses, in particular, showed that a wide ränge of EUS, MG and RSD isolates are not only conspecific, but probably constitute a single genetic clone. This strongly suggests that it is A. invadans, and not any other biological aetiology, that has spread across Asia causing ulcerative disease in fish. It is recommended that the name A. invadans is used to describe all EUS, MG and RSD pathogenic isolates. This work also shows that Aphanomyces isolates obtained from outbreaks of ulcerative mycosis (UM) of American menhaden, are distinct from A. invadans, and more similar to the saprophytic fungus Aphanomyces laevis. It is conjectured that the invasive UM pathogen has not been studied and that this may show greater similarity to A. invadans. In comparison to the other species tested, A. invadans is most similar to the crayfish plague fungus. Aphanomyces astaci, although A. invadans is shown to be unable to infect noble crayfish {Astacus astacus). Snakeheads {Channa striata) are shown to produce antibodies in response to infection by A. invadans, a finding which may have implications for the possible future development of vaccines. A. invadans is shown to be culturally and ultrastructurally less robust, and more susceptible to chemical treatment, than other saprolegniacean fungi tested, indicating that strategic water treatments, before fish are infected, should be a relatively effective means of control. It is argued that the culturally-fastidious nature of A. invadans could also indicate an inability to compete with natural saprophytes, that may act to restrict it to a pathogenic lifestyle. Possible adaptations of zoospores to pathogenicity include particular chemotactic behaviour; a capability for limited polyplanetism in the presence of a nutrient background, indirect germination, and a form of abbreviated life-cycle. An usually thin zoospore cyst wall, that appears to lack much of the encystment vesicle-derived material apparent on other saprolegniaceans, is believed to have some significance to the ecology of A. invadans, although what this may be is undetermined. Despite the obvious ability of A. invadans to degenerate muscle tissue in fish, cultures showed relatively low production of extracellular enzymes using agar diffusion techniques, indicating that protease activity may be induced. A. invadans zoospores and cysts' have distinctive lectin-binding characteristics, and of particular interest is their ability to cross-react with monoclonal antibodies raised against Phytophthora cinnamomi, a non-saprolegniacean Oomycete. Other features of A. invadans that may provide useful species-specific taxonomic markers include temperature-growth characters, a putative K body organelle with a distinctive substructure, specific electrophoretic bands, pyrolysis mass spectra (used here for the first time in Oomycete systematics), and RAPD fingerprints. Polyclonal antibodies (PAbs) proved very non-specific, but peroxidase

Tissue Doppler imaging following paediatric cardiac surgery : early patterns of change and relationship to outcome

In this study, tissue Doppler imaging (TDI) was used to assess changes in ventricular function following repair of congenital heart defects. The relationship between TDI indices, myocardial injury and clinical outcome was explored. Forty-five children were studied; 35 withcardiac lesions and 10 controls. TDI was performed preoperatively, on admission to paediatric intensive care unit (PICU) and day 1. Regional myocardial Doppler signals were acquired from the right ventricle (RV), left ventricle (LV) and septum. TDI indices included: peak systolicvelocities, isovolumetric velocities (IVV) and isovolumetric acceleration (IVA). Preoperatively, bi-ventricular TDI velocities in the study groupwere reduced compared with normal controls. Postoperatively, RV velocities were significantly reduced and this persisted to day-1 (PreOp vs. PICU and day-1: 7.7+2.2 vs. 3.4+1.0, P < 0.0001 and 3.55+1.29, P < 0.0001). LV velocities initially declined but recovered towards baseline by day-1 (PreOp vs. PICU: 5.31+1.50 vs. 3.51+1.23, P < 0.0001). Isovolumetric parameters in all regions were reduced throughout the postoperative period. Troponin-I release correlated with longer X-clamp times (r=0.82, P < 0.0001) and reduced RV velocities (r=0.42, P=0.028). Reduced pre- and postoperative LV velocities correlated with longer ventilation (PreOp: r=0.54, P=0.002; PostOp: r=0.42, P=0.026). This study identified reduced postoperative RV velocities correlated with myocardial injury while reduced LV TDI correlated with longer postoperative ventilation

Environmentally co-occurring mercury resistance plasmids are genetically and phenotypically diverse and confer variable context-dependent fitness effects

Plasmids are important mobile elements that can facilitate genetic exchange and local adaptation within microbial communities. We compared the sequences of four co-occurring pQBR-family environmental mercury resistance plasmids and measured their effects on competitive fitness of a Pseudomonas fluorescens SBW25 host, which was isolated at the same field site. Fitness effects of carriage differed between plasmids and were strongly context dependent, varying with medium, plasmid status of competitor and levels of environmental mercury. The plasmids also varied widely in their rates of conjugation and segregational loss. We found that few of the plasmid-borne accessory genes could be ascribed functions, although we identified a putative chemotaxis operon, a type IV pilus-encoding cluster, and a region encoding putative arylsulfatase enzymes, which were conserved across geographically distant isolates. One plasmid, pQBR55, conferred the ability to catabolise sucrose. Transposons, including the mercury resistance Tn5042, appeared to have been acquired by the different pQBR plasmids by recombination, indicating an important role for horizontal gene transfer in the recent evolution of pQBR plasmids. Our findings demonstrate extensive genetic and phenotypic diversity amongst co-occurring members of a plasmid community and suggest a role for environmental heterogeneity in the maintenance of plasmid diversity

PERCIVAL mission to Mars

With the downturn of the world economy, the priority of unmanned exploration of the solar system has been lowered. Instead of foregoing all missions to our neighbors in the solar system, a new philosophy of exploration mission design has evolved to insure the continued exploration of the solar system. The 'Discovery-class' design philosophy uses a low cost, limited mission, available technology spacecraft instead of the previous 'Voyager-class' design philosophy that uses a 'do-everything at any cost' spacecraft. The Percival Mission to Mars was proposed by Ares Industries as one of the new 'Discovery-class' of exploration missions. The spacecraft will be christened Percival in honor of American astronomer Percival Lowell who proposed the existence of life on Mars in the early twentieth century. The main purpose of the Percival mission to Mars is to collect and relay scientific data to Earth suitable for designing future manned and unmanned missions to Mars. The measurements and observations made by Percival will help future mission designers to choose among landing sites based on the feasibility and scientific interest of the sites. The primary measurements conducted by the Percival mission include gravity field determination, surface and atmospheric composition, sub-surface soil composition, sub-surface seismic activity, surface weather patterns, and surface imaging. These measurements will be taken from the orbiting Percival spacecraft and from surface penetrators deployed from Mars orbit. The design work for the Percival Mission to Mars was divided among four technical areas: Orbits and Propulsion System, Surface Penetrators, Gravity and Science Instruments, and Spacecraft Structure and Systems. The results for each of the technical areas is summarized and followed by a design cost analysis and recommendations for future analyses

Misclassification of Plasmodium infections by conventional microscopy and the impact of remedial training on the proficiency of laboratory technicians in species identification.

BACKGROUND: Malaria diagnosis is largely dependent on the demonstration of parasites in stained blood films by conventional microscopy. Accurate identification of the infecting Plasmodium species relies on detailed examination of parasite morphological characteristics, such as size, shape, pigment granules, besides the size and shape of the parasitized red blood cells and presence of cell inclusions. This work explores misclassifications of four Plasmodium species by conventional microscopy relative to the proficiency of microscopists and morphological characteristics of the parasites on Giemsa-stained blood films. CASE DESCRIPTION: Ten-day malaria microscopy remedial courses on parasite detection, species identification and parasite counting were conducted for public health and research laboratory personnel. Proficiency in species identification was assessed at the start (pre) and the end (post) of each course using known blood films of Plasmodium falciparum, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax infections with densities ranging from 1,000 to 30,000 parasites/μL. Outcomes were categorized as false negative, positive without speciation, P. falciparum, P. malariae, P. ovale, P. vivax and mixed infections. DISCUSSION AND EVALUATION: Reported findings are based on 1,878 P. falciparum, 483 P. malariae, 581 P. ovale and 438 P. vivax cumulative results collated from 2008 to 2010 remedial courses. Pre-training false negative and positive misclassifications without speciation were significantly lower on P. falciparum infections compared to non-falciparum infections (p < 0.0001). Post-training misclassifications decreased significantly compared to pre- training misclassifications which in turn led to significant improvements in the identification of the four species. However, P. falciparum infections were highly misclassified as mixed infections, P. ovale misclassified as P. vivax and P. vivax similarly misclassified as P. ovale (p < 0.05). CONCLUSION: These findings suggest that the misclassification of malaria species could be a common occurrence especially where non-falciparum infections are involved due to lack of requisite skills in microscopic diagnosis and variations in morphological characteristics within and between Plasmodium species. Remedial training might improve reliability of conventional light microscopy with respect to differentiation of Plasmodium infections

α-Klotho expressison in human tissue

Context: α-Klotho has emerged as a powerful regulator of the aging process. To-date, the expression profile of α-Klotho in human tissues is unknown and its existence in some human tissue types is subject to much controversy. Objective: This is the first study to characterize system-wide tissue expression of transmembrane α-Klotho in humans. We have employed next generation targeted proteomic analysis using Parallel Reaction Monitoring (PRM) in parallel with conventional antibody-based methods to determine the expression and spatial distribution of human α-Klotho expression in health. Results: The distribution of α-Klotho in human tissues from various organ systems, including arterial, epithelial, endocrine, reproductive and neuronal tissues was first identified by immunohistochemistry. Kidney tissues showed strong α-Klotho expression, while liver did not reveal a detectable signal. These results were next confirmed by western blotting of both whole tissues and primary cells. To validate our antibody-based results, α-Klotho expressing tissues were subjected to PRM mass spectrometry identifying peptides specific for the full length, transmembrane α-Klotho isoform. Conclusions: The data presented confirms α-Klotho expression in the kidney tubule and in artery, and provides evidence of α-Klotho expression across organ systems and cell-types that have not previously been described in humans.K.L. received a Genzyme-Sanofi Fellowship in Nephrology grant. T.F.H. is funded by the NIHR award to the Cambridge Biomedical Research Centre and by NIHR grant 14/49/147. The Cambridge Aorta Study is funded by the British Heart Foundation.This is the author accepted manuscript. The final version is available from the Endocrine Society via http://dx.doi.org/10.1210/jc.2015-1800

Mixed-Methods Evaluation of Real-Time Safety Reporting by Hospitalized Patients and Their Care Partners:The MySafeCare Application

OBJECTIVE: To evaluate the amount and content of data patients and carepartners reported using a real-time electronic safety tool compared to other reporting mechanisms, and understand their perspectives on safety concerns and reporting in the hospital. METHODS: Mixed-methods study including 20 month pre- and post-implementation trial evaluating MySafeCare, a web-based application which allows hospitalized patients/carepartners to report safety concerns in real-time. Comparison of MySafeCare submission rates for three hospital units (oncology acute care; vascular intermediate care; medical intensive care) to submissions rates of Patient Family Relations (PFR) Department, a hospital service to address patient/family concerns. Triangulation of quantitative data with thematic analysis of safety concern submissions and patient/carepartner interviews to understand submission content and perspectives on safety reporting. RESULTS: Thirty-two MySafeCare submissions were received with an average rate of 1.7 submissions per 1,000 patient-days and a range of 0.3 to 4.8 submissions per 1,000 patient-days across all units, indicating notable variation between units. MySafeCare submission rates were significantly higher than PFR submission rates during the post-intervention period on the vascular unit (4.3 [95% CI 2.8 – 6.5] versus 1.5 [95% CI 0.7 – 3.1], Poisson) (P=0.01). Overall trends indicated a decrease in PFR submissions after MySafeCare implementation. Triangulated data indicated patients preferred to report anonymously and did not want concerns submitted directly to their care team. CONCLUSIONS: MySafeCare evaluation confirmed the potential value of providing an electronic, anonymous reporting tool in the hospital to capture safety concerns in real-time. Such applications should be tested further as part of patient safety programs

α-Klotho Expression in Human Tissues.

CONTEXT: α-Klotho has emerged as a powerful regulator of the aging process. To date, the expression profile of α-Klotho in human tissues is unknown, and its existence in some human tissue types is subject to much controversy. OBJECTIVE: This is the first study to characterize systemwide tissue expression of transmembrane α-Klotho in humans. We have employed next-generation targeted proteomic analysis using parallel reaction monitoring in parallel with conventional antibody-based methods to determine the expression and spatial distribution of human α-Klotho expression in health. RESULTS: The distribution of α-Klotho in human tissues from various organ systems, including arterial, epithelial, endocrine, reproductive, and neuronal tissues, was first identified by immunohistochemistry. Kidney tissues showed strong α-Klotho expression, whereas liver did not reveal a detectable signal. These results were next confirmed by Western blotting of both whole tissues and primary cells. To validate our antibody-based results, α-Klotho-expressing tissues were subjected to parallel reaction monitoring mass spectrometry (data deposited at ProteomeXchange, PXD002775) identifying peptides specific for the full-length, transmembrane α-Klotho isoform. CONCLUSIONS: The data presented confirm α-Klotho expression in the kidney tubule and in the artery and provide evidence of α-Klotho expression across organ systems and cell types that has not previously been described in humans.K.L. received a Genzyme-Sanofi Fellowship in Nephrology grant. T.F.H. is funded by the NIHR award to the Cambridge Biomedical Research Centre and by NIHR grant 14/49/147. The Cambridge Aorta Study is funded by the British Heart Foundation.This is the author accepted manuscript. The final version is available from the Endocrine Society via http://dx.doi.org/10.1210/jc.2015-1800

Gene mobility promotes the spread of resistance in bacterial populations

Theory predicts that horizontal gene transfer (HGT) expands the selective conditions under which genes spread in bacterial populations. Whereas vertically inherited genes can only spread by positively selected clonal expansion, mobile genetic elements can drive fixation of genes by infectious HGT. We tested this using populations of Pseudomonas fluorescens and the conjugative mercury resistance (Hg R) plasmid pQBR57. HGT expanded the selective conditions allowing the spread of Hg R: Chromosomal Hg R only increased in frequency under positive selection, whereas plasmid-encoded Hg R reached fixation with or without positive selection. Tracking plasmid dynamics over time revealed that the mode of Hg R inheritance varied across mercury environments. Under mercury selection, the spread of Hg R was driven primarily by clonal expansion while in the absence of mercury Hg R dynamics were dominated by infectious transfer. Thus, HGT is most likely to drive the spread of resistance genes in environments where resistance is useless

Wildlife Trade and Global Disease Emergence

The global trade in wildlife provides disease transmission mechanisms that not only cause human disease outbreaks but also threaten livestock, international trade, rural livelihoods, native wildlife populations, and the health of ecosystems. Outbreaks resulting from wildlife trade have caused hundreds of billions of dollars of economic damage globally. Rather than attempting to eradicate pathogens or the wild species that may harbor them, a practical approach would include decreasing the contact rate among species, including humans, at the interface created by the wildlife trade. Since wildlife marketing functions as a system of scale-free networks with major hubs, these points provide control opportunities to maximize the effects of regulatory efforts