Detection of Pseudomonas aeruginosa in sputum samples by modified fluorescent in situ hybridization

Pseudomonas aeruginosa is the most common and dominant infectious agent that causes chronic pneumonia in patients with cystic fibrosis (CF). Fluorescent in situ hybridization (FISH) is a powerfulmolecular method for the specific and rapid diagnosis of bacteria, including the detection of P. aeruginosa in sputum samples from CF patients. High background fluorescence of viscous sputumsamples obtained from CF patients may impede detection of microorganisms by FISH. The aim of this study was to test the application of biotin during FISH technique to reduce unspecific background fluorescence in sputum samples to facilitate and improve detection of P. aeruginosa. Sixty-three sputum samples from CF patients were tested by FISH to detect P. aeruginosa. All the 63 samples were also examined by a modified FISH procedure including biotin treatment. The FISH results were compared with those of conventional culture method. The specificity of FISH was 100%. The sensitivity of FISH for detection of P. aeruginosa from samples without biotin treatment was 83.3%, whereas in biotin-treated samples was 88.1%. Biotin reduced background fluorescence of 12 sputum samples of CF patients and it did not show any adverse effect on FISH results of the remaining sputum samples. Therefore, using of biotin in FISH procedure seems to facilitate and improve the detection of respiratorytract infections by P. aeruginosa in this population

Pressure dependence of diffusion in simple glasses and supercooled liquids

Using molecular dynamics simulation, we have calculated the pressure dependence of the diffusion constant in a binary Lennard-Jones Glass. We observe four temperature regimes. The apparent activation volume drops from high values in the hot liquid to a plateau value. Near the critical temperature of the mode coupling theory it rises steeply, but in the glassy state we find again small values, similar to the ones in the liquid. The peak of the activation volume at the critical temperature is in agreement with the prediction of mode coupling theory

Distribution, expression and long range mapping of legiolysin gene (lly) specific DNA sequences in Legionellae

The legiolysin gene (lly) cloned from Legionella pneumophila Philadelphia 1 confers the phenotypes of hemolysis and browning of the culture medium. An internal Uy-specific DNA probe was used in Southern hybridizations for the detection of Uy-specific DNA in the genomes of legioneUae and other gram-negative pathogenic bacteria. Under conditi9ns of high stringency, tlie Uy DNA probe specifically reacted with DNA fragments fr9m L. pneumophiüz isolates; by reducing stringency, hybridization was also observed for all other Legionella strains tested. No hybridization occurred with DNAs isolated from bact~ria of other genera. The Uy genewas mapped by pulsed-field gel electrophoresis to the respective genomic Notl fragments of Legionelltz isolates. By using antilegiolysin monospecific polyclonal antibodies in Western blots (immunoblots), Lly proteins could be detected only in L. pneumophila isolates

A lack of dynamic triggering of slow slip and tremor indicates that the shallow Cascadia megathrust offshore Vancouver Island is likely locked

Author Posting. © American Geophysical Union, 2018. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geophysical Research Letters 45 (2018): 11,095-11,103, doi:10.1029/2018GL079519.Great subduction zone earthquakes vary considerably in the updip extent of megathrust rupture. It is unclear if this diversity reflects variations in interseismic strain accumulation owing to the limited number of subduction zones with seafloor monitoring. We use a borehole seismic‐geodetic observatory installed at the updip end of the Cascadia fault offshore Vancouver Island to show that the megathrust there does not appear to slip in triggered tremor or slow‐slip events when subjected to moderate dynamic stress transients. Borehole tilt and seismic data from recent teleseismic M7.6–8.1 earthquakes demonstrate a lack of triggered slow slip above the Mw 4.0 level and an absence of triggered tremor despite shear‐stress transients of 1–10 kPa that were sufficient to trigger tremor on the downdip end of the interface. Our observations are most consistent with a model in which the Cascadia fault offshore Vancouver Island is locked all the way to the trench.NSF Grant Numbers: OCE‐1259243, OCE‐1259718; W. M. Keck Foundation2019-04-2

Seasonal variation of Pneumocystis jirovecii infection: analysis of underlying climatic factors

AbstractPneumocystis jirovecii causes severe pneumonia (PCP) in immunocompromised patients. Seasonal changes of PCP incidence may be associated with climate changes. In this first study using multiple linear regression statistics to assess monthly climatic data and Pneumocystis, PCP incidence was positively correlated with mean temperature, but not with rainfall or wind strength

A long-term geothermal observatory across subseafloor gas hydrates, IODP Hole U1364A, Cascadia accretionary prism

© The Author(s), 2020. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in WHOI Becker, K., Davis, E. E., Heesemann, M., Collins, J. A., & McGuire, J. J. A long-term geothermal observatory across subseafloor gas hydrates, IODP Hole U1364A, Cascadia accretionary prism. Frontiers in Earth Science, 8, (2020): 568566, https://doi.org/10.3389/feart.2020.568566We report 4 years of temperature profiles collected from May 2014 to May 2018 in Integrated Ocean Drilling Program Hole U1364A in the frontal accretionary prism of the Cascadia subduction zone. The temperature data extend to depths of nearly 300 m below seafloor (mbsf), spanning the gas hydrate stability zone at the location and a clear bottom-simulating reflector (BSR) at ∼230 mbsf. When the hole was drilled in 2010, a pressure-monitoring Advanced CORK (ACORK) observatory was installed, sealed at the bottom by a bridge plug and cement below 302 mbsf. In May 2014, a temperature profile was collected by lowering a probe down the hole from the ROV ROPOS. From July 2016 through May 2018, temperature data were collected during a nearly two-year deployment of a 24-thermistor cable installed to 268 m below seafloor (mbsf). The cable and a seismic-tilt instrument package also deployed in 2016 were connected to the Ocean Networks Canada (ONC) NEPTUNE cabled observatory in June of 2017, after which the thermistor temperatures were logged by Ocean Networks Canada at one-minute intervals until failure of the main ethernet switch in the integrated seafloor control unit in May 2018. The thermistor array had been designed with concentrated vertical spacing around the bottom-simulating reflector and two pressure-monitoring screens at 203 and 244 mbsf, with wider thermistor spacing elsewhere to document the geothermal state up to seafloor. The 4 years of data show a generally linear temperature gradient of 0.055°C/m consistent with a heat flux of 61–64 mW/m2. The data show no indications of thermal transients. A slight departure from a linear gradient provides an approximate limit of ∼10−10 m/s for any possible slow upward advection of pore fluids. In-situ temperatures are ∼15.8°C at the BSR position, consistent with methane hydrate stability at that depth and pressure.KB was supported by NSF grant OCE-1259718 for construction and deployment of the thermistor cable in the hole. Construction of the seismic-strain-tilt instrumentation was supported by a Keck Foundation grant to WHOI, and deployment and recovery of the integrated sensor string was supported by NSF grant OCE-1259243 to JM and JC. Support for the pressure-monitoring instrumentation and 2014 CTD profile was provided by the Geological Survey of Canada and Ocean Networks Canada

Yersinia V–Antigen Exploits Toll-like Receptor 2 and CD14 for Interleukin 10–mediated Immunosuppression

A characteristic of the three human-pathogenic Yersinia spp. (the plague agent Yersinia pestis and the enteropathogenic Yersinia pseudotuberculosis and Yersinia enterocolitica) is the expression of the virulence (V)-antigen (LcrV). LcrV is a released protein which is involved in contact-induced secretion of yersinia antihost proteins and in evasion of the host's innate immune response. Here we report that recombinant LcrV signals in a CD14- and toll-like receptor 2 (TLR2)-dependent fashion leading to immunosuppression by interleukin 10 induction. The impact of this immunosuppressive effect for yersinia pathogenesis is underlined by the observation that TLR2-deficient mice are less susceptible to oral Y. enterocolitica infection than isogenic wild-type animals. In summary, these data demonstrate a new ligand specificity of TLR2, as LcrV is the first known secreted and nonlipidated virulence-associated protein of a Gram-negative bacterium using TLR2 for cell activation. We conclude that yersiniae might exploit host innate pattern recognition molecules and defense mechanisms to evade the host immune response

Ocean Observatories as a Tool to Advance Gas Hydrate Research

Since 2009, unprecedented comprehensive long-term gas hydrate observations have become available from Ocean Networks Canada's NEPTUNE cabled ocean observatory at the northern Cascadia margin. Several experiments demonstrate the scientific importance of permanent power and Internet connectivity to the ocean floor as they have advanced the field of gas hydrate related research. One example is the cabled crawler Wally at Barkley Canyon, enabling live in situ exploration of the hydrate mounds and its associated benthic communities through the crawler's mobility and permanent accessibility throughout the year. Another example is a bubble-imaging sonar at Clayoquot Slope, revealing the strong relationship between ebullition of natural gas and tidal pressure, without apparent correlation to earthquakes, storms, or temperature fluctuations, in year-long continuous recordings. Finally, regular observatory maintenance cruises allow additional science sampling including echo-sounder surveys to extend the observatory footprint. Long-term trends in the data are not yet apparent but can also become evident from continuous measurements, as ocean observatories such as NEPTUNE are built for a 25-year lifetime, and expansion of the observatory networks makes these findings comparable and testable. \ud Plain Language Summary Natural gas near the ocean floor creates a rapidly changing environment where it is important to collect data continuously in order to determine the magnitude, speed, and potential mechanism of change. This long-standing challenge of year-round access to the deep ocean has been tackled by Ocean Networks Canada through cabling the northern Cascadia seafloor, providing power and Internet communication-ideal for power-hungry instruments, large data volumes, and real-time access. The presence of gas influences the shape of the seafloor, animal activity, and potential escape of methane, a potent greenhouse gas. A seafloor crawler Wally was operated around deep canyon mounds of gas hydrate (a solid gas-water composite) since 2009 and helped discover environmental changes influencing sea life. Further along the continental slope, an acoustic sonar monitored rising methane bubbles where the bubbling appears to be controlled neither by earthquakes, winter storms, nor subtle temperature changes but actually strongly by tidal pressure. Regular maintenance of the observatory by ship allows more data to be collected near the cabled seafloor sites, extending the observations to a larger area. Ocean observatories are built to last decades and therefore more data for more research can be collected, potentially detecting relatively slow processes as well

Germination of phagocytosed E. cuniculi spores does not significantly contribute to parasitophorous vacuole formation in J774 cells

The obligate intracellular microsporidia have developed a unique invasion mechanism to infect their host cells. Spores explosively evert a tube-like structure and extrude the infectious spore content through this organelle into the host cell. Spores from species of the genus Encephalitozoon were also shown to be efficiently internalized by phagocytosis, which led to the hypothesis that spore germination from inside a phagosome might contribute to the infection process. Here, we challenge this hypothesis by quantifying Encephalitozoon cuniculi infection rates of J774 cells that were incubated with the phagocytosis inhibitor cytochalasin D. We demonstrate that the invasion rate in cytochalasin D-treated cells is identical to untreated controls, although phagocytic uptake of E. cuniculi spores was less than 10% of control samples. This study suggests that germination of phagocytosed spores is not a significant infection mode for E. cuniculi