Subversion of actin dynamics by EspM effectors of attaching and effacing bacterial pathogens

Rho GTPases are common targets of bacterial toxins and type III secretion system effectors. IpgB1 and IpgB2 of Shigella and Map of enteropathogenic (EPEC) and enterohemorrhagic (EHEC) Escherichia coli were recently grouped together on the basis that they share a conserved WxxxE motif. In this study, we characterized six WxxxE effectors from attaching and effacing pathogens: TrcA and EspM1 of EPEC strain B171, EspM1 and EspM2 of EHEC strain Sakai and EspM2 and EspM3 of Citrobacter rodentium. We show that EspM2 triggers formation of global parallel stress fibres, TrcA and EspM1 induce formation of localized parallel stress fibres and EspM3 triggers formation of localized radial stress fibres. Using EspM2 and EspM3 as model effectors, we report that while substituting the conserved Trp with Ala abolished activity, conservative Trp to Tyr or Glu to Asp substitutions did not affect stress-fibre formation. We show, using dominant negative constructs and chemical inhibitors, that the activity of EspM2 and EspM3 is RhoA and ROCK-dependent. Using Rhotekin pull-downs, we have shown that EspM2 and EspM3 activate RhoA; translocation of EspM2 and EspM3 triggered phosphorylation of cofilin. These results suggest that the EspM effectors modulate actin dynamics by activating the RhoA signalling pathway

Algebraic Approach to Shape Invariance

The integrability condition called shape invariance is shown to have an underlying algebraic structure and the associated Lie algebras are identified. These shape-invariance algebras transform the parameters of the potentials such as strength and range. Shape-invariance algebras, in general, are shown to be infinite-dimensional. The conditions under which they become finite-dimensional are explored.Comment: Submitted to Physical Review A. Latex file, 9 pages. Manuscript is also available at http://nucth.physics.wisc.edu/preprints

Human Salmonella Typhi exposure generates differential multifunctional cross‐reactive T‐cell memory responses against Salmonella Paratyphi and invasive nontyphoidal Salmonella

Objective There are no vaccines for most of the major invasive Salmonella strains causing severe infection in humans. We evaluated the specificity of adaptive T memory cell responses generated after Salmonella Typhi exposure in humans against other major invasive Salmonella strains sharing capacity for dissemination. Methods T memory cells from eleven volunteers who underwent controlled oral challenge with wt S. Typhi were characterised by flow cytometry for cross‐reactive cellular cytokine/chemokine effector responses or evidence of degranulation upon stimulation with autologous B‐lymphoblastoid cells infected with either S. Typhi, Salmonella Paratyphi A (PA), S. Paratyphi B (PB) or an invasive nontyphoidal Salmonella strain of the S. Typhimurium serovar (iNTSTy). Results Blood T‐cell effector memory (TEM) responses after exposure to S. Typhi in humans evolve late, peaking weeks after infection in most volunteers. Induced multifunctional CD4+ Th1 and CD8+ TEM cells elicited after S. Typhi challenge were cross‐reactive with PA, PB and iNTSTy. The magnitude of multifunctional CD4+ TEM cell responses to S. Typhi correlated with induction of cross‐reactive multifunctional CD8+ TEM cells against PA, PB and iNTSTy. Highly multifunctional subsets and T central memory and T effector memory cells that re‐express CD45 (TEMRA) demonstrated less heterologous T‐cell cross‐reactivity, and multifunctional Th17 elicited after S. Typhi challenge was not cross‐reactive against other invasive Salmonella. Conclusion Gaps in cross‐reactive immune effector functions in human T‐cell memory compartments were highly dependent on invasive Salmonella strain, underscoring the importance of strain‐dependent vaccination in the design of T‐cell‐based vaccines for invasive Salmonella

Association Between Lifetime Marijuana Use and Cognitive Function in Middle Age: The Coronary Artery Risk Development in Young Adults (CARDIA) Study.

Marijuana use is increasingly common in the United States. It is unclear whether it has long-term effects on memory and other domains of cognitive function. To study the association between cumulative lifetime exposure to marijuana use and cognitive performance in middle age. We used data from the Coronary Artery Risk Development in Young Adults (CARDIA) study, a cohort of 5115 black and white men and women aged 18 to 30 years at baseline from March 25, 1985, to June 7, 1986 (year 0), and followed up over 25 years from June 7, 1986, to August 31, 2011, to estimate cumulative years of exposure to marijuana (1 year = 365 days of marijuana use) using repeated measures and to assess associations with cognitive function at year 25. Linear regression was used to adjust for demographic factors, cardiovascular risk factors, tobacco smoking, use of alcohol and illicit drugs, physical activity, depression, and results of the mirror star tracing test (a measure of cognitive function) at year 2. Data analysis was conducted from June 7, 1986, to August 31, 2011. Three domains of cognitive function were assessed at year 25 using the Rey Auditory Verbal Learning Test (verbal memory), the Digit Symbol Substitution Test (processing speed), and the Stroop Interference Test (executive function). Among 3385 participants with cognitive function measurements at the year 25 visit, 2852 (84.3%) reported past marijuana use, but only 392 (11.6%) continued to use marijuana into middle age. Current use of marijuana was associated with worse verbal memory and processing speed; cumulative lifetime exposure was associated with worse performance in all 3 domains of cognitive function. After excluding current users and adjusting for potential confounders, cumulative lifetime exposure to marijuana remained significantly associated with worse verbal memory. For each 5 years of past exposure, verbal memory was 0.13 standardized units lower (95% CI, -0.24 to -0.02; P = .02), corresponding to a mean of 1 of 2 participants remembering 1 word fewer from a list of 15 words for every 5 years of use. After adjustment, we found no associations with lower executive function (-0.03 [95% CI, -0.12 to 0.07]; P = .56) or processing speed (-0.04 [95% CI, -0.16 to 0.08]; P = .51). Past exposure to marijuana is associated with worse verbal memory but does not appear to affect other domains of cognitive function

A radium assay technique using hydrous titanium oxide adsorbent for the Sudbury Neutrino Observatory

As photodisintegration of deuterons mimics the disintegration of deuterons by neutrinos, the accurate measurement of the radioactivity from thorium and uranium decay chains in the heavy water in the Sudbury Neutrino Observatory (SNO) is essential for the determination of the total solar neutrino flux. A radium assay technique of the required sensitivity is described that uses hydrous titanium oxide adsorbent on a filtration membrane together with a beta-alpha delayed coincidence counting system. For a 200 tonne assay the detection limit for 232Th is a concentration of 3 x 10^(-16) g Th/g water and for 238U of 3 x 10^(-16) g U/g water. Results of assays of both the heavy and light water carried out during the first two years of data collection of SNO are presented.Comment: 12 pages, 4 figure

Electroosmotic flow of biorheological micropolar fluids through microfluidic channels

An analysis is presented in this work to assess the influence of micropolar nature of fluids in fully developed flow induced by electrokinetically driven peristaltic pumping through a parallel plate microchannel. The walls of the channel are assumed as sinusoidal wavy to analyze the peristaltic flow nature. We consider that the wavelength of the wall motion is much larger as compared to the channel width to validate the lubrication theory. To simplify the Poisson Boltzmann equation, we also use the Debye-Hückel linearization (i.e. wall zeta potential ≤ 25mV). We consider governing equation for micropolar fluid in absence of body force and couple effects however external electric field is employed. The solutions for axial velocity, spin velocity, flow rate, pressure rise and stream functions subjected to given physical boundary conditions are computed. The effects of pertinent parameters like Debye length and Helmholtz-Smoluchowski velocity which characterize the EDL phenomenon and external electric field, coupling number and micropolar parameter which characterize the micropolar fluid behavior, on peristaltic pumping are discussed through the illustrations. The results show that peristaltic pumping may alter by applying external electric fields. This model can be used to design and engineer the peristalsis-lab-on-chip and micro peristaltic syringe pumps for biomedical applications

Mucosal-Associated Invariant T cells exhibit distinct functional signatures associated with protection against typhoid fever

We have previously demonstrated that Mucosal-Associated Invariant T (MAIT) cells secrete multiple cytokines after exposure to Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever in humans. However, whether cytokine secreting MAIT cells can enhance or attenuate the clinical severity of bacterial infections remain debatable. This study characterizes human MAIT cell functions in subjects participating in a wild-type S. Typhi human challenge model. Here, we found that MAIT cells exhibit distinct functional signatures associated with protection against typhoid fever. We also observed that the cytokine patterns of MAIT cell responses, rather than the average number of cytokines expressed, are more predictive of typhoid fever outcomes. These results might enable us to objectively, based on functional parameters, identify cytokine patterns that may serve as predictive biomarkers during natural infection and vaccination

Oral Wild-Type Salmonella Typhi Challenge Induces Activation of Circulating Monocytes and Dendritic Cells in Individuals Who Develop Typhoid Disease.

A new human oral challenge model with wild-type Salmonella Typhi (S. Typhi) was recently developed. In this model, ingestion of 104 CFU of Salmonella resulted in 65% of subjects developing typhoid fever (referred here as typhoid diagnosis -TD-) 5-10 days post-challenge. TD criteria included meeting clinical (oral temperature ≥38°C for ≥12 h) and/or microbiological (S. Typhi bacteremia) endpoints. One of the first lines of defense against pathogens are the cells of the innate immune system (e.g., monocytes, dendritic cells -DCs-). Various changes in circulating monocytes and DCs have been described in the murine S. Typhimurium model; however, whether similar changes are present in humans remains to be explored. To address these questions, a subset of volunteers (5 TD and 3 who did not develop typhoid despite oral challenge -NoTD-) were evaluated for changes in circulating monocytes and DCs. Expression of CD38 and CD40 were upregulated in monocytes and DCs in TD volunteers during the disease days (TD-0h to TD-96h). Moreover, integrin α4β7, a gut homing molecule, was upregulated on monocytes but not DCs. CD21 upregulation was only identified in DCs. These changes were not observed among NoTD volunteers despite the same oral challenge. Moreover, monocytes and DCs from NoTD volunteers showed increased binding to S. Typhi one day after challenge. These monocytes showed phosphorylation of p38MAPK, NFkB and Erk1/2 upon stimulation with S. Typhi-LPS-QDot micelles. In contrast, monocytes from TD volunteers showed only a moderate increase in S. Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation. This is the first study to describe different activation and migration profiles, as well as differential signaling patterns, in monocytes and DCs which relate directly to the clinical outcome following oral challenge with wild type S. Typhi

Enteropathogenic Escherichia coli (EPEC) inactivate innate immune responses prior to compromising epithelial barrier function

Enteropathogenic Escherichia coli (EPEC) infection of the human small intestine induces severe watery diarrhoea linked to a rather weak inflammatory response despite EPEC's in vivo capacity to disrupt epithelial barrier function. Here, we demonstrate that EPEC flagellin triggers the secretion of the pro-inflammatory cytokine, interleukin (IL)-8, from small (Caco-2) and large (T84) intestinal epithelia model systems. Interestingly, IL-8 secretion required basolateral infection of T84 cells implying that flagellin must penetrate the epithelial barrier. In contrast, apical infection of Caco-2 cells induced IL-8 secretion but less potently than basolateral infections. Importantly, infection of Caco-2, but not T84 cells rapidly inhibited IL-8 secretion by a mechanism dependent on the delivery of effectors through a translocation system encoded on the locus of enterocyte effacement (LEE). Moreover, EPEC prevents the phosphorylation-associated activation of multiple kinase pathways regulating IL-8 gene transcription by a mechanism apparently independent of LEE-encoded effectors and four non-LEE-encoded effectors. Crucially, our studies reveal that EPEC inhibits the capacity of the cells to secrete IL-8 in response to bacterial antigens and inflammatory cytokines prior to disrupting barrier function by a distinct mechanism. Thus, these findings also lend themselves to a plausible mechanism to explain the absence of a strong inflammatory response in EPEC-infected humans