Negative regulation of MurZ and MurA underlies the essentiality of GpsB- and StkP-mediated protein phosphorylation in Streptococcus pneumoniae D39.

GpsB links peptidoglycan synthases to other proteins that determine the shape of the respiratory pathogen Streptococcus pneumoniae (pneumococcus; Spn) and other low-GC Gram-positive bacteria. GpsB is also required for phosphorylation of proteins by the essential StkP(Spn) Ser/Thr protein kinase. Here we report three classes of frequently arising chromosomal duplications (≈21-176 genes) containing murZ (MurZ-family homolog of MurA) or murA that suppress ΔgpsB or ΔstkP. These duplications arose from three different repeated sequences and demonstrate the facility of pneumococcus to modulate gene dosage of numerous genes. Overproduction of MurZ or MurA alone or overproduction of MurZ caused by ΔkhpAB mutations suppressed ΔgpsB or ΔstkP phenotypes to varying extents. ΔgpsB and ΔstkP were also suppressed by MurZ amino-acid changes distant from the active site, including one in commonly studied laboratory strains, and by truncation or deletion of the homolog of IreB(ReoM). Unlike in other Gram-positive bacteria, MurZ is predominant to MurA in pneumococcal cells. However, ΔgpsB and ΔstkP were not suppressed by ΔclpCP, which did not alter MurZ or MurA amounts. These results support a model in which regulation of MurZ and MurA activity, likely by IreB(Spn), is the only essential requirement for StkP-mediated protein phosphorylation in exponentially growing D39 pneumococcal cells.

Chromosomal Duplications of MurZ (MurA2) or MurA (MurA1), Amino Acid Substitutions in MurZ (MurA2), and Absence of KhpAB Obviate the Requirement for Protein Phosphorylation in Streptococcus pneumoniae D39.

GpsB links peptidoglycan synthases to other proteins that determine the shape of the respiratory pathogen Streptococcus pneumoniae (pneumococcus; Spn ) and other low-GC Gram-positive bacteria. GpsB is also required for phosphorylation of proteins by the essential StkP( Spn ) Ser/Thr protein kinase. Here we report three classes of frequently arising chromosomal duplications (≈21-176 genes) containing murZ (MurZ-family homolog of MurA) or murA that suppress Δ gpsB or Δ stkP . These duplications arose from three different repeated sequences and demonstrate the facility of pneumococcus to modulate gene dosage of numerous genes. Overproduction of MurZ or MurA alone or overexpression of MurZ caused by Δ khpAB mutations suppressed Δ gpsB or Δ stkP phenotypes to varying extents. Δ gpsB and Δ stkP were also suppressed by MurZ amino-acid changes distant from the active site, including one in commonly studied laboratory strains, and by truncation or deletion of the homolog of IreB(ReoM). Unlike in other Gram-positive bacteria, MurZ is predominant to MurA in pneumococcal cells. However, Δ gpsB and Δ stkP were not suppressed by Δ clpCP , which did not alter MurZ or MurA amounts. These results support a model in which regulation of MurZ and MurA activity, likely by IreB( Spn ), is the only essential requirement for protein phosphorylation in exponentially growing D39 pneumococcal cells.

S1 Domain RNA-Binding Protein CvfD Is a New Posttranscriptional Regulator That Mediates Cold Sensitivity, Phosphate Transport, and Virulence in Streptococcus pneumoniae D39.

Posttranscriptional gene regulation often involves RNA-binding proteins that modulate mRNA translation and/or stability either directly through protein-RNA interactions or indirectly by facilitating the annealing of small regulatory RNAs (sRNAs). The human pathogen Streptococcus pneumoniae D39 (pneumococcus) does not encode homologs to RNA-binding proteins known to be involved in promoting sRNA stability and function, such as Hfq or ProQ, even though it contains genes for at least 112 sRNAs. However, the pneumococcal genome contains genes for other RNA-binding proteins, including at least six S1 domain proteins: ribosomal protein S1 (rpsA), polynucleotide phosphorylase (pnpA), RNase R (rnr), and three proteins with unknown functions. Here, we characterize the function of one of these conserved, yet uncharacterized, S1 domain proteins, SPD_1366, which we have renamed CvfD (conserved virulence factor D), since loss of the protein results in attenuation of virulence in a murine pneumonia model. We report that deletion of cvfD impacts the expression of 144 transcripts, including the pst1 operon, encoding phosphate transport system 1 in S. pneumoniae We further show that CvfD posttranscriptionally regulates the PhoU2 master regulator of the pneumococcal dual-phosphate transport system by binding phoU2 mRNA and impacting PhoU2 translation. CvfD not only controls expression of phosphate transporter genes but also functions as a pleiotropic regulator that impacts cold sensitivity and the expression of sRNAs and genes involved in diverse cellular functions, including manganese uptake and zinc efflux. Together, our data show that CvfD exerts a broad impact on pneumococcal physiology and virulence, partly by posttranscriptional gene regulation.IMPORTANCE Recent advances have led to the identification of numerous sRNAs in the major human respiratory pathogen S. pneumoniae However, little is known about the functions of most sRNAs or RNA-binding proteins involved in RNA biology in pneumococcus. In this paper, we characterize the phenotypes and one target of the S1 domain RNA-binding protein CvfD, a homolog of general stress protein 13 identified, but not extensively characterized, in other Firmicutes species. Pneumococcal CvfD is a broadly pleiotropic regulator, whose absence results in misregulation of divalent cation homeostasis, reduced translation of the PhoU2 master regulator of phosphate uptake, altered metabolism and sRNA amounts, cold sensitivity, and attenuation of virulence. These findings underscore the critical roles of RNA biology in pneumococcal physiology and virulence.

Evaluation of Optimal Timing of Expert Feedback in a Simulated Flexible Ureteroscopy Course.

Introduction: Simulation-based training (SBT) has become an increasingly popular modality to train novice surgical residents in the face of rapidly increasing innovative surgical techniques across all surgical disciplines. Recent studies have already demonstrated SBT to be effective in helping overcome the learning curve associated with new surgical techniques, especially in junior residents and endoscopic procedures. In addition, it is known that trainees benefit significantly from expert feedback; however, there is a paucity of data looking into the optimal timing of this feedback during SBT. To address this knowledge deficit, an SBT curriculum was developed for junior urology residents to assess optimal timing of feedback during SBT for flexible ureteroscopy (fURS). Materials and Methods: The SBT course consisted of a pretraining assessment, three independent practice sessions, and a post-training assessment, with residents receiving expert feedback right after their pretraining assessment (early feedback [EF]) or after their final independent training session (late feedback [LF]). Results: Fifteen trainees with similar baseline fURS experience and precourse fURS task performance score participated in the study. There was a significant difference between the pre- and post-task completion times overall (15.2 minutes vs 9.1 minutes, p < 0.001), with no difference between the early or LF groups (p = 0.884). The mean performance scores improved for both groups (18.2 vs 24.2, p < 0.001) with the EF group having a more statistically significant improvement in performance scores than the LF group (p = 0.05), and most (73%) of residents preferred EF. Conclusions: This study demonstrates that an SBT curriculum for fURS is effective for technical skills development among junior trainees, and that EF resulted in marginally better overall scores and was preferred by residents.

Intraoperative Radiographic Determination of Ureteral Length as a Method of Determining Ideal Stent Length.

INTRODUCTION: Accurate determination of ureteral length (UL) and appropriate stent length remains a challenge. The objective of this study was to describe an intraoperative technique to measure UL and determine appropriate stent length, and to compare this technique with other methods of determining appropriate stent length. METHODS: Patients undergoing ureteroscopy requiring postoperative stenting and who had a preoperative CT were prospectively identified. Gender, age, height, body mass index, L1 to L5 lumbar height on CT, and surgeon's estimate of UL were recorded. UL was measured using four methods: direct measurement with a ureteral catheter, ureteropelvic junction (UPJ) to ureterovesical junction distance on axial and coronal CT, and using a novel intraoperative radiographic technique. Radiographic measurement was performed using a radiographic nipple marker affixed to the skin over the ureteral orifice (UO) and an angiographic catheter with radiopaque markings at 1 cm intervals. UL was the distance from the UPJ to the marker at the UO measured using the catheter markers. Correlation between direct measurement and the recorded variables and methods of ureteral measurement were calculated. Stent length was chosen based on radiographic measurement. Stents were deemed of appropriate length if they showed a proximal coil in the renal pelvis and a distal coil in the bladder without crossing midline. RESULTS: Twenty-five ureters from 23 patients were included. Radiographically measured UL was strongly correlated with direct measurement. (r = 0.873, p < 0.01). Coronal and axial CT ULs were significantly associated with direct measurement (p < 0.05). Height, lumbar height, and surgeon's estimate of UL were not. Stents were deemed of appropriate length in 23/25 cases (92%). CONCLUSIONS: This new method for radiographic UL measurement is strongly correlated with directly measured UL. A length of stent chosen based on radiographic UL resulted in an appropriate stent length.

Carbon Monoxide Photoproduction from Particles and Solutes in the Delaware Estuary under Contrasting Hydrological Conditions.

Full-spectrum, ultraviolet (UV), and visible broadband apparent quantum yields (AQYs) for carbon monoxide (CO) photoproduction from chromophoric dissolved organic matter (CDOM) and particulate organic matter (POM) were determined in the Delaware Estuary in two hydrologically contrasting seasons in 2012: an unusually low flow in August and a storm-driven high flow in November. Average AQYs for CDOM and POM in November were 10 and 16 times the corresponding AQYs in August. Maximum AQYs in November occurred in a midestuary particle absorption maximum zone. Although POM AQYs were generally smaller than CDOM AQYs, the ratio of the former to the latter increased substantially from the UV to the visible. In both seasons, UV solar radiation was the primary driver for CO photoproduction from CDOM whereas visible light was the principal contributor to POM-based CO photoproduction. CDOM dominated CO photoproduction in the uppermost water layer while POM prevailed at deeper depths. On a depth-integrated basis, the Delaware Estuary shifted from a CDOM-dominated system in August to a POM-dominated system in November with respect to CO photoproduction. This study reveals that flood events may enhance photochemical cycling of terrigenous organic matter and switch the primary photochemical driver from CDOM to POM.

Voyager observations of the interaction of the heliosphere with the interstellar medium.

This paper provides a brief review and update on the Voyager observations of the interaction of the heliosphere with the interstellar medium. Voyager has found many surprises: (1) a new energetic particle component which is accelerated at the termination shock (TS) and leaks into the outer heliosphere forming a foreshock region; (2) a termination shock which is modulated by energetic particles and which transfers most of the solar wind flow energy to the pickup ions (not the thermal ions); (3) the heliosphere is asymmetric; (4) the TS does not accelerate anomalous cosmic rays at the Voyager locations; and (5) the plasma flow in the Voyagers 1 (V1) and 2 (V2) directions are very different. At V1 the flow was small after the TS and has recently slowed to near zero, whereas at V2 the speed has remained constant while the flow direction has turned tailward. V1 may have entered an extended boundary region in front of the heliopause (HP) in 2010 in which the plasma flow speeds are near zero.

Personality traits and career satisfaction of health care professionals.

Based on Holland's theorizing that vocational satisfaction arises from a good match between one's personality and career choice, one purpose of the study was to examine broad and narrow personality traits that characterize health care workers in comparison with professionals from other occupations. Also investigated were ways in which characteristic traits of health care workers were related to career satisfaction. Professionals utilizing the services of eCareerfit.com responded to online surveys that have been demonstrated to produce reliable and valid measures of broad and narrow personality traits and levels of career satisfaction. An independent sample t test was used to compare means of health care workers with those from other occupations. Pearson product-moment correlations were then computed to assess relationships between the traits and career satisfaction of health care professionals. Two traits that were particularly strong among health care workers were also significantly correlated with career satisfaction: work drive and conscientiousness. Other traits were found to be significantly related to career satisfaction in health care but were not uniquely high in the sample of health care professionals. To increase career satisfaction of health care professionals and thus to improve retention rates, administrators should consider focusing on recruiting and selecting individuals with higher levels of key personality traits.

Cool heliosheath plasma and deceleration of the upstream solar wind at the termination shock.

The solar wind blows outward from the Sun and forms a bubble of solar material in the interstellar medium. The termination shock occurs where the solar wind changes from being supersonic (with respect to the surrounding interstellar medium) to being subsonic. The shock was crossed by Voyager 1 at a heliocentric radius of 94 au (1 au is the Earth-Sun distance) in December 2004 (refs 1-3). The Voyager 2 plasma experiment observed a decrease in solar wind speed commencing on about 9 June 2007, which culminated in several crossings of the termination shock between 30 August and 1 September 2007 (refs 4-7). Since then, Voyager 2 has remained in the heliosheath, the region of shocked solar wind. Here we report observations of plasma at and near the termination shock and in the heliosheath. The heliosphere is asymmetric, pushed inward in the Voyager 2 direction relative to the Voyager 1 direction. The termination shock is a weak, quasi-perpendicular shock that heats the thermal plasma very little. An unexpected finding is that the flow is still supersonic with respect to the thermal ions downstream of the termination shock. Most of the solar wind energy is transferred to the pickup ions or other energetic particles both upstream of and at the termination shock.

The laparoscopic experience of recently trained Canadian urologists.

OBJECTIVE: While laparoscopy represents an increasingly important aspect of operative urology, the experience of Canadian urology trainees is poorly defined. The purpose of this study was to determine the level of laparoscopic training of Canadian urology trainees during residency as well as their comfort level with various laparoscopic techniques. METHODS: An e-mail-and web-based questionnaire was administered to the two most recent cohorts of Canadian-trained urologists (residencies completed in 2003 or 2004). A total of 50 questionnaires were sent. Overall laparoscopic experience and experience with specific laparoscopic tasks (e.g. colonic mobilization) were assessed. Subjects also reported on their relative comfort level (Likert scale of 1 to 5) with various laparoscopic procedures and tasks. Finally, attitudes towards the future role of laparoscopy in the practice of urology were determined. RESULTS: Thirty-six individuals (72%) responded to the questionnaire. Of the respondents, 28 (78%) were performing fellowships; of those pursuing fellowship training, 13 (36%) involved laparoscopy. Thirty-five respondents (97%) had received some laparoscopic exposure during residency; 29 (81%) had mobilized the colon, spleen or liver and 27 (75%) had dissected the renal vessels. Only 7 out of 33 respondents (21%) felt that their residency adequately prepared them to perform laparoscopy independently. There were no significant differences between the responses of the two cohorts. CONCLUSION: Most Canadian urology trainees are being exposed to laparoscopy, but more exposure is required in order to perform laparoscopy in practice. Many residents still require fellowship training to become facile in laparoscopy.