Analysis of Staphylococcus aureus Transcriptome in Pediatric Soft Tissue Abscesses and Comparison to Murine Infections.

A comprehensive understanding of how Staphylococcus aureus adapts to cause infections in humans can inform development of diagnostic, therapeutic, and preventive approaches. Expression analysis of clinical strain libraries depicts in vitro conditions that differ from those in human infection, but low bacterial burden and the requirement for reverse transcription or nucleic acid amplification complicate such analyses of bacteria causing human infection. We developed methods to evaluate the mRNA transcript signature of S. aureus in pediatric skin and soft tissue infections (SSTI) directly ex vivo Abscess drainage from 47 healthy pediatric patients undergoing drainage of a soft tissue infection was collected, and RNA was extracted from samples from patients with microbiologically confirmed S. aureus abscesses (42% due to methicillin-resistant S. aureus [MRSA]). Using the NanoString platform and primers targeting S. aureus mRNA transcripts encoding surface-expressed or secreted proteins, we measured direct counts of 188 S. aureus mRNA transcripts in abscess drainage. We further evaluated this mRNA signature in murine models of S. aureus SSTI and nasal colonization where the kinetics of the transcriptome could be determined. Heat maps of the S. aureus mRNA signatures from pediatric abscesses demonstrated consistent per-target expression across patients. While there was significant overlap with the profiles from murine SSTI and nasal colonization, important differences were noted, which can inform efforts to develop therapeutic and vaccine approaches.

T(H)17-Mediated Protection against Pneumococcal Carriage by a Whole-Cell Vaccine Is Dependent on Toll-Like Receptor 2 and Surface Lipoproteins.

A pneumococcal whole-cell vaccine (WCV) confers T(H)17-mediated immunogenicity and reduces nasopharyngeal (NP) carriage in mice. Activation of Toll-like receptor 2 (TLR2) has been shown to be important for generating T(H)17 responses, and several lipidated pneumococcal proteins have TLR2-activating properties. Here we investigated the roles of TLR2 and lipidation of proteins in WCV-induced interleukin-17A (IL-17A) responses and protection against NP carriage. Immunization of Tlr2(-/-) mice with WCV conferred significantly lower IL-17A levels and reduced protection against NP carriage, compared to wild-type (WT) mice, suggesting that host TLR2 engagement is required for effective immunity and protection elicited by WCV immunization. Using a WCV with deletion of lgt, the gene encoding the enzyme required for lipidation and membrane attachment of prolipoproteins, we show that lipidation and membrane localization of these proteins are critical for the immunogenicity and protective efficacy of the WCV. To evaluate the roles of diacylglyceryl transferase (Lgt)-mediated processes in the recall of WCV-induced protective responses, we colonized WCV-immunized animals with a strain in which lgt was deleted. WCV-immunized animals still had significantly reduced colonization burdens, compared to control animals, which suggests that lipidation and membrane localization of pneumococcal prolipoproteins are less critical for the recall of the immune responses elicited by WCV immunization than for the priming of such responses. Elucidation of underlying immune mechanisms and the optimal characteristics of WCV formulations can help guide vaccine development and enhance our understanding of host-pneumococcus interactions.

Inflammatory and immunological biomarkers are not related to survival in adults with Cystic Fibrosis.

BACKGROUND: Chronic Pseudomonas aeruginosa pulmonary infection is associated with a decline in lung function and reduced survival in people with Cystic Fibrosis (CF). Damaging inflammatory and immunological mediators released in the lungs can be used as markers of chronic infection, inflammation and lung tissue damage. METHODS: Clinical samples were collected from CF patients and healthy controls. Serum IgG and IgA anti-Pseudomonas antibodies, sputum IL-8 and TNFα, plasma IL-6 and urine TNFr1 were measured by ELISA. Sputum neutrophil elastase (NE), cathepsin S and cathepsin B were measured by spectrophotometric and fluorogenic assays. The relationship between IgG and IgA, inflammatory mediators and long-term survival was determined. RESULTS: IgG and IL-6 positively correlated with mortality. However, multivariate analysis demonstrated that after adjusting for FEV(1), IgG was not independently related to mortality. A relationship was observed between IgG and IL-6, TNFα, TNFr1 and between IgA and IL8, cathepsin S and cathepsin B. CONCLUSIONS: These data indicate that biomarkers of inflammation are not independent predictors of survival in people with CF.

Chymotrypsin-like serine proteinases are involved in the maintenance of cell viability.

An increasing number of studies have implicated serine proteinases in the development of apoptosis. In this study, we assessed the ability of a set of highly specific irreversible inhibitors (activity probes), incorporating an α-amino alkane diphenyl phosphonate moiety, to modulate cell death. In an initial assessment of the cellular toxicity of these activity probes, we discovered that one example, N-α-tetramethylrhodamine phenylalanine diphenylphosphonate {TMR-Phe(P)(OPh)(2)} caused a concentration-dependent decrease in the viability of HeLa and U251 mg cells. This reduced cell viability was associated with a time-dependent increase in caspase-3 activity, PARP cleavage and phosphatidylserine translocation, establishing apoptosis as the mechanism of cell death. SDS-PAGE analysis of cell lysates prepared from the HeLa cells treated with TMR-Phe(P)(OPh)(2), revealed the presence of a fluorescent band of molecular weight 58 kDa. Given that we have previously reported on the use of this type of activity probe to reveal active proteolytic species, we believe that we have identified a chymotrypsin-like serine proteinase activity integral to the maintenance of cell viability.

Association of airway cathepsin B and S with inflammation in cystic fibrosis.

SUMMARY: Irreversible tissue damage within the cystic fibrosis (CF) lung is mediated by proteolytic enzymes during an inflammatory response. Serine proteinases, in particular neutrophil elastase (NE), have been implicated however, members of the cysteine proteinase family may also be involved. The aim of this study was to determine cathepsin B and S levels in cystic fibrosis (CF) sputum and to assess any relationship to recognized markers of inflammation such as sputum NE, interleukin-8 (IL-8), tumor necrosis factor alpha (TNF-alpha), urine TNF receptor 1 (TNFr1), plasma IL-6, and serum C-reactive protein (CRP). Proteinase activities were measured in the sputum of 36 clinically stable CF patients using spectrophotometric and fluorogenic assays. Immunoblots were also used to confirm enzyme activity data. All other parameters were measured by ELISA. Patients had a mean age of 27.2 (8.2) years, FEV. of 1.6 (0.79) L and BMI of 20.7 (2.8). Both cathepsin B and S activities were detected in all samples, with mean concentrations of 18.0 (13.5) microg/ml and 1.6 (0.88) microg/ml, respectively and were found to correlate not only with each other but with NE, TNF-alpha and IL-8 (in all cases . < 0.05). Airway cathepsin B further correlated with circulatory IL-6 and CRP however, no relationship for either cathepsin was observed with urine TNFr1. This data indicates that cathepsin B and S may have important roles in the pathophysiology of CF lung disease and could have potential as markers of inflammation in future studies.

The emerging role of serine proteases in apoptosis.

Unregulated apoptosis can be due to a disruption in the balance and control of both intra- and inter-cellular proteolytic activities leading to various disease states. Many proteases involved in apoptotic processes are yet to be identified; however, several are already well characterized. Caspases traditionally held the predominant role as prime mediators of execution. However, latterly, evidence has accumulated that non-caspases, including calpains, cathepsins, granzymes and the proteasome have roles in mediating and promoting cell death. Increasingly, research is implicating serine proteases within apoptotic processing, particularly in the generation of nuclear events such as condensation, fragmentation and DNA degradation observed in late-stage apoptosis. Serine proteases therefore are emerging as providing additional or alternative therapeutic targets.

Inflammatory markers in cystic fibrosis patients with transmissible Pseudomonas aeruginosa.

Chronic Pseudomonas aeruginosa infection in cystic fibrosis (CF) leads to a damaging host inflammatory response. There are an increasing number of reports of P. aeruginosa cross-infection at CF centres. The clinical significance of acquisition of a transmissible strain for patients who already harbour P. aeruginosa is unclear. In this study, levels of inflammatory markers in clinically stable adult CF patients who harbour transmissible and sporadic strains of P. aeruginosa have been compared. Patients with CF and chronic P. aeruginosa infection were grouped into those who harbour a transmissible P. aeruginosa and those who harbour their own sporadic strains. Total white cell and differential counts, sputum neutrophil elastase (NE), interleukin (IL)-8, tumour necrosis factor (TNF)-alpha, plasma IL-6 and NE/alpha1-antitrypsin complexes, serum C-reactive protein, and urine TNF receptor 1 were all measured in clinically stable patients 4-6 weeks following completion of intravenous antibiotic therapy. The two groups (both n=20) were well matched for per cent predicted forced expiratory volume in one second, per cent predicted forced vital capacity and body mass index. There were no significant differences in levels of white cell counts or inflammatory markers between the two groups. At times of clinical stability, cystic fibrosis patients infected with transmissible Pseudomonas aeruginosa do not have a heightened inflammatory response above that of those harbouring sporadic strains.

Expectancy and suggestibility: are the effects of environmental enhancement due to detection?

This study replicated the effect of Wickless and Kirsch's experiential expectancy manipulation, in which lights and music from hidden sources were used to convince participants that they were responding successfully to suggestions for visual and auditory hallucinations. The hypothesis that the effect is mediated by detection of the manipulation was tested by providing some participants with cues that their experiences were due to actual changes in the physical environment rather than to their responses to suggestion. This hypothesis was not confirmed. A significant effect on suggestibility was obtained only among participants not given cues aimed at enabling detection of the manipulation, and among those provided with the cues, suspicion of the manipulation was negatively correlated with response to suggestion.

Stereoscopic head-mounted display incorporated into microsurgical procedures: technical note.

OBJECTIVE: We have previously evaluated video technologies that have allowed for the use of electronic imaging during microneurosurgical and endoscopic procedures. This stereoscopic camera set and monitor/recording system allows for the use of stereoscopic images during surgery by the primary surgical assistant and ancillary personnel. It also allows for stereoscopic recording and playback using a wide video home system format. We describe a novel prototype of a head-mounted display (HMD) that allows the surgical team to simultaneously visualize the surgical field stereoscopically and includes picture-in-picture, voice control, and stereoscopic recording capabilities. INSTRUMENTATION: A stereoscopic HMD with 640 x 480 (video graphics array) pixel resolution and the ability to display 24-bit images has been designed. This device weighs 900 g. It is interfaceable with common video display formats. RESULTS: This stereoscopic HMD is being evaluated in a prospective multicenter trial of open microsurgical and endoscopic minimally invasive procedures. In our experience to date, there have been no equipment failures or complications attributable to the use of the display system. The equipment was well accepted by users who reported significant benefits in visualization. CONCLUSION: The use of a stereoscopic HMD may result in improved efficiency and safety in both endoscopic and open microsurgical procedures. We have verified that the HMD is comfortable during the course of a surgical procedure, is reliable, and allows for accessibility to the operative field with an excellent field of view and three-dimensional perception. Positioning and dexterity within the operative field are also enhanced. Additional uses relate to surgical training, multimodal information display, and operative rehearsals.

Head-mounted display system for microneurosurgery.

Advances in video technology have enabled the incorporation of high-resolution electronic imaging of an operative field during microneurosurgical and endoscopic procedures. While current video-based monoscopic camera and monitor systems produce an excellent image, the third dimension is lacking and the operative field and image are at different locations. We describe a head-mounted display (HMD) that allows the surgical team to simultaneously visualize a surgical field and a high-resolution magnified stereoscopic image which includes a picture-in-picture capability. Stereoscopic HMDs should result in improved efficiency and safety in both endoscopic and open microsurgical procedures. Additional uses relate to surgical training and operative rehearsals.

Reporting biases in hypnosis: suggestion or compliance?

The tendency of highly hypnotizable participants to bias their retrospective perceptual reports in response to instructional demands was reexamined with the addition of low-hypnotizable control participants instructed to simulate hypnosis. Mean scores of high-hypnotizable participants and simulators did not differ, but the responses of simulators to the demand instruction was less variable than those of high-hypnotizable participants, and the shape of the response distribution was different. Unlike simulators, some high-hypnotizable participants who had reported changes in perception that were consistent with a hypnotic suggestion subsequently reported changes opposite to those suggested by a demand instruction. These data were interpreted as suggesting that the responses of high-hypnotizable participants to both the demand instruction and the preceding hypnotic suggestion were not entirely due to compliance.

Depression and memory narrative type.

Research with autobiographical memories has distinguished between memory narratives of specific events and summaries of many events blended together. Depression has been associated with a reduced ability to retrieve and orally relate specific positive memories. This study explored the hedonic bias in memory through collection of written autobiographical memories from 90 nonclinical college students whose mood was assessed for depression. Participants with higher depression scores recalled significantly more summary memories in response to a request for a positive self-defining memory than did participants with lower depression scores. There were no significant differences in the number of single-event and summary memories when participants were asked for a negative memory. We used J. A. Singer and K. H. Moffitt's (1991-1992) scoring system to distinguish between summarized and specific memory narratives.

Focus group interview with parents of children with medically complex needs: an intimate look at their perceptions and feelings.

The purpose of this paper was to identify the needs of parents of children with medically complex needs from their own perception. In order to provide in-depth information, the focus group interview technique was used. Several strong recurrent themes were identified. The most persistent need was for a general organization or framework with which the care providers could operate. Along these same lines, the fragmentation of training, needs and services was consistently stated. A general lack of information in terms of home care and how to plan for the future was identified. Support groups were universally lauded for the invaluable services provided to the care parents.

Cardiovascular mortality trends in Harris County, Texas: 1980 to 1986.

Cardiovascular diseases are the leading causes of death in Texas and in the United States. This study determines the trend in mortality rates attributed to cardiovascular diseases in Harris County from 1980 to 1986. The region of the county that does not include the City of Houston was specifically studied. Mortality of cardiovascular diseases in these two areas follow patterns similar to that of the United States in the same time period. Both the entire county and Harris County excluding Houston show declines in cardiovascular mortality rates in the 7-year period. The populations were divided into four ethnic categories (white, black, Hispanic and "other"), and each ethnic group reported significant declines in overall cardiovascular mortality except in the "other" population, which showed an increase in the male group. Significant downward trends were noticed in the white and Hispanic population in the two major subcategories of cardiovascular diseases: diseases of the heart and cerebrovascular diseases. The black population in each geographic area studied was consistently higher in cardiovascular mortality than the other three ethnic groups observed (white, Hispanic and "other"). Knowledge of cardiovascular disease mortality rates by ethnicity, sex, and age as well as temporal changes in mortality rates within Harris County are important for health planners in continuing and implementing programs aimed at awareness, prevention, and treatment of cardiovascular diseases.

Chromatic aftereffects associated with a night vision goggle simulation.

A visual perception experiment was conducted to determine the chromatic aftereffects of viewing a yellow-green field that simulated the display of current night vision goggles. Six females and two males served as subjects in a color-naming procedure. Subjects sequentially viewed an adaptation field, which was either yellow-green or white, and small colored targets presented on a CRT display. The time required to name the color of the targets was found to be dependent on the color of the adaptation field, the color of the target, and the interaction of these two variables. It was recommended that the effects of attenuation of the luminance of the night vision goggles be studied, and that color cockpit displays be redundantly coded whenever possible.

New approaches to the management of acute puerperal uterine inversion.

Six cases of acute puerperal inversion of the uterus are reported. In four of these, tocolytic agents were used to facilitate replacement of the inverted uterus; and in five, Prostin-15M was given after replacement. The use of tocolytics may obviate the need for general anesthesia for uterine replacement in some patients with uterine inversion. Prostin-15M serves to minimize bleeding and to maintain uterine position after replacement.