Concordance analysis of microbial genomes

The set of proteins which are conserved across families of microbes contain important targets of new anti-microbial agents. We have developed a simple and efficient computational tool which determines concordances of putative gene products that show sets of proteins conserved across one set of user specified genomes and not present in another set of user specified genomes. The thresholds and the homology scoring criterion are selectable to allow the user to decide the stringency of the homologies. The system uses a relational database to store protein coding regions from different genomes, and to store the results of a complete comparison of all sequences against all sequences using the FASTA program. Using Web technology, the display of all the related proteins for a given sequence and calculation of multiple sequence alignments (using CLUSTALW) can be performed with the click of a button. The current database holds 97 365 sequences from 19 complete or partial genomes and 8798905 FASTA comparison results. A example concordance is presented which demonstrates that the target of the quinolone antibiotics could have been identified using this tool.     

Specific immune induction following DNA-based immunization through in vivo transfection and activation of macrophages/antigen-presenting cells

The initiation of an adaptive immune response requires Ag presentation in combination with the appropriate activation signals. Classically, Ag presentation and immune activation occur in the lymph node and spleen, where a favorable organ architecture and rich cellular help can enhance the process. Recently, several investigators have reported the use of DNA expression cassettes to elicit cellular and humoral immunity against diverse pathogens. Although the immune mechanisms involved are still poorly understood, plasmid inoculation represents a model system for studying immune function in response to invading pathogens. In this report, we demonstrate the presence of activated macrophages or dendritic cells in the blood lymphocyte pool and peripheral tissues of animals inoculated with DNA expression cassettes. These cells are directly transfected in vivo, present Ag, and display the surface proteins CD80 and CD86. Our studies indicate that these cells function as APC and can activate naive T lymphocytes. They may represent an important first step APC in genetic immunization and natural infection.     

Rapid-fractionation preoperative chemoradiation, pancreaticoduodenectomy, and intraoperative radiation therapy for resectable pancreatic adenocarcinoma

PURPOSE: To evaluate the toxicities, radiographic and pathologic responses, and event-free outcomes with combined modality treatment that involves preoperative rapid-fractionation chemoradiation, pancreaticoduodenectomy, and electron-beam intraoperative radiation therapy (EB-IORT) for patients with resectable pancreatic adenocarcinoma. PATIENTS AND METHODS: Patients with radiographically resectable localized adenocarcinoma of the pancreatic head were entered onto a preoperative protocol that consisted of a 2-week course of fluorouracil (5-FU) 300 mg/m2 daily 5 days per week and concomitant rapid-fractionation radiation 30 Gy, 3 Gy daily 5 days per week. Radiographic restaging was performed 4 weeks after chemoradiation, and patients with localized disease underwent pancreaticoduodenectomy with EB-IORT 10 to 15 Gy. RESULTS: Thirty-five patients were entered onto the study and completed chemoradiation, 34 (97%) as outpatients. Three patients (9%) experienced grade 3 nausea and vomiting; no other grade 3 or 4 toxicities were observed. Of the 27 patients taken to surgery, 20 patients (74%) underwent pancreaticoduodenectomy with EB-IORT. All patients had a less than grade III pathologic response to preoperative chemoradiation. At a median follow-up of 37 months, the 3-year survival rate in patients who underwent combined modality therapy was 23%. CONCLUSION: Combined modality treatment with preoperative rapid-fractionation chemoradiation, pancreaticoduodenectomy, and EB-IORT is associated with minimal toxicity and excellent locoregional control. This represents one approach to maximize the proportion of patients who receive all components of combined modality therapy and avoids the toxicity of pancreaticoduodenectomy in patients found to have metastatic disease at the time of restaging.     

Assessment of regional left ventricular long-axis motion with MR velocity mapping in healthy subjects

We present a case of trichothiodystrophy associated with a hypereosinophilic syndrome. This case had been followed for 30 years. Trichothiodystrophy was characterized by lamellar ichthyosis, hair and nail dystrophies, kyphoscoliosis, congenital luxation of the hip. The hypereosinophilic syndrome was characterized by an itching urticaria-like eruption. Although the patient's general condition had remaining stable for 30 years, during the last year it worsened and the patient suddenly died. The authors discuss about the significance of this association.     

Enhanced T-cell immunogenicity and protective efficacy of a human immunodeficiency virus type 1 vaccine regimen consisting of consecutive priming with DNA and boosting with recombinant fowlpox virus

The induction of human immunodeficiency virus (HIV)-specific T-cell responses is widely seen as critical to the development of effective immunity to HIV type 1 (HIV-1). Plasmid DNA and recombinant fowlpox virus (rFPV) vaccines are among the most promising safe HIV-1 vaccine candidates. However, the immunity induced by either vaccine alone may be insufficient to provide durable protection against HIV-1 infection. We evaluated a consecutive immunization strategy involving priming with DNA and boosting with rFPV vaccines encoding common HIV-1 antigens. In mice, this approach induced greater HIV-1-specific immunity than either vector alone and protected mice from challenge with a recombinant vaccinia virus expressing HIV-1 antigens. In macaques, a dramatic boosting effect on DNA vaccine-primed HIV-1-specific helper and cytotoxic T-lymphocyte responses, but a decline in HIV-1 antibody titers, was observed following rFPV immunization. The vaccine regimen protected macaques from an intravenous HIV-1 challenge, with the resistance most likely mediated by T-cell responses. These studies suggest a safe strategy for the enhanced generation of T-cell-mediated protective immunity to HIV-1.     

Hypertonicity regulates the function of human neutrophils by modulating chemoattractant receptor signaling and activating mitogen-activated protein kinase p38

Excessive neutrophil activation causes posttraumatic complications, which may be reduced with hypertonic saline (HS) resuscitation. We tested if this is because of modulated neutrophil function by HS. Clinically relevant hypertonicity (10-25 mM) suppressed degranulation and superoxide formation in response to fMLP and blocked the activation of the mitogen activated protein kinases (MAPK) ERK1/2 and p38, but did not affect Ca2+ mobilization. HS did not suppress oxidative burst in response to phorbol myristate acetate (PMA). This indicates that HS suppresses neutrophil function by intercepting signal pathways upstream of or apart from PKC. HS activated p38 by itself and enhanced degranulation in response to PKC activation. This enhancement was reduced by inhibition of p38 with SB203580, suggesting that p38 up-regulation participates in HS-induced enhancements of degranulation. HS had similar effects on the degranulation of cells that were previously stimulated with fMLP, but had no effect on its own, suggesting that HS enhancement of degranulation requires another signal. We conclude that depending on other stimuli, HS can suppress neutrophil activation by intercepting multiple receptor signals or augment degranulation by enhancing p38 signaling. In patients HS resuscitation may reduce posttraumatic complications by preventing neutrophil activation via chemotactic factors released during reperfusion.     

Participation of phosphoinositide-specific phospholipase Cgamma1 and STAT1 protein in the formation of latent complexes of signal proteins

It is known that the growth factor activates appropriate membrane receptors which become starting points of cascades of protein-protein interactions leading to cellular response. Recent data suggest that different signalling pathways may cross-talk during the cellular response. Here we show that phosphoinositide-specific phospholipase C gamma 1, one of the key elements in phosphoinositide pathway of signal transduction, is physically associated with members of the STAT pathway. The precipitation of phospholipase C gamma 1, using polyclonal antibody in A-431 cells, leads to co-immunoprecipitation of STAT1 alpha and STAT1 beta, as well as STAT3. The formation of such complexes was observed in both unstimulated and EGF stimulated cells. The participation of SH3-domains in the formation of such complexes is discussed.     

Fitness landscapes for effects of shape on chemotaxis and other behaviors of bacteria

Data on the shapes of 218 genera of free-floating or free-swimming bacteria reveal groupings around spherical shapes and around rod-like shapes of axial ratio about 3. Motile genera are less likely to be spherical and have larger axial ratios than nonmotile genera. The effects of shape on seven possible components of biological fitness were determined, and actual fitness landscapes in phenotype space are presented. Ellipsoidal shapes were used as models, since their hydrodynamic drag coefficients can be rigorously calculated in the world of low Reynolds number, where bacteria live. Comparing various shapes of the same volume, and assuming that departures from spherical have a cost that varies with the minimum radius of curvature, led to the following conclusions. Spherical shapes have the largest random dispersal by Brownian motion. Increased surface area occurs in oblate ellipsoids (disk-like), which rarely occur. Elongation into prolate ellipsoids (rod-like) reduces sinking speed, and this may explain why some nonmotile genera are rod-like. Elongation also favors swimming efficiency (to a limited extent) and the ability to detect stimulus gradients by any of three mechanisms. By far the largest effect (several hundred-fold) is on temporal detection of stimulus gradients, and this explains why rod-like shapes and this mechanism of chemotaxis are common.     

Hearing loss in community-dwelling older persons: national prevalence data and identification using simple questions

OBJECTIVE: To estimate the prevalence of hearing loss among community-dwelling older persons according to clinical criteria and to develop a brief self-report screening instrument to detect hearing loss. DESIGN: Survey. SETTING: National probability sample of noninstitutionalized older persons. PARTICIPANTS: A total of 2506 persons aged 55 to 74 who participated in the National Health and Nutrition Examination Survey. MAIN OUTCOME MEASURES: Hearing loss as defined by Ventry and Weinstein (VW) criteria and by the High Frequency Pure-Tone Average (HFPTA) scale. RESULTS: Hearing loss by VW criteria was present in 14.2% and by HFPTA criteria in 35.1% of those surveyed. The prevalence increased with advancing age and was higher among men and those with less education. A logistic regression model identified six independent factors for hearing loss by VW criteria: age > or = 70 years (adjusted odds-ratio (AOR) 2.7, 95% confidence interval (95% CI) 1.6, 4.4), male gender (AOR 3.0, 95% CI 1.9, 4.8), < or = 12th grade education (AOR 3.8, 95% CI 1.8, 7.7), having seen a doctor for deafness or hearing loss (AOR 8.9, 95% CI 5.3, 14.9), unable to hear a whisper across a room (AOR 3.2, 95% CI 2.0, 5.1), and unable to hear a normal voice across a room (AOR 6.2, 95% CI 2.6, 14.9). A clinical scale based on the logistic model had 80% sensitivity and 80% specificity in predicting hearing loss using VW criteria and 59% sensitivity and 88% specificity in predicting hearing loss using HFPTA criteria. CONCLUSIONS: Hearing loss, as defined by two clinical criteria, is common and can be screened for accurately using simple questions that assess sociodemographic and hearing-related characteristics.