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Pulmonary arterial hypertension (PAH) is a multifactorial and chronic disease characterized by an increase in pulmonary artery (PA) resistance leading to right ventricle (RV) failure. Endothelial dysfunction and alteration NO/cGMP signaling in PA play a major role in PAH. We described the involvement of Rac1 in the control of systemic blood pressure through its involvement in NO-mediated relaxation of arterial smooth muscle cell (SMC). The aim of this study was to decipher the role of SMC Rac1 in PAH. Rac1 activation, Rac-GTP, is determined by immunofluorescence in lungs biopsies from control and PAH patients. PAH is induced by exposure of wild-type (WT) and SMC Rac1-deficient mice (SM-Rac1-KO mice) to chronic hypoxia (10% O 2 , 4 weeks). PAH is assessed by the measurement of RV systolic pressure and hypertrophy. The proliferation of pulmonary arterial SMC (PASMC) in hypoxic condition is measured by EdU staining. Oxidative stress is assessed by immunofluorescence using the DHE probe and by electron spin resonance (ESR). In biopsies from PAH patients, we detected an increase (3-fold) of Rac-GTP in PA compared to control (n = 4–6). To decipher the role of Rac activation in PAH, WT and SM-Rac1-KO mice were exposed to hypoxia to induce PAH. The elevation of systolic pressure in RV and RV hypertrophy were decreased by 32.5% and 60% (n = 5–7) in SM-Rac1-KO mice. Deletion of Rac1 in SMC reduced the hypoxia-induced remodeling. In vitro, we observed that hypoxia-induced PASMC proliferation is decreased by 50% by the Rac1 inhibitor, EHT1864. We demonstrated that Rac1 inhibition or deletion in SMC prevent ROS production in PASMC and limit PASMC proliferation and vascular remodeling. Our results suggest that Rac1 activity in PASMC plays a causal role in PAH by favoring ROS dependent PA remodelling induced by chronic hypoxia. Therefore, Rac1 deletion or inhibition protect against hypoxic PAH. [ABSTRACT FROM AUTHOR]

s. perelle, f. dilasser, j. grout and p. fach. 2005.The aims of the study were to identify the specific genes of O-antigen gene cluster from Shiga toxin-producingEscherichia coli(STEC) O103 and to provide the basis for a specific real-time PCR test for rapid detection ofE. coliO103.The published primers complementary to JUMPstart andgndgene, the conserved flanking sequences of O-antigen genes clusters inE. coliand related species, were used to amplify the 12-kbp O103 O-antigen biosynthesis locus of STEC O103. A DNA library representative of this cluster allowed two O103-specific probes to be identified in the flippase (wzx) and UDP-galactose-4-epimerase (galE) genes. Two specific O103 serotyping real-time PCR tests based on these two genes were successfully developed.These results confirm that the O-antigen gene cluster sequences ofE. coliallow rapidly a specific O-antigen real-time PCR assay to be designed.These findings increase the number of real-time PCR-assays available to replace the classical O-serotyping amongE. coliO-antigen. [ABSTRACT FROM AUTHOR]

Aims: This paper provides identification of a DNA sequence derived from Shiga toxin-producing Escherichia coli (STEC) O157:H7 and information on its utilization for detecting STEC O157 by PCR. Methods and Results: Random Amplified Polymorphic DNA and DNA library were used to identify in STEC O157:H7 (strain EDL 933) a 2634-bp S mall I nserted L ocus, designated SILO157 . Analysis of 211 bacterial strains showed that the PCR assays amplifying the SILO157 region could be used to detect STEC O157 with a good specificity. Conclusions: Characterization of a novel locus in STEC O157 is attractive since the serotype O157:H7 of STEC is still by far the most important serotype associated with more serious diseases. This island encodes putative proteins and especially one that is predicted to be an outer membrane protein designated IHP1. Significance and Impact of the Study: Further investigations could now be developed to appreciate the role of the SILO157 in pathogenicity. [ABSTRACT FROM AUTHOR]

Abstract Background Enterobacter sakazakii is the causative agent of rare but severe food-borne infections associated with meningitis, necrotizing enterocolitis and sepsis in infants. Rehydrated powdered infant formulae have been implicated as the source of infection in several outbreaks and sporadic cases. In this work, a real time fluorescence resonance energy transfer PCR assay incorporating an internal amplification control (IAC) was developed for the specific detection of E. sakazakii in foods. Performance of the assay, coupled to an automated DNA extraction system and the E. sakazakii ISO-IDF (TS 22964/RM 210) enrichment procedure, was evaluated on infant formulae and samples from production environment. Results The real-time PCR assay had 100% specificity as assessed using 35 E. sakazakii and 184 non-E. sakazakii strains. According to the E. sakazakii strains tested, the detection limits ranged from 5 to 25 genomic copies. Assays on pure cultures (including real-time PCR and DNA extraction) gave a sensitivity of about 102 to 103 CFU/ml. Out of 41 naturally contaminated infant formulae and environmental samples analysed for the presence of E. sakazakii, 23 were positive by real-time PCR and 22 by the conventional culture method, giving 97.5% concordance with the ISO-IDF reference method. Conclusion This method, combining specific real-time PCR, automated DNA extraction and ISO-IDF standard enrichments, provides a useful tool for rapid screening of E. sakazakii in food and environmental matrices.

Aims: This paper provides identification of a DNA sequence derived from Shiga toxin-producing Escherichia coli (STEC) O157:H7 and information on its utilization for detecting STEC O157 by PCR. Methods and Results: Random Amplified Polymorphic DNA and DNA library were used to identify in STEC O157:H7 (strain EDL 933) a 2634-bp S mall I nserted L ocus, designated SILO157 . Analysis of 211 bacterial strains showed that the PCR assays amplifying the SILO157 region could be used to detect STEC O157 with a good specificity. Conclusions: Characterization of a novel locus in STEC O157 is attractive since the serotype O157:H7 of STEC is still by far the most important serotype associated with more serious diseases. This island encodes putative proteins and especially one that is predicted to be an outer membrane protein designated IHP1. Significance and Impact of the Study: Further investigations could now be developed to appreciate the role of the SILO157 in pathogenicity. [ABSTRACT FROM AUTHOR]