A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

Résistance aux antibiotiques chez les pneumocoques.

In 1875, 7 years prior to the description of the Koch bacillus, Klebs visualized the first Streptococcus pneumoniae in pleural fluid. Since then, this organism has played a decisive role in biomedical science. From a biological point of view, it was extensively involved in the development of passive and active immunization by serotherapy and vaccination respectively. Genetic transformation was also first observed in S. pneumoniae, leading to the discovery of DNA. From a clinical point of view, S. pneumoniae is today still a prime cause of otitis media in children and of pneumonia in all age groups, as well as a predominant cause of meningitis and bacteremia. In adults, bacteremia still has a mortality of over 25%. Although S. pneumoniae remained very sensitive to penicillin for many years, penicillin-resistant strains have emerged and increased dramatically over the last 15 years. During this period the frequency of penicillin-resistant isolates has increased from < or = 1% to frequencies varying from 20 to 60% in geographic areas as diverse as South Africa, Spain, France, Hungary, Iceland, Alaska, and numerous regions of the United States and South America. In Switzerland, the current frequency of penicillin-resistant pneumococci ranges between 5 and > or = 10%. The increase in penicillin-resistant pneumococci correlates with the intensive use of beta-lactam antibiotics. The mechanism of resistance is not due to bacterial production of penicillinase but to an alteration of the bacterial target of penicillin, the so-called penicillin-binding proteins. Resistance is subdivided into (1) intermediate level resistance (minimal inhibitory concentration [MIC] of penicillin of 0.1-1 mg/l) and (2) high level resistance (MCI > or = 2 mg/l). The clinical significance of intermediate resistance remains poorly defined. On the other hand, highly resistant strains have been responsible for numerous therapeutic failures, especially in cases of meningitis. Antibiotics recommended against penicillin-resistant pneumococci include cefotaxime, ceftriaxone, imipenem and in some instances vancomycin. However, penicillin-resistant pneumococci tend to present cross-resistances to all the antibiotics of the beta-lactam family and could even become resistant to the last resort drugs mentioned above. Thus, the explosion of resistance to penicillin in pneumococci is a ubiquitous phenomenon which must be fought against by (1) avoiding excessive use of antibiotics, (2) the practice of microbiological sampling of infected foci before treatment, (3) the systematic surveillance of resistance profiles of pneumococci against antibiotics and (4) adequate vaccination of populations at risk.

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

Metabolomic assays of amoxicillin, cephapirin and ceftiofur in chicken muscle. Application to treated chicken samples by liquid chromatography quadrupole time-of-flight mass spectrometry

The aim of this study was to identity metabolites and transformation products (TPs) in chicken muscle from amoxicillin (AMX), cephapirin (PIR) and ceftiofur (TIO), which are antibiotics of the β-lactam family. Liquid chromatography coupled to quadrupole time-of-flight (QqTOF) mass spectrometry was utilized due to its high resolution, high mass accuracy and MS/MS capacity for elemental composition determination and structural elucidation. Amoxicilloic acid (AMA) and amoxicillin diketopiperazine (DKP) were found as transformation products from AMX. Desacetylcephapirin (DAC) was detected as a metabolite of PIR. Desfuroylceftiofur (DFC) and its conjugated compound with cysteine (DFC-S-Cys) were detected as a result of TIO in contact with chicken muscle tissue. The metabolites and transformation products were also monitored during the in vivo AMX treatment and slaughtering period. It was found that two days were enough to eliminate AMX and associated metabolites/transformation products after the end of administration.

Inserção C-H de carbenóides de ródio em água e reutilização do catalisador

A five-session laboratory experiment is described for the synthesis of a beta-lactam via Rh(II) catalysed intramolecular C-H insertion of a alpha-diazo-alpha-ethoxycarbonylacetamide. The metallo-carbene, responsible for the C-H bond activation, was generated from the diazo substrate and the catalyst Rh2(OAc)4. The high stability and solubility of the catalyst and the exclusive C-H insertion of the Rh-carbene allows the synthesis of this important heterocycle in water and the catalyst reutilization.

Regulation of excitotoxicity in thiamine deficiency : role of glutamate transporters.

L’excitotoxicité est un mécanisme physiopathologique majeur impliqué dans la pathogenèse de la déficience en thiamine (DT). Dans les régions cérébrales vulnérables à la DT, on observe une mort cellulaire induite par excitotoxicité dont l’origine semble être la conséquence d’une perturbation du métabolisme énergétique mitochondrial, d’une dépolarisation membranaire soutenue et d’une diminution de l’absorption du glutamate par les astrocytes suite à la diminution de l’expression des transporteurs EAAT1 et EAAT2. Il est clairement établi que le glutamate joue un rôle central dans l’excitotoxicité lors de la DT. Ainsi, la mise en évidence des mécanismes impliqués dans la diminution de l’expression des transporteurs du glutamate est essentielle à la compréhension de la physiopathologie de la DT. L’objectif de cette thèse consiste en l’étude de la régulation des transporteurs astrocytaires du glutamate et la mise au point de stratégies thérapeutiques ciblant la pathogenèse de l’excitotoxicité lors de l’encéphalopathie consécutive à la DT. Les principaux résultats de cette thèse démontrent des perturbations des transporteurs du glutamate à la fois dans des modèles animaux de DT et dans des astrocytes en culture soumis à une DT. La DT se caractérise par la perte du variant d’épissage GLT-1b codant pour un transporteur du glutamate dans le thalamus et le colliculus inférieur, les régions cérébrales affectées lors d’une DT, en l’absence de modification des niveaux d’ARNm. Ces résultats suggèrent une régulation post-transcriptionnelle de l’expression des transporteurs du glutamate en condition de DT. Les études basées sur l’utilisation d’inhibiteurs spécifiques des facteurs de transcription NFkB et de l’enzyme nucléaire poly(ADP)ribose polymérase-1 (PARP-1) démontrent que la régulation de l’expression du transporteur GLT-1 est sous le contrôle de voies de signalisation NFkB dépendantes de PARP-1. Cette étude démontre une augmentation de l’activation de PARP-1 et de NFkB dans les régions vulnérables chez le rat soumis à une DT et en culture d’astrocytes DT. L’inhibition pharmacologique du facteur de transcription NFkB par le PDTC induit une augmentation des niveaux d’expression de GLT-1, tandis que l’inhibition de PARP-1 par le DPQ conduit à l’inhibition de l’hyperactivation de NFkB observée lors de DT. L’ensemble de ces résultats met en évidence un nouveau mécanisme de régulation des transporteurs du glutamate par l’activation de PARP-1. L’accumulation de lactate est une caractéristique de la DT. Un traitement avec le milieu de culture d’astrocytes en condition de DT sur des cultures d’astrocytes naïfs induit une diminution de l’expression de GLT-1 ainsi qu’une inhibition de la capacité d’absorption du glutamate par les astrocytes naïfs. En revanche, l’administration de lactate exogène ne modifie pas le niveau d’expression protéique de GLT-1. Ainsi, des facteurs solubles autres que le lactate sont sécrétés par des astrocytes en condition de perturbation métabolique et peuvent potentiellement réguler l’activité des transporteurs du glutamate et contribuer à la pathogenèse du syncytium astroglial. En outre, la ceftriaxone, un antibiotique de la famille des β-lactamines, augmente de façon différentielle l’expression du variant-d’épissage GLT-1 dans le colliculus inférieur chez le rat DT et en culture d’astrocytes DT. Ces résultats suggèrent que la ceftriaxone peut constituer une avenue thérapeutique dans la régulation de l’activité des transporteurs du glutamate lors de DT. Pour conclure, la mort cellulaire d’origine excitotoxique lors de DT survient en conséquence d’une dysfonction mitochondriale associée à une perturbation du métabolisme énergétique cérébral. La modification de l’expression des transporteurs du gluatamate est sous le contrôle des voies de signalisation NFkB dépendantes du facteur PARP-1. De plus, l’inhibition métabolique et l’augmentation des sécrétions de lactate observées lors de DT peuvent également constituer un autre mécanisme physiopathologique expliquant la diminution d’expression des transporteurs de glutamate. Enfin, la ceftriaxone pourrait représenter une stratégie thérapeutique potentielle dans le traitement de la régulation de l’expression des transporteurs du glutamate et de la perte neuronale associés à l’excitotoxicité observée lors de DT.

Complete sequence and molecular epidemiology of IncK epidemic plasmid encoding bla(CTX-M-14)

Antimicrobial drug resistance is a global challenge for the 21st century with the emergence of resistant bacterial strains worldwide. Transferable resistance to beta-lactam antimicrobial drugs, mediated by production of extended-spectrum beta-lactamases (ESBLs), is of particular concern. In 2004, an ESBL-carrying IncK plasmid (pCT) was isolated from cattle in the United Kingdom. The sequence was a 93,629-bp plasmid encoding a single antimicrobial drug resistance gene, bla(CTX-M-14). From this information, PCRs identifying novel features of pCT were designed and applied to isolates from several countries, showing that the plasmid has disseminated worldwide in bacteria from humans and animals. Complete DNA sequences can be used as a platform to develop rapid epidemiologic tools to identify and trace the spread of plasmids in clinically relevant pathogens, thus facilitating a better understanding of their distribution and ability to transfer between bacteria of humans and animals.

Vibrational properties, crystal X-ray diffraction structure and quantum chemical calculations on a divalent sulfur substituted phthalimide: 1H-Isoindole-1,3(2H)-dione, 2-[(methoxycarbonyl)thio]

Structural and conformational properties of 1H-Isoindole-1,3(2H)-dione, 2-[(methoxycarbonyl)thio] (S-phthalimido O-methyl thiocarbonate) are analyzed using a combined approach including X-ray diffraction, vibrational spectra and theoretical calculation methods. The vibrational properties have been studied by infrared and Raman spectroscopies along with quantum chemical calculations (B3LYP and B3PW91 functional in connection with the 6-311++G** and aug-cc-pVDZ basis sets). The crystal structure was determined by X-ray diffraction methods. The substance crystallizes in the monoclinic P2(1)/c space group with a = 6.795(1), b = 5.109(1), c = 30.011(3) angstrom, beta = 90.310(3)degrees and Z = 4 molecules per unit cell. The conformation adopted by the N-S-C=O group is syn (C=O double bond in synperiplanar orientation with respect to the N-S single bond). The experimental molecular structure is well reproduced by the MP2/aug-cc-pVDZ method. (C) 2010 Elsevier B.V. All rights reserved.

Effect of the addition of invert sugar on the production of cephalosporin C in a fed-batch bioreactor

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)