Kinetic characterization of GES-22 beta-lactamase harboring the M169L clinical mutation

The class A p-lactamase GES-22 has been identified in Acinetobacter baumannii isolates in Turkey, and subsequently shown to differ from GES-11 by a single substitution (M169L). Because M169 is part of the omega loop, a structure that is known to have major effects on substrate selectivity in class A beta-lactamases, we expressed, purified and kinetically characterized this novel variant. Our results show that compared with GES-11(6xHis), GES-22(6xHis) displays more efficient hydrolysis of penicillins, and aztreonam, but a loss of efficiency against ceftazidime. In addition, the M169L substitution confers on GES-22 more efficient hydrolysis of the mechanistic inhibitors clavulanic acid and sulbactam. These effects are highly similar to other mutations at the homologous position in other class A beta-lactamases, suggesting that this methionine has a key structural role in aligning active site residues and in substrate selectivity across the class.Recep Tayyip Erdogan University:BAP-2013.102.03.12 Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK): TUBITAK-113Z054 United States Department of Health & Human Services National Institutes of Health (NIH) - USA 1R15AI082416 Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) 2214-

Acinetobacter baumannii in intensive care unit: A novel system to study clonal relationship among the isolates

<p>Abstract</p> <p>Background</p> <p>The nosocomial infections surveillance system must be strongly effective especially in highly critic areas, such as Intensive Care Units (ICU). These areas are frequently an epidemiological epicentre for transmission of multi-resistant pathogens, like <it>Acinetobacter baumannii</it>. As an epidemic outbreak occurs it is very important to confirm or exclude the genetic relationship among the isolates in a short time. There are several molecular typing systems used with this aim. The Repetitive sequence-based PCR (REP-PCR) has been recognized as an effective method and it was recently adapted to an automated format known as the DiversiLab system.</p> <p>Methods</p> <p>In the present study we have evaluated the combination of a newly introduced software package for the control of hospital infection (VIGI@ct) with the DiversiLab system. In order to evaluate the reliability of the DiversiLab its results were also compared with those obtained using f-AFLP.</p> <p>Results</p> <p>The combination of VIGI@ct and DiversiLab enabled an earlier identification of an <it>A. baumannii </it>epidemic cluster, through the confirmation of the genetic relationship among the isolates. This cluster regards 56 multi-drug-resistant <it>A. baumannii </it>isolates from several specimens collected from 13 different patients admitted to the ICU in a ten month period. The <it>A. baumannii </it>isolates were clonally related being their similarity included between 97 and 100%. The results of the DiversiLab were confirmed by f-AFLP analysis.</p> <p>Conclusion</p> <p>The early identification of the outbreak has led to the prompt application of operative procedures and precautions to avoid the spread of pathogen. To date, 6 months after the last <it>A. baumannii </it>isolate, no other related case has been identified.</p

Pathogenesis of adolescent idiopathic scoliosis in girls - a double neuro-osseous theory involving disharmony between two nervous systems, somatic and autonomic expressed in the spine and trunk: possible dependency on sympathetic nervous system and hormones with implications for medical therapy

Anthropometric data from three groups of adolescent girls - preoperative adolescent idiopathic scoliosis (AIS), screened for scoliosis and normals were analysed by comparing skeletal data between higher and lower body mass index subsets. Unexpected findings for each of skeletal maturation, asymmetries and overgrowth are not explained by prevailing theories of AIS pathogenesis. A speculative pathogenetic theory for girls is formulated after surveying evidence including: (1) the thoracospinal concept for right thoracic AIS in girls; (2) the new neuroskeletal biology relating the sympathetic nervous system to bone formation/resorption and bone growth; (3) white adipose tissue storing triglycerides and the adiposity hormone leptin which functions as satiety hormone and sentinel of energy balance to the hypothalamus for long-term adiposity; and (4) central leptin resistance in obesity and possibly in healthy females. The new theory states that AIS in girls results from developmental disharmony expressed in spine and trunk between autonomic and somatic nervous systems. The autonomic component of this double neuro-osseous theory for AIS pathogenesis in girls involves selectively increased sensitivity of the hypothalamus to circulating leptin (genetically-determined up-regulation possibly involving inhibitory or sensitizing intracellular molecules, such as SOC3, PTP-1B and SH2B1 respectively), with asymmetry as an adverse response (hormesis); this asymmetry is routed bilaterally via the sympathetic nervous system to the growing axial skeleton where it may initiate the scoliosis deformity (leptin-hypothalamic-sympathetic nervous system concept = LHS concept). In some younger preoperative AIS girls, the hypothalamic up-regulation to circulating leptin also involves the somatotropic (growth hormone/IGF) axis which exaggerates the sympathetically-induced asymmetric skeletal effects and contributes to curve progression, a concept with therapeutic implications. In the somatic nervous system, dysfunction of a postural mechanism involving the CNS body schema fails to control, or may induce, the spinal deformity of AIS in girls (escalator concept). Biomechanical factors affecting ribs and/or vertebrae and spinal cord during growth may localize AIS to the thoracic spine and contribute to sagittal spinal shape alterations. The developmental disharmony in spine and trunk is compounded by any osteopenia, biomechanical spinal growth modulation, disc degeneration and platelet calmodulin dysfunction. Methods for testing the theory are outlined. Implications are discussed for neuroendocrine dysfunctions, osteopontin, sympathoactivation, medical therapy, Rett and Prader-Willi syndromes, infantile idiopathic scoliosis, and human evolution. AIS pathogenesis in girls is predicated on two putative normal mechanisms involved in trunk growth, each acquired in evolution and unique to humans

Antibiotico-resistenza e Batteri ambientali

La diffusione di patogeni batterici altamente resistenti alle maggiori classi di farmaci antibatterici rappresenta un importante problema per la sanità pubblica mondiale. La straordinaria capacità dei microrganismi a sviluppare meccanismi di resistenza si è sempre tradotta, di seguito all’introduzione di nuovi farmaci antibatterici, nell’emergenza e nella diffusione di isolati clinici resistenti. In questa prospettiva, lo studio complessivo del resistoma, mediante genomica e metagenomica, di specie batteriche prevalentemente ambientali ha rivelato la presenza di numerosi determinanti di resistenza agli antibiotici, talvolta capaci di conferire resistenza ai farmaci antibatterici di più recente sviluppo (e. g. carbapenemi, daptomicina). Lo studio molecolare di tali determinanti di resistenza, oltre ad offrire un interessante paradigma per la comprensione dei meccanismi alla base dell’evoluzione e della diversificazione della funzione delle proteine, rivelano altri aspetti interessanti. In particolare, è stato osservato che alcuni fattori di resistenza prodotti da batteri ambientali sarebbero enzimi apparentemente dotati di attività promiscua, indicando che la loro funzione primaria potrebbe essere diversa dalla resistenza agli antibiotici e che i meccanismi di resistenza potrebbe anche essere il frutto di evoluzione convergente. Rimane controverso l’impatto dell’utilizzo di composti antibatterici legato ad attività umane sull’evoluzione e sulla presenza di meccanismi di resistenza in batteri ambientali. Tuttavia, è stata dimostrata l’origine ambientale di alcuni determinanti di resistenza recentemente comparsi in isolati clinici, sottolineando l’importanza di batteri ambientali nell’evoluzione complessiva del resistoma di patogeni umani. Una maggiore conoscenza del resistoma ambientale e della sua evoluzione è quindi importante anche in vista dello sviluppo di nuovi farmaci antibatterici

Clinically-relevant β-lactamases: Structure-function relationships and inhibition.

β-lactamases are important resistance factors to β-lactam antibiotics, especially in Gram-negative bacteria, that show an extraordinary diversity in terms of structure, mechanism of hydrolysis and substrate profile and susceptibility to inhibitors. Two families of β-lactamases are currently known: (a) the active site serine β-lactamases which perform hydrolysis of β-lactams via an acylation-deacylation mechanism and (b) the metallo-β-lactamases, which require one or two zinc ion(s) for activity. The current knowledge on β-lactamase structure-function relationships will be reviewed, with a particular focus on carbapenemases, such as KPC-2, VIM-2 and NDM-1 metallo-β-lactamases and OXA-type carbapenem-hydrolyzing enzymes, which currently represent among the most worrisome resistance determinants currently emerging in Pseudomonas aeruginosa, Acinetobacter baumannii and Enterobacteriaceae. The identification and development of new β-lactamase inhibitors, whose clinical need is stronger than ever, is an active field of research. The properties of novel β-lactamase inhibitors, some of which are under clinical development (e. g. NXL104), will be discussed

Characterization of a novel 3-N-aminoglycoside acetyltransferase gene, aac(3)-Ic, from a Pseudomonas aeruginosa integron.

Antimicrob Agents Chemother. 2003 May; 47(5):1746-8