Shared and Unique Evolutionary Trajectories to Ciprofloxacin Resistance in Gram-Negative Bacterial Pathogens.

Resistance to the broad-spectrum antibiotic ciprofloxacin is detected at high rates for a wide range of bacterial pathogens. To investigate the dynamics of ciprofloxacin resistance development, we applied a comparative resistomics workflow for three clinically relevant species of Gram-negative bacteria: Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa. We combined experimental evolution in a morbidostat with deep sequencing of evolving bacterial populations in time series to reveal both shared and unique aspects of evolutionary trajectories. Representative clone characterization by sequencing and MIC measurements enabled direct assessment of the impact of mutations on the extent of acquired drug resistance. In all three species, we observed a two-stage evolution: (i) early ciprofloxacin resistance reaching 4- to 16-fold the MIC for the wild type, commonly as a result of single mutations in DNA gyrase target genes (gyrA or gyrB), and (ii) additional genetic alterations affecting the transcriptional control of the drug efflux machinery or secondary target genes (DNA topoisomerase parC or parE). IMPORTANCE The challenge of spreading antibiotic resistance calls for systematic efforts to develop more "irresistible" drugs based on a deeper understanding of dynamics and mechanisms of antibiotic resistance acquisition. To address this challenge, we have established a comparative resistomics approach which combines experimental evolution in a continuous-culturing device, the morbidostat, with ultradeep sequencing of evolving microbial populations to identify evolutionary trajectories (mutations and genome rearrangements) leading to antibiotic resistance over a range of target pathogens. Here, we report the comparative resistomics study of three Gram-negative bacteria (Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa), which revealed shared and species-specific aspects of the evolutionary landscape leading to robust resistance against the clinically important antibiotic ciprofloxacin. Despite some differences between morbidostat-deduced mutation profiles and those observed in clinical isolates of individual species, a cross-species comparative resistomics approach allowed us to recapitulate all types of clinically relevant ciprofloxacin resistance mechanisms. This observation supports the anticipated utility of this approach in guiding rational optimization of treatment regimens for current antibiotics and the development of novel antibiotics with minimized resistance propensities.

Effect of ispronicline, a neuronal nicotinic acetylcholine receptor partial agonist, in subjects with age associated memory impairment (AAMI).

Cognitive decline seen in the normal elderly is associated with selective loss of neuronal nicotinic acetylcholine receptors (nAChRs). Nicotine given either by inhalation or transdermally helps cognition, but unacceptable side effects limit its utility. The present study assessed the safety, tolerability and effect on cognition of ispronicline, a highly selective partial agonist at the 4beta2 nAChR, in elderly subjects (n =76) with age associated memory impairment (AAMI). This double-blind, placebo-controlled cross-over study explored ascending oral doses of ispronicline in the range 50-150 mg given as a single morning dose for a period of 3 weeks. Pharmacokinetics (PK) were assessed, as well as cognitive function measured by means of the Cognitive Drug Research (CDR) computerized test battery. Ispronicline had a favourable safety profile and was well tolerated at doses below 150 mg. No effect of clinical importance was seen on biochemistry, haematology, urine analysis, vital signs, electrocardiogram (ECG) or Holter monitoring. The most frequent drug induced adverse event was light-headedness (dizziness). A beneficial effect was seen on cognition across the dose range. This was most marked at 50 mg on factors measuring attention and episodic memory. PK analysis indicated a plasma Cmax range of 5-25/35 ng/ml ispronicline was associated with the most beneficial effect. These early results demonstrate ispronicline was well tolerated and did not display the side effects typical of nicotine. Ispronicline also had a beneficial effect on cognition in subjects with AAMI. This was seen most strongly in a Cmax range that had been predicted from pre-clinical animal studies.

Effects on monoamine levels in rat CNS after chronic administration of cocaine / Efecto de la administración crónica de cocaína sobre los niveles de monoaminas en el sistema nervioso central de la rata

We have previously reported time dependent and dose dependent changes in the rat dopaminergic receptor system following chronic administration of cocaine (upregulation of cacaina, D1, and DA-uptake sites). We have now evaluated the effects of chronic cacaine exposure on the central catecholamine/indolamine neurotransmitter systems. Groups of rats were injected with cocaine (15 mg/kg, i.p, b.i.d.) or saline for 1,3,7,14 or 21 days. Cortical and striatal tissues were analysed for morepinephrine, dopamine, serotomin and their primary metabolites using a HPLC-ECD method. Chronic administration of cacaine did not change the cortical and striatal concentrations of the neurotransmitters under study; except, for a transient increase in the cortical MHPG concentration on day 3. These results suggest that changes in the dopaminergic receptor system following chronic cocaine exposure are not to changes in the neurotransmitter concentrations

Intoxicación crónica con manganeso: cuantificación autorradiográfica de los receptores colinérgicos muscarínicos en el cerebro de ratón / Chronic manganese poisoning: autoradiograohic cuantitation of muscarinic cholinergic receptors in mouse brain

El tratamiento crónico con cloruro de manganeso (5 mg Mn/kg/día), durante 9 semanas, no afectó la unión del radioligando [3H]-quinuclidinil benzilato a los receptores colinérgicos muscarínicos en el cerebro de ratón. Mediante técnica autorradiográfica se determinó la localización anatómica precisa de los receptores y se procedió a la cuantificació de los mismos en los cortes coronales del bulbo olfatorio y del cerebro medio. A la luz de los resultados obtenidos podemos concluir que, en nuestras condiciones experimentales, no se producen alteraciones en la densidad de los receptores colinérgicos muscarínicos en el cerebro de ratones intoxicados con manganeso