Performance of a System for Rapid Phenotypic Antimicrobial Susceptibility Testing of Gram-Negative Bacteria Directly from Positive Blood Culture Bottles.

The rapid administration of optimal antimicrobial treatment is paramount for the treatment of bloodstream infections (BSIs), and rapid antimicrobial susceptibility testing (AST) results are essential. Q-linea has developed the ASTar system, a rapid phenotypic AST device. Here, we report the performance of the ASTar BC G- (Gram-negative) kit when assessed according to the ISO 20776-2:2007 standard for performance evaluation of in vitro diagnostic AST devices. The evaluated ASTar BC G- kit uses a broad panel of 23 antimicrobials for the treatment of BSIs caused by Gram-negative fastidious and nonfastidious bacteria across a range of 6 to 14 2-fold dilutions, including cefoxitin as a screening agent for AmpC-producing Enterobacterales. The ASTar system processes blood culture samples to generate data on MICs and susceptible, intermediate, or resistant (SIR) category. The automated protocol includes concentration determination and concentration adjustment to enable a controlled inoculum, followed by broth microdilution (BMD) and microscopy performed continuously to generate MIC values within approximately 6 h once the test is run on the ASTar system. The performance of the ASTar system was assessed against the ISO 20776-2:2007 standard BMD reference method. Testing was performed across three sites, with results from 412 contrived blood cultures and 74 fresh clinical blood cultures. The ASTar system was also tested for reproducibility, with triplicate testing of 11 strains. The accuracy study comprised 8,650 data points of bacterium-antimicrobial tests. The ASTar system demonstrated an overall essential agreement (EA) of 95.8% (8,283/8,650) and a categorical agreement (CA) of 97.6% (8,433/8,639) compared to the reference BMD method. The overall rate of major discrepancies (MDs) was 0.9% (62/6,845), and that of very major discrepancies (VMDs) was 2.4% (30/1,239). This study shows that the ASTar system delivers reproducible results with overall EA and CA of >95%.

O6-Methylguanine-DNA methyltransferase status in neuroendocrine tumours: prognostic relevance and association with response to alkylating agents.

BACKGROUND: O(6)-Methylguanine-DNA methyltransferase (MGMT) loss of expression has been suggested to be predictive of response to temozolomide in neuroendocrine tumours (NETs), but so far, only limited data are available. We evaluated the prognostic and predictive value of MGMT status, assessed by two molecular methods and immunohistochemistry, in a large series of NETs of different origins. METHODS: A total of 107 patients, including 53 treated by alkylants (temozolomide, dacarbazine or streptozotocin), were retrospectively studied. In each case, we used methyl-specific PCR (MS-PCR) and pyrosequencing for evaluation of promoter methylation and immunohistochemistry for evaluation of protein status. RESULTS: MGMT promoter methylation was detected in 12 out of 99 (12%) interpretable cases by MS-PCR and in 24 out of 99 (24%) by pyrosequencing. O(6)-Methylguanine-DNA methyltransferase loss of expression was observed in 29 out of 89 (33%) interpretable cases. Status of MGMT was not correlated with overall survival (OS) from diagnosis. Progression-free survival and OS from first alkylant use (temozolomide, dacarbazine and streptozotocin) were higher in patients with MGMT protein loss (respectively, 20.2 vs 7.6 months, P<0.001 and 105 vs 34 months, P=0.006) or MGMT promoter methylation assessed by pyrosequencing (respectively, 26.4 vs 10.8 months, P=0.002 and 77 vs 43 months, P=0.026). CONCLUSIONS: Our results suggest that MGMT status is associated with response to alkylant-based chemotherapy in NETs.

Radical ions with nearly degenerate ground state: correlation between the rate of spin-lattice relaxation and the structure of adiabatic potential energy surface.

Paramagnetic spin-lattice relaxation (SLR) in radical cations (RCs) of the cycloalkane series in liquid solution was studied and analyzed from the point of view of the correlation between the relaxation rate and the structure of the adiabatic potential energy surface (PES) of the RCs. SLR rates in the RCs formed in x-ray irradiated n-hexane solutions of the cycloalkanes studied were measured with the method of time-resolved magnetic field effect in the recombination fluorescence of spin-correlated radical ion pairs. Temperature and, for some cycloalkanes, magnetic field dependences of the relaxation rate were determined. It was found that the conventional Redfield theory of the paramagnetic relaxation as applied to the results on cyclohexane RC, gave a value of about 0.2 ps for the correlation time of the perturbation together with an unrealistically high value of 0.1 T in field units for the matrix element of the relaxation transition. The PES structure was obtained with the DFT quantum-chemical calculations. It was found that for all of the cycloalkanes RCs considered, including low symmetric alkyl-substituted ones, the adiabatic PESes were surfaces of pseudorotation due to avoided crossing. In the RCs studied, a correlation between the SLR rate and the calculated barrier height to the pseudorotation was revealed. For RCs with a higher relaxation rate, the apparent activation energies for the SLR were similar to the calculated heights of the barrier. To rationalize the data obtained it was assumed that the vibronic states degeneracy, which is specific for Jahn-Teller active cyclohexane RC, was approximately kept in the RCs of substituted cycloalkanes for the vibronic states with the energies above and close to the barrier height to the pseudorotation. It was proposed that the effective spin-lattice relaxation in a radical with nearly degenerate low-lying vibronic states originated from stochastic crossings of the vibronic levels that occur due to fluctuations of the interaction between the radical and the solvent. The magnitude of these fluctuations, ~100 cm(-1), determines the upper scale of the unperturbed splitting between the vibronic states, for which the manifestation of this paramagnetic relaxation mechanism could be expected. Our estimate for the relaxation rate derived using standard Landau-Zener model of nonadiabatic transitions at the level crossing agrees with the experimental data. This paramagnetic relaxation mechanism can also be operative in paramagnetic species of other types such as linear radicals, radicals with threefold degeneracy, paramagnetic centers in crystals, etc. It looks likely that the proposed SLR mechanism can be quenched by a fast vibrational relaxation in radicals.

Radical cations of branched alkanes as observed in irradiated solutions by the method of time-resolved magnetic field effect.

Spin dynamics in radical ion pairs formed under ionizing irradiation of n-hexane solutions of two branched alkanes 2,3-dimethylbutane and 2,2,4-trimethylpentane has been studied by the method of time-resolved magnetic field effect in recombination fluorescence. Experimental curves of the magnetic field effect are satisfactorily described by assuming that the spin dynamics is determined by the hyperfine interactions in the radical cation (RC) of branched alkane under study with hyperfine coupling (HFC) constants averaged by internal rotations of RC fragments. The HFC constants determined from the magnetic field effect curves are close to those estimated within DFT B3LYP approach. Analysis of the results indicates that at room temperature the lifetimes of the RC of the studied branched alkanes amount to, at least, tens of nanoseconds.

Detection of the kinetics of biochemical reactions with oxygen using exchange broadening in the ESR spectra of nitroxide radicals.

To detect changes in the oxygen concentration during biochemical reactions, the exchange broadening in the ESR spectra of nitroxide radicals caused by the dissolved oxygen, has been used. The measurements have been carried out using changes in the width either of the proton hyperfine structure components or of the nitrogen hyperfine structure line with an unresolved proton structure. Detection of mitochondrial respiration in a volume of about 10(-3) cm 3 and respiration for 100 +/- 5 liver cells in a volume of about 10(-4) cm3 has been carried out.