Measurement of the branching fraction and CP content for the decay B0-->D(*+)D(*-).

We report a measurement of the branching fraction of the decay B0-->D(*+)D(*-) and of the CP-odd component of its final state using the BABAR detector. With data corresponding to an integrated luminosity of 20.4 fb (-1) collected at the Upsilon(4S) resonance during 1999-2000, we have reconstructed 38 candidate signal events in the mode B0-->D(*+)D(*-) with an estimated background of 6.2+/-0.5 events. From these events, we determine the branching fraction to be B(B0-->D(*+)D(*-))=[8.3+/-1.6(stat)+/-1.2(syst)]x10(-4). The measured CP-odd fraction of the final state is 0.22+/-0.18(stat)+/-0.03(syst).

Measurement of the B(0) lifetime with partially reconstructed B(0)-->D(-)l(+)nu(l) decays.

The B(0) lifetime was measured with a sample of 23 million BB pairs collected by the BABAR detector at the PEP-II e(+)e(-) storage ring during 1999 and 2000. Events from the semileptonic decay B(0)-->D(*-)l(+)nu(l) have been selected with a partial reconstruction method in which only the charged lepton and the slow pi from the D*--->D(0)pi(-) decay are reconstructed. The result is tau(B(0)) = 1.529+/-0.012(stat)+/-0.029(syst) ps.

Search for the rare decays B-->Kl(+)l(-) and B-->K(*)l(+)l(-).

We present results from a search for the flavor-changing neutral current decays B-->Kl(+)l(-) and B-->K(*)l(+)l(-), where l(+)l(-) is either an e(+)e(-) or mu(+)mu(-) pair. The data sample comprises 22.7 x 10(6) Upsilon(4S)-->B(-)B decays collected with the BABAR detector at the PEP-II B Factory. We obtain the 90% C.L. upper limits B(B-->Kl(+)l(-))<0.51 x 10(-6) and B(B-->K(*)l(+)l(-))<3.1 x 10(-6), close to standard model predictions for these branching fractions. We have also obtained limits on the lepton-family-violating decays B-->Ke+/-mu(-/+) and B-->K(*)e(+/-)mu(-/+).

Search for T and CP violation in B0-B-0 mixing with inclusive dilepton events.

We report the results of a search for T and CP violation in B0-B-0 mixing using an inclusive dilepton sample collected by the BABAR experiment at the PEP-II B Factory. The asymmetry between l+l+ and l-l- events allows us to compare the probabilities for B-0-->B0 and B0-->B-0 oscillations and thus probe T and CP invariance. Using a sample of 23 x 10(6) BB- pairs, we measure a same-sign dilepton asymmetry of A(T/CP) = [0.5+/-1.2(stat)+/-1.4(syst)]%. For the modulus of the ratio of complex mixing parameters p and q, we obtain q/p = 0.998+/-0.006(stat)+/-0.007(syst).

Measurement of B0-B-0 flavor oscillations in hadronic B0 decays.

Flavor oscillations of neutral B mesons have been studied in e+e- annihilation data collected with the BABAR detector at center-of-mass energies near the upsilon(4S) resonance. The data sample used for this purpose consists of events in which one B0 meson is reconstructed in a hadronic decay mode, while the flavor of the recoiling B0 is determined with a tagging algorithm that exploits the correlation between the flavor of the heavy quark and the charges of its decay products. From the time development of the observed mixed and unmixed final states, we determine the B0-B-0 oscillation frequency deltamd to be 0.516+/-0.016(stat)+/-0.010(syst) ps-1.

Measurement of the B0-B-0 oscillation frequency with inclusive dilepton events.

The B0-B-0 oscillation frequency has been measured with a sample of 23 x 10(6) BB- pairs collected with the BABAR detector at the PEP-II asymmetric B Factory at SLAC. In this sample, we select events in which both B mesons decay semileptonically and use the charge of the leptons to identify the flavor of each B meson. A simultaneous fit to the decay time difference distributions for opposite- and same-sign dilepton events gives deltamd = 0.493+/-0.012(stat)+/-0.009(syst) ps-1.

Dimerization and membrane anchors in extracellular targeting of vancomycin group antibiotics.

Antibiotics of the vancomycin group are shown to enhance their affinities for the bacterial cell wall by the devices of either dimerization (vancomycin and other glycopeptides which dimerize even more strongly) or use of a membrane anchor (teicoplanin); a chelate mechanism is suggested in both cases, as supported by antagonism experiments with the cell wall analog di-N-acetyl-L-Lys-D-Ala-D-Ala. These results may have implications for other binding processes which occur near membrane surfaces.

Kinetic and physical studies of beta-lactamase inhibition by a novel penem, BRL 42715.

The interactions of Staphylococcus aureus, Bacillus cereus I, TEM, Klebsiella pneumoniae K1 and Enterobacter cloacae P99 beta-lactamases with the novel penem inhibitor BRL 42715 were investigated kinetically and, in some cases, by electrospray mass spectrometry (e.s.m.s.). All the beta-lactamases were rapidly inactivated by BRL 42715, with second-order rate constants ranging from 0.17 to 6.4 microM-1.s-1. The initial stoichiometry of beta-lactamase inhibition was essentially 1:1, with the exception of the K1 enzyme. In this instance about 20 molecules of BRL 42715 were hydrolysed before the enzyme was completely inhibited. Inhibited beta-lactamases did not readily regain activity in the absence of BRL 42715, the half-lives for regeneration of free enzyme ranging from 5 min for the K1 beta-lactamase to over 2 days for the staphylococcal enzyme. Recovery of activity was incomplete for TEM-1, K1 and P99 beta-lactamases, suggesting partitioning of the inhibited enzymes to give a permanently (or at least very stable) inactivated species. Examination of the interactions of the penem with TEM, B. cereus I and P99 beta-lactamases by e.s.m.s. also showed rapid and stoichiometric binding of the inhibitor. In all cases a mass increase of 264 Da over the native enzyme was observed, corresponding to the molecular mass of BRL 42715, showing that no fragmentation of the penem occurred on reaction with the beta-lactamases.

Comparative activities of clavulanic acid, sulbactam, and tazobactam against clinically important beta-lactamases.

Clavulanic acid, sulbactam, and tazobactam are inhibitors of a variety of plasmid-mediated beta-lactamases. However, inhibition data for these three inhibitors with a wide range of different plasmid-mediated beta-lactamases have not yet been compared under the same experimental conditions. A number of groups have inferred that clavulanic acid inhibits extended-spectrum TEM and SHV beta-lactamases, but inhibition data have rarely been published. In this study, the 50% inhibitory concentrations of these three beta-lactamase inhibitors for 35 plasmid-mediated beta-lactamases have been determined. Of these 35 beta-lactamases, 20 were extended-spectrum TEM- or SHV-derived beta-lactamases. The other 15 enzymes were conventional-spectrum beta-lactamases such as TEM-1 and SHV-1. Clavulanic acid was a more potent inhibitor than sulbactam for 32 of the 35 plasmid-mediated beta-lactamases tested. In particular, clavulanic acid was 60 and 580 times more potent than sulbactam against TEM-1 and SHV-1, respectively, currently the two most clinically prevalent gram-negative plasmid-mediated beta-lactamases. Statistical analysis of the data of the 50% inhibitory concentrations showed that clavulanic acid was 20 times more active overall than sulbactam against the conventional-spectrum enzymes. In addition, clavulanic acid was 14 times more potent than sulbactam at inhibiting the extended-spectrum enzymes. Tazobactam also showed significantly greater activity than sulbactam against the two groups of beta-lactamases. There were no significant differences between the overall activities of tazobactam and clavulanic acid against the extended-spectrum TEM and SHV enzymes and conventional-spectrum enzymes, although differences in their inhibition profiles were observed.

beta-Lactamase ragged ends detected by electrospray mass spectrometry correlates poorly with multiple banding on isoelectric focusing.

Purified preparations of TEM-2, P99, Bacillus cereus I and B. cereus II beta-lactamases were examined by electrospray (ES) mass spectrometry. The ES mass spectra of the B. cereus enzymes revealed the presence of four to five components of different mass, corresponding to the loss of different numbers of N-terminal amino acids (ragged ends). The ES mass spectra of both TEM-2 and P99 consisted of a single component with no evidence of ragged ends. All four beta-lactamase preparations were visualized on isoelectric focusing (IEF) gels stained with nitrocefin to investigate a possible correlation between IEF patterns and ragged ends. Multiple banding patterns were seen with each beta-lactamase preparation. Although these may correlate with the presence of ragged ends in the two B. cereus preparations, the satellite bands seen with P99 and TEM-2 were not associated with differences detected by ES mass spectrometry. In this study we have shown for the first time that beta-lactamase satellite bands seen on IEF are not always associated with ragged ends. Furthermore, we have illustrated the use of ES mass spectrometry to characterize the extent of ragged end formation in protein samples. This is of particular significance if the sample is required for detailed biochemical or crystallography experiments.

The metabolic cost of backpack and shoulder load carriage.

Eleven healthy male volunteer soldiers (mean [SD] age 24.0 [2.8] years, stature 174.1 [5.2] cm, body weight 73.2 [10.8] kg, body fat 14.2 [5.0]% and maximal oxygen uptake 4.1 [0.4] 1 min-1) walked at 4.8 km h-1 on a motor driven treadmill for 5 min at each of three gradients (0, 2.5 and 5%) whilst carrying a two-part 26 kg load either on each shoulder or strapped to a backpack frame. The load was made up of two cylinders, one weighing 18.4 kg and the other weighing 7.6 kg. For all treadmill gradients the mean (SD) backpacking heart rates and oxygen uptakes (0% gradient, 122 [10] beats min-1, 1.51 [0.11] 1 min-1; 2.5% gradient, 135 [10] beats min-1, 1.81 [0.13] 1 min-1; 5% gradient, 155 [7] beats min-1, 2.21 [0.11] 1 min-1) were significantly (p less than 0.001) lower than for shoulder load carriage (0% gradient, 130 [9] beats min-1, 1.70 [0.12] 1 min-1, 2.5% gradient, 147 [8] beats min-1; 2.01 [0.10] 1 min-1; 5% gradient 164 [9] beats min-1, 2.39 [0.11] 1 min-1). The relative oxygen cost of backpacking was 4.3-4.7% VO2 max lower than for shoulder load carriage. It is concluded that the metabolic cost of backpacking an asymmetric two part 26 kg load was significantly less than for shoulder load carriage when walking at 4.8 km h-1 on a treadmill over gradients of 0-5%.(ABSTRACT TRUNCATED AT 250 WORDS)

Synthesis and antibacterial activity of C-2 alkenylthio-carbapenem derivatives.

A number of C-2 aminoalkenylthio-carbapenem derivatives possessing both the (5R,6R, 8S)- and the (5R,6S,8R)-stereochemistries have been prepared from the olivanic acids MM 22382 and MM 22383, respectively. Certain members of this new class of compounds displayed potent, broad spectrum antibacterial activity as well as improved stability to human kidney homogenate.

MM 45289, a potent glycopeptide antibiotic which interacts weakly with diacetyl-L-lysyl-D-alanyl-D-alanine.

MM 45289 (A82846A, eremomycin), a glycopeptide antibiotic of the vancomycin type, was confirmed to have improved antibacterial activity over vancomycin. However its affinity (Ka) for the target site peptide mimetic diacetyl-L-lysyl-D-alanyl-D-alanine (DALAA) was 23-fold lower. Concentrations of DALAA required to reverse the antibacterial activity of MM 45289 were in the order of 10 to 50-fold higher than for vancomycin. These results have implications for both mode of action studies and mechanism-based screening strategies for this class of antibiotic.

Biological properties of ER 42859, a novel erythromycin derivative.

The antimicrobial activity of a new semi-synthetic oral erythromycin derivative, ER 42859, was evaluated in vitro and in vivo in comparison with erythromycin, spiramycin, josamycin, oleandomycin and the newer semi-synthetic derivatives flurithromycin, roxithromycin and A-56268. MIC values of ER 42859 were superior to those of roxithromycin, oleandomycin, josamycin and spiramycin but generally 2-fold poorer than those of erythromycin. The activity equalled that of erythromycin against Haemophilus influenzae and was superior to that of roxithromycin and A-56268 against this organism. MIC values of the compound were greatly influenced by pH due to the dibasic nature of the molecule. ER 42859 had markedly superior activity to erythromycin, spiramycin, josamycin, oleandomycin and flurithromycin against experimental infections in mice and similar activity to roxithromycin and A-56268. Blood and tissue levels were high and prolonged in rodents. In volunteers, blood levels were prolonged but inferior to those of erythromycin.

9,11-cyclic acetal derivatives of (9S)-9-dihydroerythromycin A.

A series of 9,11-cyclic acetal derivatives of (9S)-9-dihydroerythromycin A (4) have been prepared and their antibacterial activities compared to those of erythromycin A and 9-dihydroerythromycin A. Many of the cyclic acetal derivatives showed better antibacterial activity than their parent 4. In particular, the acetaldehyde acetal (9-O,11-O-ethylidene-9-dihydroerythromycin A) (8b) showed good antibacterial activity in comparison with erythromycin A but was not sufficiently improved in vivo to warrant progression.

Erythromycin A 11,12-methylene acetal.

Erythromycin A 11,12-methylene acetal (5) and the corresponding 9-methoxime, 9-dihydro, and 8-hydroxy derivatives have been prepared and their antibacterial activities compared with those of erythromycin A and its 11,12-cyclic carbonate. The simple methylene acetal 5 showed excellent activity against Gram-positive organisms in vitro.

Uptake of erythromycin by McCoy and HEp2 cells: its dependence on cellular pH gradients.

The accumulation of erythromycin, a weak lipophilic base, was studied in two mammalian cell lines. Uptake was similar in both cell types, being highly pH dependent and not saturated even at external concentrations of 1000 mg/l. At pH 6.7 the ratio of cellular:extracellular erythromycin concentrations (C/E) was only 2.0, whereas at pH 7.5 it was as high as 7.0. Carbonylcyanide-m-chlorophenyl hydrazone, an uncoupler of oxidative phosphorylation which was used to collapse cellular pH gradients, caused a 50% reduction in C/E ratio within five minutes when the external pH was greater than 6.9 but had no effect at or below pH 6.7. These results suggest that ion-trapping plays a major role in the accumulation of erythromycin by these cells.

A simple rapid technique to measure neutrophil or serum bactericidal activity.

A rapid and simple technique to measure both serum and neutrophil bactericidal activity is described. It is based on microtitre equipment and avoids the cumbersome and time consuming serial dilution and plate counting of conventional experiments. A constant concentration of neutrophils and/or serum is incubated with a series of dilutions of bacterial suspensions. Bacterial survival is estimated at various times from their ability to form colonies on agar and the results conveniently expressed as the largest bacterial population that can be eliminated by either neutrophils or serum alone. The technique is as simple and quick as a standard microtitre MBC test commonly used for antibiotics and scoring of the results takes only a few seconds per sample. Data are presented showing the close correlation between results obtained using this technique and those from conventional tests of neutrophil bactericidal activity and phagocytosis-associated chemiluminescence.