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Antibacterial Effects of Amoxicillin-Clavulanate against Streptococcus pneumoniae and Haemophilus influenzae Strains for Which MICs Are High, in an In Vitro Pharmacokinetic Model. Alasdair P. MacGowan, 2004.The antibacterial effect of amoxicillin-clavulanate in two formulations, pharmacokinetically enhanced 16:1 amoxicillin-clavulanate twice a day (b.i.d.) and standard 7:1 amoxicillin-clavulanate b.i.d., were studied in an in vitro pharmacokinetic model of infection . Five strains of Streptococcus pneumoniae and two of Haemophilus influenzae, all associated with raised MICs (2 to 8 mg/liter), were used . The antibacterial effect was measured over 24 h by the area under the bacterial kill curve (AUBKC) and the log change in viable count at 24 h (  24) . A high 108 CFU/ml and low 106 CFU/ml initial inocula were used . Employing the
24 effect measure, the time above MIC (T>MIC) 50% maximum effect (EC50) for S . pneumoniae was in the range 21 to 28% with an 80% maximal response of 41 to 51%, for the AUBKC measure, the value was 26 to 39%, irrespective of inoculum . For H . influenzae, the T>MIC EC50 was 28 to 37%, and the 80% maximum response was 32 to 48% for the
24 measure and 20 to 48% for AUBKC . The maximum response occurred at a T>MIC of 50 to 60% for both species and inocula . The S . pneumoniae data were analyzed by analysis of variance to assess the effect of inoculum, formulation, and MIC on antibacterial effect . Standard and enhanced formulations had different effects depending on MIC, with the standard formulation less effective at higher amoxicillin-clavulanate MICs . This is explained by the greater T>MICs of the enhanced formulation . Although resistant to amoxicillin-clavulanate by conventional breakpoints, S . pneumoniae and H . influenzae strains for which MICs are 2 or 4 mg/liter may well respond to therapy with pharmacokinetically enhanced formulation amoxicillin-clavulanate .
Evolutionary Relationships of Primary Prokaryotic Endosymbionts of Whiteflies and Their Hosts. MyLo Ly Thao, 2004.Whiteflies (Hemiptera: Sternorrhyncha: Aleyrodidae) are plant sap-sucking insects that harbor prokaryotic primary endosymbionts (P-endosymbionts) within specialized cells located in their body cavity . Four-kilobase DNA fragments containing 16S-23S ribosomal DNA (rDNA) were amplified from the P-endosymbiont of 24 whiteflies from 22 different species of 2 whitefly subfamilies . In addition, 3-kb DNA fragments containing mitochondrial cytB, nd1, and large-subunit rDNA (LrDNA) were amplified from 17 whitefly species . Comparisons of the P-endosymbiont (16S-23S rDNA) and host (cytB-nd1-LrDNA) phylogenetic trees indicated overall congruence consistent with a single infection of a whitefly ancestor with a bacterium and subsequent cospeciation (cocladogenesis) of the host and the P-endosymbiont . On the basis of both the P-endosymbiont and host trees, the whiteflies could be subdivided into at least five clusters . The major subdivision was between the subfamilies Aleyrodinae and Aleurodicinae . Unlike the P-endosymbionts of may other insects, the P-endosymbionts of whiteflies were related to Pseudomonas and possibly to the P-endosymbionts of psyllids . The lineage consisting of the P-endosymbionts of whiteflies is given the designation "Candidatus Portiera" gen . nov., with a single species, "Candidatus Portiera aleyrodidarum" sp . nov . Salt-Dependent Expression of Glucosylglycerol-Phosphate Synthase, Involved in Osmolyte Synthesis in the Cyanobacterium Synechocystis sp . Strain PCC 6803. Kay Marin, 2002.The cyanobacterium Synechocystis sp . strain PCC 6803 is able to acclimate to levels of salinity ranging from freshwater to twice the seawater concentrations of salt by accumulating the compatible solute glucosylglycerol (GG) . Expression of the ggpS gene coding for the key enzyme (glucosylglycerol-phosphate synthase) in GG synthesis was examined in detail . Under control conditions, the GgpS protein is stable, so that weak constitutive transcription of the ggpS gene resulted in a significant protein content . However, the enzyme activity was biochemically switched off, and no GG was detectable . After a salt shock, an immediate increase in mRNA content proportional to the salt content occurred, while the GgpS protein and GG contents rose in a linear manner . Furthermore, the stability of the ggpS mRNA increased transiently . In salt-acclimated cells expression of the ggpS gene, the GgpS protein content, and the amount of accumulated GG depended linearly on the external salt concentration . Mapping of the 5' end of the ggpS transcript revealed a long nontranslated 5' sequence and a putative typical cyanobacterial promoter, which did not show any obvious salt-regulatory element . The alternative  factor
F was found to be involved in salt-dependent regulation of ggpS, since in a
F mutant induction of this gene was strongly reduced . The present study demonstrated that in addition to biochemical regulation of GgpS activity, alterations of ggpS expression are involved in regulation of GG synthesis in Synechocystis sp . strain PCC 6803 . A model showing the interaction of the two regulatory levels is presented .
Deficiency of a Sinorhizobium meliloti bacA Mutant in Alfalfa Symbiosis Correlates with Alteration of the Cell Envelope. Gail P. Ferguson, 2002.The BacA protein is essential for the long-term survival of Sinorhizobium meliloti and Brucella abortus within acidic compartments in plant and animal cells, respectively . Since both the S . meliloti and B . abortus bacA mutants have an increased resistance to bleomycin, it was hypothesized that BacA was a transporter of bleomycin and bleomycin-like compounds into the bacterial cell . However, our finding that the S . meliloti bacA mutant also has an increased sensitivity to detergents, a hydrophobic dye, ethanol, and acid pH supported a model in which BacA function affects the bacterial cell envelope . In addition, an S . meliloti lpsB mutant that is defective at a stage in infection of the host similar to that found for a bacA mutant is also sensitive to the same agents, and the carbohydrate content of its lipopolysaccharide (LPS) is altered . However, analysis of crude preparations of the bacA mutant LPS suggested that, unlike that for LpsB, BacA function did not affect the carbohydrate composition of the LPS . Rather, we found that at least one function of BacA is to affect the distribution of LPS fatty acids, including a very-long-chain fatty acid thought to be unique to the  -proteobacteria, including B . abortus .
AniA Regulates Reserve Polymer Accumulation and Global Protein Expression in Rhizobium etli. Sergio Encarnación, 2002.Previously, it was reported that the oxidative capacity and ability to grow on carbon sources such as pyruvate and glucose were severely diminished in the Rhizobium etli phaC::  Smr/Spr mutant CAR1, which is unable to synthesize poly-ß-hydroxybutyric acid (PHB) (M . A . Cevallos, S . Encarnación, A . Leija, Y . Mora, and J . Mora, J . Bacteriol . 178:1646-1654, 1996) . By random Tn5 mutagenesis of the phaC strain, we isolated the mutants VEM57 and VEM58, both of which contained single Tn5 insertions and had recovered the ability to grow on pyruvate or glucose . Nucleotide sequencing of the region surrounding the Tn5 insertions showed that they had interrupted an open reading frame designated aniA based on its high deduced amino acid sequence identity to the aniA gene product of Sinorhizobium meliloti . R . etli aniA was located adjacent to and divergently transcribed from genes encoding the PHB biosynthetic enzymes ß-ketothiolase (PhaA) and acetoacetyl coenzyme A reductase (PhaB) . An aniA::Tn5 mutant (VEM5854) was constructed and found to synthesize only 40% of the wild type level of PHB . Both VEM58 and VEM5854 produced significantly more extracellular polysaccharide than the wild type . Organic acid excretion and levels of intracellular reduced nucleotides were lowered to wild-type levels in VEM58 and VEM5854, in contrast to those of strain CAR1, which were significantly elevated . Proteome analysis of VEM58 showed a drastic alteration of protein expression, including the absence of a protein identified as PhaB . We propose that the aniA gene product plays an important role in directing carbon flow in R . etli.
Hydrogen-Oxidizing Capabilities of Helicobacter hepaticus and In Vivo Availability of the Substrate. Robert J. Maier, 2003.Hydrogen-oxidizing hydrogenase activity was detected in Helicobacter hepaticus and compared to the activity in Helicobacter pylori for characteristics associated with hydrogen uptake respiratory hydrogenases . Intact whole cells could couple H2 oxidation to oxygen uptake, and no H2 uptake was observed without oxygen available to complete the respiratory pathway . The H . hepaticus enzyme coupled H2 oxidation to reduction of many positive potential acceptors, and it underwent anaerobic or reductive activation . H . hepaticus had a strong affinity for molecular H2 (apparent Km of 2.5 µM), and microelectrode measurements on the livers of live mice demonstrated that H2 is available in the host tissue at levels 20-fold greater than the apparent whole-cell Km value . Identification and Application of Plasmids Suitable for Transfer of Foreign DNA to Members of the Genus Gordonia. Matthias Arenskötter, 2003.Gene transfer systems for Gordonia polyisoprenivorans strains VH2 and Y2K based on electroporation and conjugation, respectively, were established . Several parameters were optimized, resulting in transformation efficiencies of >4 x 105 CFU/µg of plasmid DNA . In contrast to most previously described electroporation protocols, the highest efficiencies were obtained by applying a heat shock after the intrinsic electroporation . Under these conditions, transfer and autonomous replication of plasmid pNC9503 was also demonstrated to proceed in G . alkanivorans DSM44187, G . nitida DSM44499T, G . rubropertincta DSM43197T, G . rubropertincta DSM46038, and G . terrae DSM43249T . Conjugational plasmid DNA transfer to G . polyisoprenivorans resulted in transfer frequencies of up to 5 x 10-6 of the recipient cells . Recombinant strains capable of polyhydroxyalkanoate synthesis from alkanes were constructed .
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