Improved medium for recovery and enumeration of Pseudomonas aeruginosa from water using membrane filters.
A modified mPA medium, designated mPA-C, was shown to recover Pseudomonas aeruginosa from a variety of water sources with results comparable to those with mPA-B and within the confidence limits of a most-probable-number technique. Enumeration of P. aeruginosa on mPA-C was possible after only 24 h of incubation at 41.5 degrees C, compared with 72 h of incubation required for mPA-B and 96 h of incubation for a presumptive most probable number. (+info)
Automated food microbiology: potential for the hydrophobic grid-membrane filter.
Bacterial counts obtained on hydrophobic grid-membrane filters were comparable to conventional plate counts for Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus in homogenates from a range of foods. The wide numerical operating range of the hydrophobic grid-membrane filters allowed sequential diluting to be reduced or even eliminated, making them attractive as components in automated systems of analysis. Food debris could be rinsed completely from the unincubated hydrophobic grid-membrane filter surface without affecting the subsequent count, thus eliminating the possibility of counting food particles, a common source of error in electronic counting systems. (+info)
Cytokine removal during continuous hemofiltration in septic patients.
A potential application of the continuous renal replacement therapies is the extracorporeal removal of inflammatory mediators in septic patients. Cytokine elimination with continuous renal replacement therapies has been demonstrated in several clinical studies, but so far without important effects on their serum concentrations. Improved knowledge of the cytokine removal mechanisms could lead to the development of more efficient treatment strategies. In the present study, 15 patients with septic shock and acute renal failure were observed during the first 24 h of treatment with continuous venovenous hemofiltration (CVVH) with an AN69 membrane. After 12 h, the hemofilter was replaced and the blood flow rate (QB) was switched from 100 ml/min to 200 ml/min or vice versa. Pre- and postfilter plasma and ultrafiltrate concentrations of selected inflammatory and anti-inflammatory cytokines were measured at several time points allowing the calculation of a mass balance. Cytokine removal was highest 1 h after the start of CVVH and after the change of the membrane (ranging from 25 to 43% of the prefilter amount), corresponding with a significant fall in the serum concentration of all cytokines. The inhibitors of inflammation were removed to the same extent as the inflammatory cytokines. Adsorption to the AN69 membrane appeared to be the main clearance mechanism, being most pronounced immediately after installation of a new membrane and decreasing steadily thereafter, indicating rapid saturation of the membrane. A QB of 200 ml/min was associated with a 75% increase of the ultrafiltration rate and a significantly higher convective elimination and membrane adsorption than at a QB of 100 ml/min. The results indicate that optimal cytokine removal with CVVH with an AN69 membrane could be achieved with a combination of a high QB/ultrafiltration rate and frequent membrane changes. (+info)
Effect of leukocytapheresis therapy using a leukocyte removal filter in Crohn's disease.
Eighteen patients with active Crohn's disease were treated with one leukocytapheresis session per week for a five-week intensive therapy, decreasing to one leukocytapheresis session per month for five sessions of initial maintenance therapy. Nutritional indices, inflammatory reactions, flow cytometry profiles, and cytokine production were also assessed before and after the intensive and initial maintenance therapy. Nine of the patients (50%) attained remission at the end of the intensive therapy. The nine non-remission patients had exhibited longer periods of suffering and more severely affected sites prior to the therapy. In 14 of 18 patients (77.8%), the nutritional indices, Internal Organization of Inflammatory Bowel Disease (IOIBD) score and Crohn's Disease Activity Index (CDAI) improved from the pretherapy levels, but only the remission group (50%) showed improvement in C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR). The remission group showed significantly higher pretherapy CD4+ CD45+ cell ratios and interleukin-2 (IL-2) production than the non-remission group, and significantly lower activated cells. (+info)
Transduction-like gene transfer in the methanogen Methanococcus voltae.
Strain PS of Methanococcus voltae (a methanogenic, anaerobic archaebacterium) was shown to generate spontaneously 4.4-kbp chromosomal DNA fragments that are fully protected from DNase and that, upon contact with a cell, transform it genetically. This activity, here called VTA (voltae transfer agent), affects all markers tested: three different auxotrophies (histidine, purine, and cobalamin) and resistance to BES (2-bromoethanesulfonate, an inhibitor of methanogenesis). VTA was most effectively prepared by culture filtration. This process disrupted a fraction of the M. voltae cells (which have only an S-layer covering their cytoplasmic membrane). VTA was rapidly inactivated upon storage. VTA particles were present in cultures at concentrations of approximately two per cell. Gene transfer activity varied from a minimum of 2 x 10(-5) (BES resistance) to a maximum of 10(-3) (histidine independence) per donor cell. Very little VTA was found free in culture supernatants. The phenomenon is functionally similar to generalized transduction, but there is no evidence, for the time being, of intrinsically viral (i.e., containing a complete viral genome) particles. Consideration of VTA DNA size makes the existence of such viral particles unlikely. If they exist, they must be relatively few in number;perhaps they differ from VTA particles in size and other properties and thus escaped detection. Digestion of VTA DNA with the AluI restriction enzyme suggests that it is a random sample of the bacterial DNA, except for a 0.9-kbp sequence which is amplified relative to the rest of the bacterial chromosome. A VTA-sized DNA fraction was demonstrated in a few other isolates of M. voltae. (+info)
Virus passage through track-etch membranes modified by salinity and a nonionic surfactant.
Why do viruses sometimes not pass through larger pores in track-etch filters? Increasing the salinity (0.8 to 160 mM Na+) decreased phiX174 and PRD1 passage through track-etch polycarbonate membranes (sodium dodecyl sulfate coated but not polyvinylpyrrolidone coated) and PRD1 passage through polyester membranes. Undiminished passage when 0.1% Tween 80 was added implied that nonionic virus adsorption occurred and indicated that high levels of salinity decreased virus passage by decreasing electrostatic repulsion that prevented adsorption. (+info)
A cell-free assay allows reconstitution of Vps33p-dependent transport to the yeast vacuole/lysosome.
We report a cell-free system that measures transport-coupled maturation of carboxypeptidase Y (CPY). Yeast spheroplasts are lysed by extrusion through polycarbonate filters. After differential centrifugation, a 125,000-g pellet is enriched for radiolabeled proCPY and is used as "donor" membranes. A 15,000-g pellet, harvested from nonradiolabeled cells and enriched for vacuoles, is used as "acceptor" membranes. When these membranes are incubated together with ATP and cytosolic extracts, approximately 50% of the radiolabeled proCPY is processed to mature CPY. Maturation was inhibited by dilution of donor and acceptor membranes during incubation, showed a 15-min lag period, and was temperature sensitive. Efficient proCPY maturation was possible when donor membranes were from a yeast strain deleted for the PEP4 gene (which encodes the principal CPY processing enzyme, proteinase A) and acceptor membranes from a PEP4 yeast strain, indicating intercompartmental transfer. Cytosol made from a yeast strain deleted for the VPS33 gene was less efficient at driving transport. Moreover, antibodies against Vps33p (a Sec1 homologue) and Vam3p (a Q-SNARE) inhibited transport >90%. Cytosolic extracts from yeast cells overexpressing Vps33p restored transport to antibody-inhibited assays. This cell-free system has allowed the demonstration of reconstituted intercompartmental transport coupled to the function of a VPS gene product. (+info)
Effect of methodology on the tuberculocidal activity of a glutaraldehyde-based disinfectant.
Although official guidelines recommend a plate counting method for testing the susceptibility of mycobacteria to disinfectants, manufacturers usually prefer to employ the BACTEC procedure. Data showing that the BACTEC method overestimates the activity of a glutaraldehyde-based disinfectant against Mycobacterium tuberculosis in comparison with a conventional plate counting procedure are presented. (+info)