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At least 811 records · Page 45Linked to original sources
PubMed1993

Insertional inactivation of genes to isolate mutants of Synechococcus sp. strain PCC 7942: isolation of filamentous strains.

We have developed a simple procedure for generating mutants of the cyanobacterium Synechococcus sp. strain PCC 7942 in which the site of the lesion can be readily identified. This procedure involves transforming Synechococcus sp. strain PCC 7942 with a library of its own DNA that was fully digested with Sau3A and ligated into the plasmid vector pUC8. The homologous integration of the recombinant plasmid into the genome will often result in the disruption of a gene and the loss of gene function. We have used this method to generate many mutants of Synechococcus sp. strain PCC 7942 which grow as multicellular filaments rather than as unicells. Since the gene harboring the lesion was tagged with pUC8, it was easily isolated. In this paper, we discuss the usefulness of this procedure for the generation of mutants, and we characterize one mutant in which the lesion may be in an operon involved in the assembly of lipopolysaccharides.

Amino Acid Sequence↗
PubMed1994

Selection of candidate quality control isolates and tentative quality control ranges for in vitro susceptibility testing of yeast isolates by National Committee for Clinical Laboratory Standards proposed standard methods.

The National Committee for Clinical Laboratory Standards has developed a proposed standard method for in vitro antifungal susceptibility testing of yeast isolates (National Committee for Clinical Laboratory Standards, document M27-P, 1992). In order for antifungal testing by the M27-P method to be accepted, reliable quality control (QC) performance criteria must be developed. In the present study, five laboratories tested 10 candidate QC strains 20 times each against three antifungal agents: amphotericin B, fluconazole, and 5-fluorocytosine. All sites conformed to the M27-P standards and used a common lot of tube dilution reagents and RPMI 1640 broth medium. Overall, 98% of MIC results with amphotericin B, 95% with fluconazole, and 92% with 5-fluorocytosine fell within the desired 3-log2 dilution range (mode +/- 1 log2 dilution). Excellent performance with all three antifungal agents was observed for six strains: Candida albicans ATCC 90028, Candida parapsilosis ATCC 90018, C. parapsilosis ATCC 22019, Candida krusei ATCC 6258, Candida tropicalis ATCC 750, and Saccharomyces cerevisiae ATCC 9763. With these strains, 3-log2 dilution ranges encompassing 94 to 100% of MICs for all three drugs were established. Additional studies with multiple lots of RPMI 1640 test medium will be required to establish definitive QC ranges.

Amphotericin B↗
PubMed2002

Prospective evaluation of the new chromogenic medium Candida ID, in comparison with Candiselect, for isolation of molds and isolation and presumptive identification of yeast species.

We conducted a prospective evaluation of Candida ID chromogenic medium (bioMérieux, Marcy l'Etoile, France) with 786 clinical specimens in comparison with Candiselect medium (Bio-Rad, Marnes la Coquette, France). Candida ID chromogenic medium identified 97.7% of Candida albicans strains; enabled presumptive identification of C. tropicalis, C. lusitaniae, C. guillermondii, and C. kefyr and better detection of yeast combinations (11.4% more often); and was more sensitive for the isolation of filamentous fungi (17.7% more often). However, Candida ID chromogenic medium appeared to be less selective vis-à-vis bacteria, with bacterial colonies sometimes pigmented blue.

Candida↗
PubMed2003

Rapid detection of rifampin resistance in Mycobacterium tuberculosis isolates by heteroduplex analysis and determination of rifamycin cross-resistance in rifampin-resistant isolates.

Direct heteroduplex analysis and a universal heteroduplex generator assay were performed to detect rifampin resistance rapidly in Turkish Mycobacterium tuberculosis isolates. Cross-resistance to rifapentine, rifabutin, and rifalazil was investigated. A relationship between specific mutations and resistance patterns, which can guide the choice of an appropriate therapeutic regimen for tuberculosis patients, was identified.

Antibiotics, Antitubercular↗
PubMed1996

Neutralization serotypes of human immunodeficiency virus type 1 field isolates are not predicted by genetic subtype. The WHO Network for HIV Isolation and Characterization.

Human immunodeficiency virus type 1 (HIV-1) primary isolates from four geographical locations in Thailand, Brazil, Rwanda, and Uganda, representing genetic subtypes A, B, C, D, and E, were examined for autologous and heterologous neutralization by panels of human HIV+ polyclonal plasma. In independent linked experiments in three laboratories using diverse methodologies and common reagents, no defined pattern of genetic subtype-specific neutralization was observed. Most plasma tested were broadly cross-neutralizing across two or more genetic subtypes, although the titer of neutralization varied across a wide range. We conclude that the genetic subtypes of HIV-1 are not classical neutralization serotypes.

HIV Infections↗
PubMed1999

Mutagenesis of CXCR4 identifies important domains for human immunodeficiency virus type 1 X4 isolate envelope-mediated membrane fusion and virus entry and reveals cryptic coreceptor activity for R5 isolates.

CXCR4 is a chemokine receptor and a coreceptor for T-cell-line-tropic (X4) and dual-tropic (R5X4) human immunodeficiency virus type 1 (HIV-1) isolates. Cells coexpressing CXCR4 and CD4 will fuse with appropriate HIV-1 envelope glycoprotein (Env)-expressing cells. The delineation of the critical regions involved in the interactions within the Env-CD4-coreceptor complex are presently under intensive investigation, and the use of chimeras of coreceptor molecules has provided valuable information. To define these regions in greater detail, we have employed a strategy involving alanine-scanning mutagenesis of the extracellular domains of CXCR4 coupled with a highly sensitive reporter gene assay for HIV-1 Env-mediated membrane fusion. Using a panel of 41 different CXCR4 mutants, we have identified several charged residues that appear important for coreceptor activity for X4 Envs; the mutations E15A (in which the glutamic acid residue at position 15 is replaced by alanine) and E32A in the N terminus, D97A in extracellular loop 1 (ecl-1), and R188A in ecl-2 impaired coreceptor activity for X4 and R5X4 Envs. In addition, substitution of alanine for any of the four extracellular cysteines alone resulted in conformational changes of various degrees, while mutants with paired cysteine deletions partially retained their structure. Our data support the notion that all four cysteines are involved in disulfide bond formation. We have also identified substitutions which greatly enhance or convert CXCR4's coreceptor activity to support R5 Env-mediated fusion (N11A, R30A, D187A, and D193A), and together our data suggest the presence of conserved extracellular elements, common to both CXCR4 and CCR5, involved in their coreceptor activities. These data will help us to better detail the CXCR4 structural requirements exhibited by different HIV-1 strains and will direct further mutagenesis efforts aimed at better defining the domains in CXCR4 involved in the HIV-1 Env-mediated fusion process.

3T3 Cells↗
PubMed1978

Type C oncornavirus isolation studies in systemic lupus erythematosus. III. Isolation of a putative retravirus by triple cell fusion.

A putative retravirus was isolated from a cell culture derived by fusion of mink and dog cell lines with cultured placental cells from a patient with systemic lupus erythematosus. The virus was detected by reverse transcriptase assay of supernatant medium after prolonged subculturing. Its characterisation, including species of origin, is in progress, but it is probably a new type C virus. Its role, if any, in lupus erythematosus in unknown.

Cell Line↗
PubMed1998

Physiological effects of a pea protein isolate in gnotobiotic rats: comparison with a soybean isolate and meat.

Pea proteins have been considered for the introduction into the human diet only recently. This protein source was tested on nutritional and digestive parameters in heteroxenic male Fischer rats inoculated with a human faecal microflora from a methane producer. Compared to soybean proteins, pea proteins have similar effects on the rat's endogenous and bacterial digestive patterns. Compared to the pea proteins, a diet containing a standard meat meal enhanced the pH and the production of ammonia, while a lyophilized beef meat enhanced that of urea. The diet containing the standard meat decreases short-chain fatty acids and modifies the ratio of caecal short-chain fatty acids. Both animal diets decreased the specific activities of pancreatic proteases such as chymotrypsin (EC 3.4.21.1), trypsin (EC 3.4.21.4), and carboxypeptidase A (EC 3.4.17.1) when compared to the diet containing the pea isolate. In conclusion, the whole composition of the diet, more than the origin of the dietary protein, influences the rat's digestive pattern.

Animal Nutritional Physiological Phenomena↗
PubMed1991

Genomic heterogeneities in bovine herpesvirus type 1 viral isolates: a major variant selected from a field isolate.

We have analyzed viral DNA patterns, obtained following digestion of six different infectious bovine rhinotracheitis isolates, using a panel of restriction enzymes. Several differences in profiles were observed, particularly for a Canadian viral strain which presumably contained a genome that was larger than the other five infectious bovine rhinotracheitis genomes. Also, an insertion/deletion of a HindIII restriction site was found in the DNA of a Danish strain; this additional HindIII site was localized precisely within the original HindIII E fragment of the infectious bovine rhinotracheitis virus. Interestingly, results could be used to distinguish each viral strain examined from the others by simple digestion with the appropriate enzyme.

DNA, Viral↗
PubMed2005

Efficacy of repeat isolated limb perfusions with tumor necrosis factor alpha and melphalan for multiple in-transit metastases in patients with prior isolated limb perfusion failure.

BACKGROUND: Isolated limb perfusion (ILP) is an effective treatment modality for multiple in-transit melanoma metastases confined to the limb. Recurrences after ILP, however, occur in approximately 50% of patients and are a challenge for further treatment. The efficacy of repeat ILPs to prolong local control in this patient category is evaluated in this article. METHODS: We used a prospective database in a tertiary referral center. Out of 100 tumor necrosis factor (TNF)-based ILPs with TNF and melphalan (TM-ILPs) in melanoma patients between March 1991 and July 2003, 25 repeat ILP procedures were performed in 21 patients in whom prior ILP treatment failed. All patients had bulky and/or numerous lesions and were treated with mild hyperthermic TM-ILP by using 2 to 4 mg of TNF and 10 to 13 mg/L of limb volume for the leg and arm, respectively. RESULTS: The complete response rate was 76%, a partial response occurred in 20%, and no change was recorded in 4%. There was no difference in the complete response rate or local toxicity between first and repeat perfusions. Local recurrence occurred in 72%; the median time to local progression was 14 months. The 5-year survival rate was 47%, which compares favorably with known survival rates of stage IIIA/AB patients. The median follow-up of the patients was 26 months. CONCLUSIONS: Patients who experience treatment failure after previous ILP treatment respond very well to repeat perfusion, and prolonged local control can thus be obtained. The subgroup of patients qualifying for repeat ILP represents a relatively favorable biological behavior of the melanoma.

Adult↗
PubMed1999

Relaxant effects of pyranocoumarin compounds isolated from a Chinese medical plant, Bai-Hua Qian-Hu, on isolated rabbit tracheas and pulmonary arteries.

Qian-Hu is a well-known traditional Chinese medicine used for the treatment of respiratory diseases and pulmonary hypertension. We compared the relaxant effects of pyranocoumarin compounds, including (+)-praeruptorin A (Pra-C), Pd-Ia (=(+/-)-praeruptorin A), pteryxin, peucedanocoumarin II (P-II) and 8-methoxy-psoralen (8-MOP) purified from Bai-Hua Qian-Hu (BQ) in isolated rabbit tracheas and pulmonary arteries. Pra-C, pteryxin and Pd-Ia produced significant relaxant effects in tracheal preparations constricted with 40 mM KCl or 10 microM acetylcholine. The relaxant response to Pra-C, pteryxin or Pd-Ia in preparations constricted with KCl was significantly more potent than that in preparations constricted with acetylcholine. Pra-C, pteryxin or Pd-Ia at a concentration of 30 microM completely relaxed tracheas constricted with 40 mM KCl, whereas P-II at the same concentration showed only partial relaxation. In pulmonary arterial preparations, 8-MOP produced a significant relaxant effect on contractions by 10 microM phenylephrine, without any effect on the contraction by 40 mM KCl. These results suggest that Pd-Ia, pteryxin and Pra-C for their calcium antagonistic action, and 8-MOP for its inhibitory effect on contraction induced by phenylephrine, may be the active principles of BQ for relaxing the smooth muscle of tracheas and pulmonary arteries, and the principle may produce a synergistic effect.

Animals↗
PubMed1976

Isolation of cells that retain differentiated functions in vitro: properties of clonally isolated type II alveolar pneymonocytes.

We have isolated byclonal culture techniques a diploid cell strain (L-2) from adult rat lung. These cells appear to retain differentiated functions that are present in type II alveolar epithelial cells of intact lung. The L-2 cells are diploid, epithelial cells, they contain osmiophilic lamellar bodies in their cytoplasm and they synthesize lecithin by the same de novo pathways as whole lung.

Animals↗
PubMed1982

Isolation and characterization of mucoid and non-mucoid Salmonella oranienburg isolated from pleural effusion of a patient with hemolytic anemia.

Salmonella oranienburg, mucoid and non-mucoid strains, were simultaneously isolated as causative agents from the pleural effusion of a patient with hereditary spherocytosis. The mucoid-typed Salmonella oranienburg was characterized by serological examination, taking advantage of the high frequency of mucoid (M) to non-mucoid (N) variation.

Anti-Bacterial Agents↗
PubMed2005

Adaptive immune responses are dispensable for isolated lymphoid follicle formation: antigen-naive, lymphotoxin-sufficient B lymphocytes drive the formation of mature isolated lymphoid follicles.

Isolated lymphoid follicles (ILFs) are recently appreciated members of the mucosal immune system. The architecture, composition, and inducible nature of these structures indicates that these structures are tertiary lymphoid structures. The process leading to the formation of tertiary lymphoid structures, lymphoid neogenesis, has been observed in a number of inflammatory and autoimmune conditions. Given this association, there is considerable interest in identifying the factors promoting lymphoid neogenesis, and understanding the steps in this process. Using murine ILF formation as a model, we have examined the roles of different cellular sources of lymphotoxin (LT) and the adaptive immune response in lymphoid neogenesis. In this study, we report that, although other cellular sources of LT may supplant B lymphocytes in the formation of immature ILFs (loosely organized clusters of B lymphocytes), LT-sufficient B lymphocytes are required for the progression of immature ILFs to mature ILFs (organized lymphoid aggregates with a follicle-associated epithelium). ILF formation occurs in the absence of T lymphocytes and Ag-specific B lymphocyte responses, and ILF B lymphocytes express elevated levels of LT in the absence of antigenic stimulation. Consistent with a role for chemokines inducing LT expression in Ag-naive B lymphocytes, and a chemokine-driven positive-feedback loop driving mature ILF formation, mature ILFs express elevated levels of B lymphocyte chemoattractant in the absence of Ag-specific B lymphocyte stimulation. These observations indicate that ILFs contain Ag-naive lymphocytes, and suggest that events occurring within ILFs shape subsequent immune responses mediated by these lymphocytes.

Animals↗
PubMed2005

Endo-1,4-beta-xylanase B from Aspergillus cf. niger BCC14405 isolated in Thailand: purification, characterization and gene isolation.

During the screening of xylanolytic enzymes from locally isolated fungi, one strain BCC14405, exhibited high enzyme activity with thermostability. This fugal strain was identified as Aspergillus cf. niger based on its morphological characteristics and internal transcribed spacer (ITS) sequences. An enzyme with xylanolytic activity from BCC14405 was later purified and characterized. It was found to have a molecular mass of ca. 21 kDa, an optimal pH of 5.0, and an optimal temperature of 55 degrees C. When tested using xylan from birchwood, it showed K(m) and V(max) values of 8.9 mg/ml and 11,100 U/mg, respectively. The enzyme was inhibited by CuSO(4) EDTA, and by FeSO(4) The homology of the 20-residue N-terminal protein sequence showed that the enzyme was an endo-1,4-beta-xylanase. The full-length gene encoding endo-1,4-beta-xylanase from BCC14405 was obtained by PCR amplification of its cDNA. The gene contained an open reading frame of 678 bp, encoding a 225 amino acid protein, which was identical to the endo-1,4-a-xylanase B previously identified in A. niger.

Amino Acid Sequence↗
PubMed1987

FK-506, a novel immunosuppressant isolated from a Streptomyces. I. Fermentation, isolation, and physico-chemical and biological characteristics.

FK-506, a novel immunosuppressant, has been isolated from the fermentation broth of Streptomyces tsukubaenis No. 9993 as colorless prism and the molecular formula was determined as C44H69NO12.H2O. The compound suppressed immune responses in vitro and in vivo with mice. This immunosuppressive effect was more potent than that of ciclosporin.

Animals↗
PubMed1988

FR-900520 and FR-900523, novel immunosuppressants isolated from a Streptomyces. II. Fermentation, isolation and physico-chemical and biological characteristics.

FR-900520 and FR-900523, novel neutral macrolide immunosuppressants, were isolated from the cultured broth of Streptomyces hygroscopicus subsp. yakushimaensis No. 7238. Their molecular formulae were determined as C43H69NO12 and C42H67NO12, respectively. The compounds suppressed immune response in vitro. IC50 values of FR-900520 and FR-900523 for mouse mixed lymphocyte reaction were 0.55 nM and 1.6 nM, respectively. FR-900520, the major component, clearly prolonged skin allograft survival in rats.

Animals↗