Isolation and characterization of filamentous bacteria present in bulking activated sludge

Summary Several types of filamentous microorganisms were observed and identified in samples of poorly settling (bulking) activated sludge. The major types encountered and the frequency (percentage) of appearance in the total of all treatment plants sampled were: Eikelboom type 0041 (60), type 1701 (45), Haliscomenobacter hydrossis (35), type 021N (30), Thiothrix spp. (20), and Sphaerotilus natans (20). Isolation techniques and culture media were developed and used to recover 42 axenic strains of filamentous bacteria from sludge samples collected. The isolates were identified as strains of Thiothrix, Beggiatoa, S. natans, and Eikelboom types 021N, 1701, 0041, and 0803. Nutritional and differential characterization of the bacteria was important to the differentiation of groups which could not be easily distinguished on the basis of morphology. Although certain treatment plant operating parameters (organic loading) seemed associated with the presence of specific filamentous organism types, possible interaction among factors precluded definite establishment of a cause and effect relationship for most of the treatment plant characteristics and organisms observed.     

Human 2-5A synthetase: characterization of a novel cDNA and corresponding gene structure.

The enzyme 2-5A synthetase is induced in cultured cells in response to interferon (IFN) treatment. A lambda gt10 cDNA library of mRNA from IFN-induced Daudi lymphoblastoid cells was screened with oligonucleotide probes. Several overlapping cDNAs were isolated and shown to be derived from the human synthetase gene using filter selection and oocyte microinjection assays. The nucleotide sequence of one of these, cDNA 8-2, extended the 2-5A synthetase sequence already described 72 bp in the 5' direction but was found to differ significantly in coding sequence at the 3' end. The longest cDNA isolated (6-2) was approximately 1.4 kb. By Northern hybridization analysis single mRNAs of 1.7 kb were detected in Daudi and T98G (glioblastoma) cells. However, in HeLa cells, four mRNAs ranging in size from 1.5 to 3.5 kb were found, one of which differed at the 3' end. Analysis of both phage and cosmid genomic clones and comparison with genomic DNA indicate that there is a single gene for 2-5A synthetase, comprising at least six exons and five introns, which can undergo a novel form of alternative RNA processing depending on cell type.     

Sympathetic control of the forearm blood flow in man during brief isometric contractions

Experiments were performed to assess the possible neurally mediated constriction in active skeletal muscle during isometric hand-grip contractions. Forearm blood flow was measured by venous occlusion plethysmography on 5 volunteers who exerted a series of repeated contractions of 4 s duration every 12 s at 60% of their maximum strength of fatigue. The blood flows increased initially, but then remained constant at 20-24 ml X min(-1) X 100 ml(-1) throughout the exercise even though mean arterial blood pressure reached 21-23 kPa (160-170 mm Hg). When the same exercise was performed after arterial infusion of phentolamine, forearm blood flow increased steadily to near maximal levels of 38.7 +/- 1.4 ml X min(-1) X 100 ml(-1). Venous catecholamines, principally norepinephrine, increased throughout exercise, reaching peak values of 983 +/- 258 pg X ml(-1) at fatigue. Of the vasoactive substances measured, the concentration of K+ and osmolarity in venous plasma also increased initially and reached a steady-state during the exercise but ATP increased steadily throughout the exercise. These data indicate a continually increasing alpha-adrenergic constriction to the vascular beds in active muscles in the human forearm during isometric exercise, that is only partially counteracted by vasoactive metabolites.     

Evidence from cDNA clones that the rat leukocyte-common antigen (T200) spans the lipid bilayer and contains a cytoplasmic domain of 80,000 Mr

The leukocyte-common antigen (L-CA or T200) includes a family of lymphoid and myeloid cell surface glycoproteins with apparent molecular weights from 180,000 to 240,000. We report a partial protein sequence for thymocyte L-CA containing 1073 amino acids predicted from cDNA clones isolated using an oligonucleotide probe. Only one segment (residues 347-368) is likely to cross the membrane, and peptide data suggest that sequences N-terminal to this are outside the cell, with residues 369-1073 inside. The cytoplasmic domain includes possible phosphorylation sites and an internal homology between residues 385-671 and 676-986. Analysis of B lymphocyte cDNA clones suggests that B cell and thymocyte mRNAs are identical in 3' sequences, but size differences in Northern blots suggest 5' sequences may differ.     

Protein kinase activity associated with pancreatic zymogen granules.

Purified zymogen granules were prepared from rat pancreas by using an iso-osmotic Percoll gradient. In the presence of [gamma-32P]ATP, phosphorylation of several granule proteins was induced by Ca2+, most notably a Mr-13 000 protein, whereas addition of cyclic AMP was without effect. When phosphatidylserine was also added, Ca2+ increased the phosphorylation of additional proteins, with the largest effect on a protein of Mr 62 000. Purified granules were also able to phosphorylate exogenous substrates. Ca2+-induced phosphorylation of lysine-rich histone was enhanced over 3-fold in the presence of phosphatidylserine, and cyclic AMP-activated protein kinase activity was revealed with mixed histone as substrate. The concentrations of free Ca2+ and cyclic AMP required for half-maximal phosphorylation of both endogenous and exogenous proteins were 1-3 microM and 57 nM respectively. Treatment of granules with 0.25 M-KCl resulted in the release of phosphatidylserine-dependent kinase activity into a high-speed granule supernatant. In contrast, granule-protein substrates of Ca2+-activated kinase activity were resistant to KCl extraction, and in fact were present in purified granule membranes. Kinase activity activated by cyclic AMP was not extracted by KCl treatment. It is concluded that phosphorylation of integral membrane proteins in the zymogen granule can be induced by one or more Ca2+-activated protein kinases. Such a reaction is a potential mechanism by which exocytosis may be regulated in the exocrine pancreas by Ca2+-mediated secretagogues.     

Butyrate formation from glucose by the rumen protozoon Dasytricha ruminantium.

Production of butyrate by the holotrich protozoon Dasytricha ruminantium involves the enzymes of glycolysis, pyruvate:ferredoxin oxidoreductase, acetyl-CoA:acetyl-CoA C-acetyltransferase, 3-hydroxybutyryl-CoA dehydrogenase, 3-hydroxyacyl-CoA hydro-lyase, 3-hydroxyacyl-CoA reductase, phosphate butyryltransferase and butyrate kinase. Subcellular fractionation by differential and density-gradient centrifugation on sucrose gradients indicated that all those enzymes except pyruvate:ferredoxin oxidoreductase were non-sedimentable at 6 X 10(6) g-min. Butyrate kinase and phosphate butyryltransferase were associated with the large- and small-granule fractions. Thus, although metabolic reactions necessary for butyrate production proceed predominantly in the cytosol, hydrogenosomes play a key role in the conversion of pyruvate into acetyl-CoA.     

Male-to-male transmission of the velo-cardio-facial syndrome: a case report and review of 60 cases

The velo-cardio-facial syndrome is one of the most common syndromes of clefting. Previous reports have shown vertical pedigree transmission, but in all cases the gene was maternally transmitted. The genetics of this syndrome had been suspected as autosomal dominant, but X-linked dominant inheritance could not be ruled out. This report describes an instance of male-to-male transmission of the velo-cardio-facial syndrome. In addition, the clinical findings in 60 cases are reported to further delineate the phenotypic spectrum of the syndrome.     

NIH/3T3 cells transfected with human tumor DNA containing activated ras oncogenes express the metastatic phenotype in nude mice.

NIH/3T3 cells transfected with DNA from malignant human tumors produced experimental and spontaneous metastases in nude mice. In contrast, parent or spontaneously transformed NIH/3T3 cells failed to metastasize. The transfected clones contained either activated c-Harvey-ras or N-ras oncogenes. A representative clone (T71-17SA2) which was used to assess selected cellular and host factors relevant to the metastatic process produced lung metastases in 100% of the NIH nude mice recipients, secreted augmented levels of type IV collagenase, and invaded human amnion basement membrane in vitro. Expression of the metastatic phenotype was not related to decreased sensitivity to natural killer cells or macrophage-mediated cytotoxicity. Analysis of the cellular DNA from the T71-17SA2 transfectant and its corresponding metastases, both of which contained activated N-ras oncogenes, revealed a twofold increase in the N-ras-specific DNA sequences in the metastatic cells. Thus, transfection with human tumor DNA containing activated ras oncogenes can induce the complete metastatic phenotype in NIH/3T3 cells by a mechanism apparently unrelated to immune cell killing.     

Inheritance of extrachromosomal ribosomal DNA during the asexual life cycle of Dictyostelium discoideum: examination by use of DNA polymorphisms.

Wild-type isolates of Dictyostelium discoideum exhibited differences in the size of restriction fragments of the extrachromosomal 88-kilobase ribosomal DNA (rDNA) palindrome. Polymorphisms in rDNA also were found among strains derived solely from the NC4 wild-type isolate. These variations involved EcoRI fragments II, III, and V; they included loss of the EcoRI site separating fragments II and V and deletion and insertion of DNA. More than one rDNA form can coexist in the same diploid or haploid cell. However, one or another parental rDNA tended to predominate in diploids constructed, using the parasexual cycle, between haploid NC4-derived strains and haploid wild-type isolates. In some cases, most if not all of the rDNA of such diploids were of one form after ca. 50 generations of growth. Segregant haploids, derived from diploids that possessed predominantly a single rDNA allele, possessed the same allele as the diploid and did not recover the other form. This evidence implies that replication does not proceed from a single chromosomal or extrachromosomal copy of the rDNA during the asexual life cycle of D. discoideum.