Sequence variations in small-subunit ribosomal RNAs of Hartmannella vermiformis and their phylogenetic implications

Evidence of associations between free-living amoebas and human disease has been increasing in recent years. Knowledge about phylogenetic relationships that may be important for the understanding of pathogenicity in the genera involved is very limited at present. Consequently, we have begun to study these relationships and report here on the phylogeny of Hartmannella vermiformis, a free-living amoeba that can harbor the etiologic agent of Legionnaires' disease. Our analysis is based on studies of small-subunit ribosomal RNA genes (srDNA). Nucleotide sequences were determined for nuclear srDNA from three strains of H. vermiformis isolated from the United Kingdom, Germany, and the United States. These sequences then were compared with a sequence previously obtained for a North American isolate by J. H. Gunderson and M. L. Sogin. The four genes are 1,840 bp long, with an average GC content of 49.6%. Sequence differences among the strains range are 0.38%-0.76%. Variation occurs at 19 positions and includes 2 single-base indels plus 14 monotypic and 3 ditypic single-base substitutions. Variation is limited to eight helix/loop structures according to a current model for srRNA secondary structure. Parsimony, distance, and bootstrap analyses used to examine phylogenetic relationships between the srDNA sequences of H. vermiformis and other eukaryotes indicated that Hartmannella sequences were most closely related to those of Acanthamoeba and the alga Cryptomonas. All ditypic sites were consistent with a separation between European and North American strains of Hartmannella, but results of other tests of this relationship were statistically inconclusive.      

Identification of different sites of expression for spo loci by transformation of Bacillus subtilis.

Asporogenous mutants of Bacillus subtilis were rendered capable of forming heat-resistant spores by transformation with wild-type (spo+) DNA at, or near, the start of sporulation. For several mutants up to about 50% of the colonies derived from heat-resistant spores, formed as a result of the transformation, remained genetically asporogenous (spo). This was thought to indicate that the genome of the mother cell, but not that of the forespore, was transformed to spo+ and that correct expression of the spo locus in the mother cell was sufficient for spore formation. At the end of the process the mother cell was destroyed, leaving a mature heat-resistant spore that was genetically asporogenous. It is concluded that the loci spoIIID, spIVA, spoVB and spoVE are expressed in the mother cell. For one mutant more than 99% of the colonies derived from heat-resistant spores were genetically spo+. It is concluded that the locus involved, spoVA, had to be expressed in the forespore. Thus different sporulation-specific loci are expressed in the mother cell and in the forespore. The loci expressed in the mother cell are expressed in one cell type so that another cell type, the forespore, can develop into a heat-resistant spore. Other unselected donor markers could be introduced into the recipient during transformation provided high concentrations of DNA were used. The frequency of congression was the same for spo survivors as for spo+ survivors. This implies that there was no correlation between the DNA strand into which the selected spo+ and the unselected donor markers integrated.     

Ouabain-insensitive salt and water movements in duck red cells. I. Kinetics of cation transport under hypertonic conditions

Duck red cells in hypertonic media experience rapid osmotic shrinkage followed by gradual reswelling back toward their original volume. This uptake of salt and water is self limiting and demands a specific ionic composition of the external solution. Although ouabain (10(-4)M) alters the pattern of cation accumulation from predominantly potassium to sodium, it does not affect the rate of the reaction, or the total amount of salt or water taken up. To study the response without the complications of active Na-K transport, ouabain was added to most incubations. All water accumulated by the cells can be accounted for by net salt uptake. Specific external cation requirements for reswelling include: sufficient sodium (more than 23 mM), and elevated potassium (more than 7 mM). In the absence of external potassium cells lose potassium without gaining sodium and continue to shrink instead of reswelling. Adding rubidium to the potassium- free solution promotes an even greater loss of cell potassium, yet causes swelling due to a net uptake of sodium and rubidium followed by chloride. The diuretic furosemide (10(-3)M) inhibits net sodium uptake which depends on potassium (or rubidium), as well as inhibits net sodium uptake which depends on sodium. As a result, cell volume is stabilized in the presence of this drug by inhibition of shrinkage, at low, and of swelling at high external potassium. The response has a high apparent energy of activation (15-20 kcal/mol). We propose that net salt and water movements in hypertonic solutions containing ouabain are mediated by direct coupling or cis-interaction, between sodium and potassium so that the uphill movement of one is driven by the downhill movement of the other in the same direction.     

Isolation and characterization of a recombinant plasmid carrying a functional part of the Bacillus subtilis spoIIA locus

Plasmid pHM2 consists of a 3.3 kb insert of Bacillus subtilis DNA in the chimeric plasmid pHV33, and can replicate in Escherichia coli and B. subtilis. In B. subtilis, pHM2 complements the defects resulting from mutations spo-42, spo-50, spo-69 and sas-1 in the spoIIA locus. This complementation can occur in recE4 strains where recombination of the plasmid with the chromosome is prevented, and the chromosome retains the mutant allele. Thus the plasmid carries a functional part of the spoIIA locus; it does not contain the complete locus as it cannot complement several other spoIIA mutations. It is likely that the locus is complex, containing at least two genes.     

Bex,the Bacillus subtilis homolog of the essential Escherichia coli GTPase Era,is required for normal cell division and spore formation

The Bacillus subtilis bex gene complemented the defect in an Escherichia coli era mutant. The Bex protein showed 39 percent identity and 67 percent similarity to the E. coli Era GTPase. In contrast to era, bex was not essential in all strains. bex mutant cells were elongated and filled with diffuse nucleoid material. They grew slowly and exhibited severely impaired spore formation.     

The involvement of interleukin-1 and interleukin-4 in the response of human annulus fibrosus cells to cyclic tensile strain: an altered mechanotransduction pathway with degeneration

Introduction  

Recent evidence suggests that intervertebral disc (IVD) cells derived from degenerative tissue are unable to respond to physiologically relevant mechanical stimuli in the 'normal' anabolic manner, but instead respond by increasing matrix catabolism. Understanding the nature of the biological processes which allow disc cells to sense and respond to mechanical stimuli (a process termed 'mechanotransduction') is important to ascertain whether these signalling pathways differ with disease. The aim here was to investigate the involvement of interleukin (IL)-1 and IL-4 in the response of annulus fibrosus (AF) cells derived from nondegenerative and degenerative tissue to cyclic tensile strain to determine whether cytokine involvement differed with IVD degeneration.     

Avian biodiversity in the coastal wetlands of the Okahukura Peninsula

The Tapora Landcare Group, operating on the Okahukura Peninsula, has the long-term goal of making this region predator fenced. The aim of this study was to obtain information on the current status of avian biodiversity and the bird community across the band of coastal wetlands on the Okahukura Peninsula. Bird counts were conducted and playback lures used to detect three cryptic wetland species: fernbirds (Bowdleria punctata); spotless crakes (Porzana tabuensis); and banded rails (Gallirallus philippensis). Fernbirds and banded rails were detected at seven of the eight wetland sites sampled whereas spotless crakes were detected at two sites. The native species with the highest relative abundance across the eight sites were silvereyes (Zosterops lateralis) and South Island pied oystercatchers (Haematopus finschi). Changes in avian biodiversity over time in the region can now be monitored, and comprehensive long-term data on the status of avian biodiversity over time obtained.