Segregation of proteinase-negative mutants from heterozygous Candida albicans

The extracellular acidic proteinase (EC 3.4.23.6) produced by Candida albicans has been reported to be a virulence factor. In studying the role of this proteinase in human disease, we determined the optimum conditions for stimulating proteinase production in order to isolate proteinase-negative (Prt-) mutants. We found that in liquid medium containing bovine serum albumin (BSA) as the sole nitrogen source, at pH 4 and 27 degrees C, the sensitivity of proteinase detection was considerably greater than when assayed on BSA agar at 37 degrees C. This observation is due, in part, to temperature sensitivity of proteinase induction. Nitrogen starvation did not induce proteinase. Proteinase production on agar was increased by adding 0.01% yeast extract (YE) to BSA medium. Using BSA + YE agar to isolate mutants, it was discovered that C. albicans ATCC 28366 was heterozygous for a Prt- mutation. Spontaneous Prt- mutants occurred at a frequency of 2 x 10(-3). Ultraviolet light increased the mitotic segregation of Prt- cells to a frequency of 1 x 10(-2). The Prt- phenotype showed a large inoculum effect, Prt- segregants reverted with a high frequency, and the revertants were unstable.     

Adenosine release from isolated rat adipocytes: influence of fat cell concentration and cell size

The release of adenosine by isolated rat adipocytes into the incubation medium was studied in relation to fat cell size and concentration. Incubations were carried out for 60 min at 37 degrees C in Krebs-Ringer bicarbonate-albumin medium containing 6 mM glucose. 2'-Deoxycoformycin was added to inhibit endogenous adenosine deaminase activity (maximal suppression was achieved at 0.8 microM concentration of the inhibitor). The data show that (a) the amount of adenosine released into the medium was similar for the first and second 30-min incubation periods; (b) increasing adipocyte concentration markedly inhibited adenosine release; and (c) large fat cells (volume greater than 300 pl) released significantly more adenosine (per fat cell) into the medium than smaller fat cells (volume less than 180 pl) when incubated at concentrations of less than or equal to 350,000 cells/ml. Above this cell concentration, differences between adenosine release and cell size were not noted. Adenosine release by isolated rat adipocytes appears to be a precisely regulated process which is exquisitly sensitive to the number of fat cells in the incubation medium and, to a certain extent, to the adipocyte size.     

Effect of varying concentrations of linoleic acid on alpha-adrenoceptor responses in spontaneously hypertensive rats

The effect of increased intake of linoleic acid on the alpha-adrenergic system was assessed by safflower oil supplementation to spontaneously hypertensive rats. Linoleic acid-enriched intake at 5%, 15% and 30% by weight of total food intake for 12 wk was associated with a reduction in resting arterial blood pressure, while heart rate and heart to body weight ratios were similar to control group values. A dose-response analysis to norepinephrine bitartrate administered intravenously indicated a significant reduction in the vascular reactivity to this alpha-adrenergic agonist in all groups given linoleic acid. Direct assessment of alpha-adrenoceptor number (Bmax) and affinity (KD) in cardiac sarcolemma with [3H]-prazosin indicated that receptor binding properties were not affected by linoleic acid intake. Our results suggest that short-term linoleic acid supplementation in the established hypertensive state may lower blood pressure through effects upon alpha-adrenergic reactivity in vascular tissue, without associated effects in cardiac tissue.     

Molecular phylogenetics of the burrowing crayfish genus Fallicambarus (Decapoda: Cambaridae)

Ainscough, B.J., Breinholt, J.W., Robison, H.W. & Crandall, K.A. (2013). Molecular phylogenetics of the burrowing crayfish genus Fallicambarus (Decapoda: Cambaridae). —Zoologica Scripta, 42, 306–316. The crayfish genus Fallicambarus contains 19 species of primary burrowing freshwater crayfish divided into two distinct subgenera. We test current hypotheses of the phylogenetic relationships among species within the genus as well as the monophyly of the genus. Our study samples all 19 species for five gene regions (both nuclear and mitochondrial) to estimate a robust phylogenetic hypothesis for the genus. We show that the genus is not a monophyletic group. The subgenus Creaserinus does fall out as a monophyletic group, but distinct from the subgenus Fallicambarus. The subgenus Fallicambarus appears to be monophyletic with the exception of the species Procambarus (Tenuicambarus) tenuis, which falls in the midst of this subgenus suggesting that it might be better classified as a Fallicambarus species. We also show that the species Fallicambarus fodiens is a species complex with distinct evolutionary lineages that are regionalized to different geographic areas.     

Pilocarpine-induced sweat gland function in individuals with multiple sclerosis.

This investigation tested the hypothesis that cholinergic sweat function of individuals with multiple sclerosis (MS) (MS-Con; n = 10) is diminished relative to matched healthy control subjects (Con; n = 10). In addition, cholinergic sweat function was determined before and after 15 wk of aerobic training in a subgroup of individuals with MS (MS-Ex; n = 7). Cholinergic sweating responses were assessed via pilocarpine iontophoresis on ventral forearm skin. A collection disk placed over the stimulated area collected sweat for 15 min. Sweat rate (SR) was calculated by dividing sweat collector volume by collection area and time. Iodine-treated paper was applied to the stimulated area to measure number of activated sweat glands (ASG). Sweat gland output (SGO) was calculated by dividing SR by density of glands under the collector. Sweat gland function was determined in MS-Ex to test the hypothesis that exercise training would increase sweating responses. No differences in ASG were observed between MS-Con and Con. SR and SGO in MS-Con [0.18 mg.cm(-2).min(-1) (SD 0.08); 1.74 microg.gland(-1).min(-1) (SD 0.79), respectively] were significantly lower (P < or = 0.05) than in Con [0.27 mg.cm(-2).min(-1) (SD 0.10); 2.43 microg.gland(-1).min(-1) (SD 0.69)]. Aerobic exercise training significantly (P < or = 0.05) increased peak aerobic capacity in MS-Ex [1.86 (SD 0.75) vs. 2.10 (SD 0.67) l/min] with no changes in ASG, SR, and SGO. Sweat gland function in individuals with MS is impaired relative to healthy controls. Fifteen weeks of aerobic training did not increase stimulated sweating responses in individuals with MS. Diminished peripheral sweating responses may be a consequence of impairments in autonomic control of sudomotor function.     

The Molecular Evolution of Visual Pigments of Freshwater Crayfishes (Decapoda: Cambaridae)

This study examines the diverse maximum wavelength absorption (λ_max) found in crayfishes (Decapoda: Cambaridae and Parastacidae) and the associated genetic variation in their opsin locus. We measured the wavelength absorption in the photoreceptors of six species that inhabit environments of different light intensities (i.e., burrows, streams, standing waters, and subterranean waters). Our results indicate that there is relatively little variation in λ_max (522–530 nm) among species from different genera and families. The existing variation did not correlate with the habitat differences of the crayfishes studied. We simultaneously sequenced the rhodopsin gene to identify the amino acid replacements that affect shifts in maximum wavelength absorption. We then related these to changes that correlated with shifts in λ_max by reconstructing ancestral character states using a maximum-likelihood approach. Using amino acid sequences obtained from five species (all were 301 amino acids in length), we identified a number of candidates for producing shifts of 4 to 8 nm in λ_max. These amino acid replacements occurred in similar regions to those involved in spectral shifts in vertebrates. Received: 12 March 1997 / Accepted: 3 June 1997