Characterization of groups of the zygomycete genusRhizopus

Rhizopus is a zygomycetous genus. Several species of this taxon may infect humans and lower animals. Seventeen isolates ofRhizopus species in three distinct morphological groups were studied: the stolonifer group (sporangiophores greater than 1 mm in height, sporangial diameters of 100–275 µm, branched rhizoids); the arrhizus group (sporangiophores greater than 1 mm in height, branched rhizoids, sporangial diameters of 100–240 µm); and the microsporus group (sporangiophores less than 0.8 mm in height, sporangial diameters less than 100 µm, simple rhizoids). Maximal growth temperatures were characteristic: the stolonifer group grew at 30°C, the arrhizus group grew at 36°C, and the microsporus group grew at 45°C. The DNA mol% G + C base composition of all isolates ranged from 34.9 to 40.2% Species within the three groups were grouped by DNA differences. The arrhizus group was most distinctive with a value of 34.9–36.3%; the stolonifer and microsporus groups had G + C values of 37.0–39.3% and 37.8–40.2%, respectively. Our research clarifies and defines the G-C values of the three important groups ofRhizopus species.     

Substitutions of the Conserved Thr42 Increased the Roles of the ε-Subunit of Maize CF1 as CF1 Inhibitor and Proton Gate

The conserved residue Thr42 of -subunit of the chloroplast ATP synthase of maize (Zea mays L.) was substituted with Cys, Arg, and Ile, respectively, through site-directed mutagenesis. The over-expressed and refolded -proteins were purified by chromatography on DEAE-cellulose and FPLC on mono-Q column, which were as biologically active (inhibiting Ca²⁺-ATPase activity and blocking proton gate) as the native subunit isolated from chloroplasts. The T42C and T42R showed higher inhibitory activities on the soluble CF₁(–) Ca²⁺-ATPase than the WT. The T42I inhibited the Ca²⁺-ATPase activity of soluble CF₁ and restored photophosphorylation activity of membrane-bound CF₁ deficient in the most efficiently. Far-ultraviolet CD spectra showed that the portions of -helix and -sheet structures of the three mutants were somewhat different from WT. Thus the conserved residue Thr42 may be important for maintaining the structure and function of the -subunit and the basic functions of the -subunit as far as an inhibitor of Ca²⁺-ATPase and the proton gate are related.     

A new metabolic pathway from n-dodecane to α, ω-dodecanedioic acid in a mutant of Candida tropicalis

Summary The metabolic formation of ,-dodecanedioic acid via ,-dodecanediol from n-dodecane using a mutant S76 of Candida tropicalis was studied.It was found that resting cells of S76 produce ,-dodecanediol from n-dodecane. This intermediate was identified by different analytical methods. With n-dodecanol as substrate the quantitative changes in the concentrations of ,-dodecanediol as well as other intermediates, e.g. monoacid, -hydroxy acid and ,-dioic acid produced by resting cells of S76 for different periods of time were determined. With ,-dodecanediol as the sole carbon source, quantitative changes of -hydroxy acid and ,-dioic acid produced by S76 were also recorded.The results confirm the existence of a new metabolic pathway via ,-diol in the course of ,-dioic acid formation from n-alkane in the mutant S76 of C. tropicalis.     

Phytoplankton dynamics and sedimentation processes during spring and summer in Balsfjord,Northern Norway

Summary Chlorophyll a, phytoplankton species composition and carbon (PPC) estimated from cell-counts, were monitored together with hydrographic parameters and nutrients in the upper 50 m of Balsfjord (ca. 70° N), northern Norway between 08 February and 29 June 1982. Sediment traps were placed at 10, 50, 100, and 170 m (10 m above bottom) for intervals of 5–20 days during the study period. Trap contents were analyzed for phytoplankton as above; dry weight, particulate organic material (POM), particulate organic nitrogen and carbon (PON and POC), ash, and particulate phosphorus were also measured. The phytoplankton community exhibited three main phases: During the first (02–15 April, chiefly surface biomass) and the second (20 April–10 May, deep biomass-maximum and spring bloom peak) periods, Phaeocystis pouchetii dominated biomass (ca. 50% of PPC) followed by vegetative cells of Chaetoceros socialis. In the third period (10 May onwards, characterized by surface estuarinecir-culation), dino- and microflagellates dominated the low post-bloom biomass. Protozooplankton comprising tintinnids, other ciliates and heterotrophic dinoflagellates increased in abundance. Vegetative cells of phytoplankton were scarce in trap collections at 50 m or below; resting cells of Chaetoceros comprised nearly all the intact sedimenting phytoplankton. Krill faeces accounted for >90% by volume of the total faecal material trapped, despite a >21 biomass dominance of copepods in the fjord. The greatest sedimentation rates of krill faeces were at > 100 m, reflecting the downward migration of krill during the day. In all, 2–3 g Cm^–2 of krill faeces were collected, representing ca. twice that from intact phytoplankton cells. POC in the traps at 50 m was ca. 11 gm^–2, accounting for ca. 17% of the estimated primary production during the study period. As the secondary production is high, a large proportion of the production of P. pouchetii must be grazed by herbivores. Copepod faeces are probably remineralized in the euphotic zone, while those of krill provide the major coupling between the pelagial and the benthos. The implications of such a sedimentation model for partitioning energy flow between the pelagial and the benthos is discussed.     

Structure and properties of aCephalosporium acremonium α-galactosidase

The amino acid and sugar composition of the enzyme protein, the effect of urea, sodium dodecyl sulphate and Concanavalin A on the purified -galactosidase (EC 3.2.1.22) from the moldCephalosporium acremonium has been studied. The results obtained by gas liquid chromatography indicated the presence ofN-acetylglucosamine, mannose, galactose andN-acetylneuramic acid in the molar proportions 27311. The presence of two types of Asn-linked oligosaccharide structures in the enzyme molecule is assumed. The -galactosidase liberates (1–3), (1–4) and (1–6)-linkedd-galactose units from various synthetic and natural substrates which have been tested. The effects of pH, substrate concentration and temperature on the catalytic activity of the enzyme are described. The purified -galactosidase also exhibited a lectin activity with an affinity towards glucose, and to some extent mannose.Abbreviations p-NPG p-nitrophenyl--d-galactopyranoside - 4-MUG 4-methylumbelliferyl--d-galactopyranoside - HU hemagglutinin unit - PBS phosphate buffered saline - SDS sodium dodecyl sulphate - ConA Concanavalin A - WGA wheat germ agglutinin - LCA Lens culinaris agglutinin - PHA phytohemagglutinin fromPhaseolus vulgaris     

The contribution of size-fractionated plankton to biomass and primary production of phytoplankton in saline–alkaline ponds

Primary productivity, biomass and chlorophyll-a of size fractionated phytoplankton (<0.22 m, <3 m, <8 m, <10 m, <40 m, <64 m, <112 m and <200 m) were estimated in 6 ponds and 5 experimental enclosures. The results showed that the planktonic algae less than 10 m are important in the biomass and production of phytoplankton in saline–alkaline ponds. The production of size fractionated phytoplankton corresponding to <112 m, <10 m and <3 m in saline–alkaline ponds were 10.5 ± 6.6 , 8.6 ± 5.4 and 0.33 ± 0.1 mgC l^–1 d^–1, respectively. Mean community respiration rate was 1.80 ± 0.73, 1.69 ± 0.90 and 1.38 ± 1.12 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) were 1.61, 8.30 and 0.33 mgC l^–1 d^–1, respectively. The ratio of those to the total phytoplankton production was 15%, 79% and 3%, respectively. The mean respiration rate of the different size groups was 0.11, 0.31 and 1.38 mgC l^–1 d^–1; the ratio of those to total respiration of phytoplankton was 6%, 17% and 77%, respectively. The production of size-fractionated phytoplankton corresponding to <200 m, <10 m and <3 m in enclosures was 2.19 ± 1.63, 2.08 ± 1.75 and 0.22 ± 0.08 mgC l^–1 d^-1, respectively. Mean community respiration rates were 1.25 ± 1.55, 1.17 ± 1.42 and 0.47 ± 0.32 mgC l^–1 d^–1, respectively. The average production of phytoplankton corresponding to micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton was 0.11, 1.86 and 0.22 mgC l^–1 d^–1, respectively. The ratio of those to the total production of phytoplankton was 5%, 85% and 10%, respectively. The mean respiration rate of different size groups were 0.08, 0.72 and 0.46 mgC l^–1 d^–1, the ratio of those to total respiration of phytoplankton was 6%, 57% and 37%, respectively. The concentrations of chlorophyll-a of the phytoplankton in the corresponding size of micro- (10–112 m), nano- (3–10 m) and pico- (<3 m) plankton in the experimental ponds were 19.3, 98.2 and 11. 9 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 15%, 76% and 9%, respectively. The concentrations of chlorophyll-a of phytoplankton micro- (10–200 m), nano- (3–10 m) and pico- (<3 m) plankton in enclosures were 1.7, 34.3 and 3.0 g l^–1, respectively. The ratio of those to the total chlorophyll-a was 4%, 88% and 8%, respectively.     

Expression in Bacillus subtilis of the gene for human urogastrone using synthetic ribosome binding sites

Summary A chemically synthesised gene coding for human urogastrone which was earlier cloned in E. coli (Smith et al. 1982) has now been cloned into expression vectors for Bacillus subtilis Two types of constructs have been made, one giving production of methionylurogastrone and the other giving rise to a methionyl-urogastrone- galactosidase fusion polypeptide facilitating quantification of expression levels.The ribosome binding sites used in the expression plasmids are synthetically made oligonucleotides residing on short restriction fragments to allow easy replacement by other ribosome binding sites.Using shuttle vectors and constitutive promoters from Bacillus phages 105 and SPP1, we were able to detect levels of expression amounting to a few thousand molecules per cell during logarithmic growth in both E. coli and B. subtilis.     

Studies of α- and βL-Crystallin Complex Formation in Solution at 60°C

Studies of molecular mechanisms of chaperone-like activity of -crystallin became an active field of research over last years. However, fine interactions between -crystallin and the damaged protein and their complex organization remain largely uncovered. Complexation between - and _L-crystallins was studied during thermal denaturation of _L-crystallin at 60°C using small-angle X-ray scattering (SAXS), light scattering, gel-permeation chromatography, and electrophoresis. A mixed solution of - and _L-crystallins at concentrations about 10 mg/ml incubated at 60°C was found to contain their soluble complexes with a mean radius of gyration 14 nm, mean molecular mass 4 MDa and maximal size over 40 nm. In pure _L-crystallin solution, no complexes were observed at 60°C. In SAXS studies, transitions in the -crystallin quaternary structure at 60°C were shown to occur and result in doubling of the molecular weight. This suggests that during the temperature-induced denaturation of _L-crystallin it binds with modified -crystallin or, alternatively, _L-crystallin complexation and -crystallin modifications are concurrent. Estimates of the -_L-crystallin complex size and relative contents of - and -_L-crystallins in the complex suggest that several -crystallin molecules are involved in complex formation.     

Expression of bioactive human interferon-gamma in transgenic rice cell suspension cultures

We investigated the possibility of producing the therapeutic recombinant cytokine, Interferon-gamma (IFN-), in transgenic rice cell (Oryza sativa, cultivar TNG67) suspension cultures. We tested expression of two vector constructs, each harboring an Amy3 leader peptide and a C-terminus His 6 tag fused to a human IFN- cDNA, one driven by a sucrose-starvation inducible promoter (rice Amy3 promoter) and the other by a constitutive maize ubiquitin promoter, in rice cell suspensions, introduced via Agrobacterium tumefaciens. There was a significant difference in the amounts of recombinant IFN- protein produced by the Ups and Amy cell lines, as cytosolic and secretory proteins respectively. Immunological analysis of IFN- recombinant protein conferred a dose-dependent anti-dengue virus activity in human A549 cells, similar to the commercial product. We discuss the attractive attributes of using rice cell suspension system for the expression of therapeutic recombinant IFN-.     

Electrotonic spread of current in monolayer cultures of neonatal rat heart cells

Summary The passive electrical properties of neonatal rat heart cells grown in monolayer cultures were determined. Hyperpolarizing current pulses were injected through one microelectrode via an active bridge circuit. Membrane voltage displacements caused by the injected current pulses were measured at various distances from the first with a second microelectrode. Using a modified least-squares method the experimental results were fitted to a Bessel function, which is the steady-state solution of the differential equation describing the relation between membrane voltage caused by current injection and interelectrode distance in a very large and very thin plane cell. Best fit was obtained with a space constant of 360 m and an internal resistivity of 500 cm. From these figures, specific membrane resistance was calculated to be 1,300 cm², assuming all current to leave through the upper surface of the monolayer.The time constant of the membrane was measured from the time course of the current-induced membrane voltage displacements. From its value of 1.7 msec a membrane capacity of 1.3 F/cm² was calculated.From these results and some literature data on nexus distribution (A. W. Spira,J. Ultrastruct. Res. 34:409, 1971) specific nexus resistance was calculated to range between 0.25 and 1.25 cm², depending on the amount of folding of the intercalated discs. The results suggest that spread of activation in monolayer cultures of heart cells by means of local circuit currents is very likely.     

Problems in estimating growth rates of marine phytoplankton from short-term14C assays

Growth rate estimates () of phytoplankton populations that were sampled from nitrogen-limited continuous cultures and then incubated for short durations in batch culture with added¹⁴C-HCO₃ ^– were significantly different than steady-state growth rates () for 3 of 5 marine phytoplankton species. Two diatoms,Thalassiosira weissflogii andChaetoceros simplex, displayed virtually identical growth rates (=) over a wide range of, whereas for a third diatom,Phaeodactylum tricornutum, was overestimated by an average of 40% compared to. In contrast, was underestimated by the¹⁴C technique for the two remaining species: up to 40% at a steady-state of 1.0 day^–1 for the chlorophyteDunaliella tertiolecta and up to 100% at of 1.4 day^–1 for the haptophytePavlova lutheri. For the latter two species the divergence between and appeared to increase with increasing steady-state. A simple model of labeled and total carbon flow between the aqueous phase and cellular biomass was constructed to demonstrate that respiration was negligible when=, but was significant when>. In the cases in which<, a rapid physiological alteration presumably took place once the steady state was disturbed and cells were placed in the incubation chambers, which perhaps was related to the nutritional state of the cultures at the time of sampling. Questions thus are raised regarding our ability to measure accurately primary productivity from shipboard experiments with confined samples of phytoplankton from nutrient-impoverished waters that probably are less hardy than the laboratory cultures used in these studies.     

Stimulation of β-carotene synthesis by hydrogen peroxide in Blakeslea trispora

-Carotene synthesis was increased from a negligible amount to 152 mg (g-dry cells)^–1 and H₂O₂ was accumulated up to 16.7 M during 2.5 day-culture of Blakeslea trispora. When cells were cultivated in 250 ml flasks containing various volumes (25–150 ml) of the medium, not only H₂O₂ accumulation but also -carotene synthesis increased as culture volume decreased. Addition of H₂O₂ (10 M) to the 1.5-day old cultures of B. trispora resulted in 46% higher -carotene synthesis than that without addition. All these results indicate that -carotene biosynthesis is stimulated by H₂O₂ in B. trispora.     

Vertical distribution and diurnal migrations of krill — Euphausia superba Dana — from hydroacoustical observations,SIBEX, December 1983/January 1984

Summary The depth distributions of krill density with the resolution of 0.435 m has been obtained on the basis of SIBEX hydroacoustic data. Krill was observed to be mainly in a layer from 20 to 100 m with the maximum of biomass migrating from the near surface area at night to greater depths during day. The krill migration pattern can be described by the function: H(t)=A+Bcos (2t/T+), where H is depth of the mass center of biomass, t-time, A-mean depth of krill occurrence, B-amplitude of diurnal changes, T=24 h and -phase of migration process. The parameters of migration pattern: A, B, T and depend on the body length of krill.     

Cell specificity in auxin- and ethylene-induced ‘supergrowth’ in Riella helicophylla

Summary In gemmalings of Riella helicophylla, auxin and ethylene stimulate elongation growth, especially of pillar cells. When the two hormones are supplied simultaneously, the effects are additive, i.e. the result is supergrowth. In the cells of the meristem, elongation is enhanced by auxin, but not by ethylene when given alone. However, these cells also respond with supergrowth to a combined treatment with auxin and ethylene. The antiauxin p-chlorophenoxyisobutyric acid suppresses both the ethylene stimulation of cell growth and the additive supergrowth. The results support the concept that auxin pre-conditions the cells to the ethylene-dependent growth event. We suggest that the response elicited by the specific cell types could be related to differences in their level of endogenous auxin.Abbreviations IAA indole-3-acetic acid - PCIB p-chlorophenoxyisobutyric acid     

Erratum: Cloning and sequencing of the phycocyanin gene from Spirulina maxima and its evolutionary analysis

The genes were cloned for the two apoprotein subunits, and ,of phycocyanin from the cyanobacterium Spirulina maxima = Arthrospiramaxima) strain F3. The - and -subunit gene-coding regionscontain 489 bp and 519 bp, respectively. The -subunit gene is upstreamfrom the -subunit gene, with a 111-bp segment separating them.Similarities between the -subunits of S. maxima and nine othercyanobacteria were between 58% and 99%, as were those between the -subunits. The maximum similarity between the - and -subunits from S. maxima was 27%.     

Beiträge zur Züchtung moniliaresistenter Sauerkirschen

Zusammenfassung Es wurden 7 Sauerkirschsorten während der Zeit der Vollblüte im Frühjahr 1958 mit drei Herkünften vonSclerotinia laxa und einer Herkunft vonSclerolinia fructigena infiziert. Bei allen Sorten wurden Zweiginfektionen, Blüteninfektionen im Freiland und Blüteninfektionen an abgeschnittenen Zweigen im Gewächshaus durchgeführt.Als sicherste und wenig arbeitsaufwendige Infektionsmethode stellte sich die Zweiginfektion heraus. Nach Infektion der Narben ist das Fortschreiten des Pilzwachstums wahrscheinlich von der Witterung abhängig. Blüteninfektionen im Gewächshaus an abgeschnittenen Zweigen eigneten sich unter den angewendeten Infektionsbedingungen nicht, da die Resistenzunterschiede der Sorten zu undeutlich zum Ausdruck kamen. Beim Vergleich der Ergebnisse von Zweig-und Blüteninfektionen zeigte sich in den Extremen übereinstimmung.Die Schattenmorelle erwies sich als anfälligste Sorte. Köröser Weichsel, Ostheimer Weichsel und Gubens Ehre waren ebenfalls anfällig. Als etwas resistent stellte sich Podbielski heraus. Die Sorte Großer Gobet was weitgehend resistent.Die drei Herkünfte vonSclerotinia laxa zeigten auf den Sorten unterschiedliche Virulenz, so daß zukünftige Resistenzprüfungen mit verschiedenen Isolierungen durchgeführt werden müssen. Die vonSclerotinia fructigena verwendete Herkunft konnte in keinem Fall Infloreszenzen infizieren; der nach Triebinfektion erzeugte Befall war ebenfalls sehr gering.Mit 5 Abbildungen     

The inheritance of β-amylase null in storage roots of sweet potato,Ipomoea batatas (L.) Lam.

Summary Several sweet potato genotypes were found to lack completely or to have only traces of-amylase in their storage roots. Such genotypes do not increase in sweetness during cooking because, without a sufficient amount of-amylase, the normal hydrolysis of starch to maltose does not occur in the cooking process. In order to study the inheritance of this biochemical variant in the genotype, 41 families were generated. The following conclusions were drawn from analyzing these families. (1) This trait is controlled by one recessive allele (designated-amy) (2) It is inherited in a hexasomic or tetradisomic manner, but not disomically or tetrasomically. This conclusion supports previous cytological data that sweet potato is an autohexaploid or has two identical genomes plus one genome which is somewhat different. (3) The-amy allele appears to exist at a high frequency in cultivated germplasm. (4) Breeding sweet potato for low-amylase activity is relatively easy. New types of sweet potato without normal-amylase activity have great potential for processing and as a staple food.     

Moulting interval and growth of juvenile Antarctic krill (Euphausia superba) fed different concentrations of the diatom Phaeodactylum tricornutum in the laboratory

Summary Juvenile Antarctic krill (Euphausia superba) were fed the diatom Phaeodactylum tricornutum at concentrations ranging from 0.0–5.0 mgCl^-1. Intermoult period (IP) decreased, but an increment of body length per moult (BL) of juvenile krill increased, up to a concentration of 1.0 mgCl^-1. No further effect of food concentrations on IP or BL was seen at concentration beyond 1.0 mgCl^-1. IP plateaued at 23.8 days and BL, 1.14 mm. The maximum daily growth rate (BL/IP) of juvenile krill was calculated to be 0.047 mm day^-1. BL and IP were correlated and the relationship is expressed as BL=-0.0066IP+2.47 (r=0.58, n=141, P<0.01). Growth conditions of krill in the Southern Ocean are discussed in terms of available food concentration in there.     

Effects of photoperiod,temperature, food and relative humidity on the induction of diapause in the predatory miteAmblyseius potentillae

Aspects of the induction of diapause were studied in a Dutch strain of the phytoseiid miteAmblyseius potentillae. The photoperiodic response curve was of the long-day type, with a sharply defined critical daylength of 14.5 h. Critical daylength varied only little at temperatures between 15.0 and 22.5°C.All post-embryonic and possibly even late-embryonic stages of development were found to be sensitive to photoperiod; sensitivity appeared to be maximal during the protonymphal stage.It is shown that -carotene is necessary for some early step in the physiological mechanism of photoperiodic induction, and not (or not exclusively) for the expression of the diapause response.Two points of sensitivity to light could be demonstrated in the nights ofld 1311 andld 1212 long-night regimes, using 1-h night interruptions. These results are similar to those obtained in lightbreak experiments with spider mites and insects. However, no effect was found with light interruptions applied during the dark phase of anld 1014 long-night regime.In resonance experiments with a constant photophase (12 h) and a variable scotophase, a weak rhythmic response was found at 22.5°C; at 19.0°C this effect was completely absent.The relative humidity experienced by the mites during diapause induction as well as during diapause development influenced the rate of diapause completion under long days (ld 168). Diapause duration appeared to be shortest when the mites experienced low relative humidity (35±5%) during diapause induction and high relative humidity (75±5%) during diapause termination, and longest under the reverse conditions.     

Optimal foraging by a suspension-feeding copepod: responses to short-term and seasonal variation in food resources

Laboratory radioisotope experiments were used to investigate the effects of phytoplankton seasonal succession on the selectivity and clearance rates of a suspension-feeding copepod in two Indiana lakes. Responses to particle size and quality were tested by allowing adult female Diaptomus birgei feeding in natural seston to select between a small (6×7 m) flagellate (Chlamydomonas reinhardii) and a large, poor quality food (heat-killed Carteria olivieri, 22×25 m). Short-term responses were tested in one lake by additional treatments in which copepods were acclimated for 5–6 h in filtered lake water (starved) or natural seston with added Chlamydomonas (enriched). Copepods from both lakes fed selectively on the small live flagellate during the spring bloom of edible phytoplankton but fed selectively on the larger, poor quality particle during the clear water phase when food was scarce. These results are interpreted as an interaction between the concentration-dependent selectivity for high quality foods predicted by optimal diet theory and a perceptual bias for large-sized particles. Selectivity for high-quality food was intermediate and clearance rates were depressed when total phytoplankton abundance was high but dominated by filamentous cyanobacteria. In each experiment copepods also responded to the short-term manipulations by exhibiting weaker discrimination against the poor quality particle in the starvation treatment and stronger discrimination in the enriched treatment. A two-way mixed model ANOVA revealed substantial short term (37%) and seasonal (53%) components to the total variance in selectivity. Clearance rates were also influenced by both phytoplankton succession and the short-term resource manipulations. As expected, clearance rates on the poor quality food were more sensitive to the abundance of alternative foods. These results show how the feeding behavior of a freshwater copepod is modulated by both seasonal and short-term variation in natural food.