Purification and characterization of thermostable pectate-lyases from a newly isolated thermophilic bacterium, Thermoanaerobacter italicus sp. nov.

A novel thermophilic spore-forming anaerobic microorganism (strain Ab9) able to grow on citrus pectin and polygalacturonic acid (pectate) was isolated from a thermal spa in Italy. The newly isolated strain grows optimally at 70°C with a growth rate of 0.23 h⁻¹ with pectin and 0.12 h⁻¹ with pectate as substrates. Xylan, starch, and glycogen are also utilized as carbon sources and thermoactive xylanolytic (highest activity at 70°–75°C), amylolytic as well as pullulolytic enzymes (highest activity at 80°–85°C) are formed. Two thermoactive pectate lyases were isolated from the supernatant of a 300-l culture of isolate Ab9 after growth on citrus pectin. The two enzymes (lyases a and b) were purified to homogeneity by ammonium sulfate treatment, anion exchange chromatography, hydrophobic chromatography and finally by preparative gel electrophoresis. After sodium dodecylsulfate (SDS) gel electrophoresis, lyase a appeared as a single polypeptide with a molecular mass of 135 000 Da whereas lyase b consisted of two subunits with molecular masses of 93 000 Da and 158 000 Da. Both enzymes displayed similar catalytic properties with optimal activity at pH 9.0 and 80°C. The enzymes were very stable at 70°C and at 80°C with a half-life of more than 60 min. The maximal activity of the purified lyases was observed with orange pectate (100%) and pectate-sodium salt (90%), whereas pectin was attacked to a much lesser extent (50%). The K _m values of both lyases for pectate and citrus pectin were 0.5 g·l⁻¹ and 5.0 g·l⁻¹, respectively. After incubation with polygalacturonic acid, mono-, di-, and tri-galacturonate were detected as final products. A 2.5-fold increase of activity was obtained when pectate lyases were incubated in the presence of 1 mM Ca²⁺. The addition of 1 mM ethylenediaminetetraacetic acid (EDTA) resulted in complete inhibition of the enzymes. These heat-stable enzymes represent the first pectate-lyases isolated and characterized from a thermophilic anaerobic bacterium. On the basis of the results of the 16S rRNA sequence comparisons and the observed phenotypic differences, we propose strain Ab9 as a new species of Thermoanaerobacter, namely Thermoanaerobacter italicus sp. nov. Received: May 25, 1997 / Accepted: June 5, 1997     

Growth of Nitrobacter by dissimilatoric nitrate reduction

Abstract Eight strains of the genus Nitrobacter grew under anaerobic conditions in the presence of nitrate. The growth was inhibited by nitrate concentrations above 0.5 mM. By a special culture technique inhibition caused by nitrite was abolished. Nitrate oxidizing cells grew in gas tight culture flasks as a biofilm on a gas-permeable silicone tubing. The biofilm allowed nitrate-reducing cells to grow at a low nitrite concentration. These cells grew either actively motile in the anaerobic medium, or in anaerobic zones of the biofilm. They produced nitrite and ammonia. Nitrogen balance calculations established a loss of inorganic nitrogen for 5 of 8 strains. This implies that nitrate-reducing cells produced furthermore volatile nitrogen compounds. N₂O was detected by gas chromatography.     

Effects of Alternaria alternata f.sp. lycopersici toxins on pollen

Summary Effects of the phytotoxic compounds (AAL-toxins) isolated from cell-free culture filtrates of Alternaria alternata f.sp. lycopersici on in vitro pollen development were studied. AAL-toxins inhibited both germination and tube growth of pollen from several Lycopersicon genotypes. Pollen from susceptible genotypes, however, was more sensitive for AAL-toxins than pollen from resistant plants, while pollen of species not belonging to the host range of the fungus was not significantly affected by the tested toxin concentrations. AAL-toxins elicit symptoms in detached leaf bioassays indistinguishable from those observed on leaves of fungal infected tomato plants, and toxins play a major role in the pathogenesis. Apparently, pathogenesis-related processes and mechanisms involved in disease resistance are expressed in both vegetative and generative tissues. This overlap in gene expression between the sporophytic and gametophytic level of a plant may be advantageously utilized in plant breeding programmes. Pollen may be used to distinguish susceptible and resistant plants and to select for resistances and tolerances against phytotoxins and other selective agents.     

云南省(虫齿)目二新种和灰背蛇(虫齿)的记述((虫齿)目:厚(虫齿)科、叉(虫齿)科、围(虫齿)科)

本文记述了云南省(虫齿)目二新种,Tapinella bannana sp.n.和Peripsocus plurimaculatus sp.n.及一新种记录种Ophiodopelma semicets Lee and Thornton,其雄虫为首次记载。     

Initial reactions in the mineralization of 2-sulfobenzoate by Pseudomonas sp. RW611

Abstract Pseudomonas sp. strain RW611 utilized the ammonium salt of 2-sulfobenzoate as sole source of carbon, sulfur, nitrogen, and energy. The xenobiotic sulfo substituent was dioxygenolytically eliminated as sulfite, which was then slowly oxidized to sulfate. 2,3-Dihydroxybenzoate, which resulted from desulfonation underwent meta -cleavage, mediated by 2,3-dihydroxybenzoate 3,4-dioxygenase activity. This enzyme was inhibited by 3-chlorocatechol and 2,3,4-trihydroxybenzoate.     

Control of nitrogenase inAzospirillum sp.

Many N₂-fixing organisms can turn off nitrogenase activity in the presence of NH₄ ⁺ and turn it on again when the NH₄ ⁺ is exhausted. One of the most interesting systems for accomplishing this is by covalent modification of one subunit of dinitrogenase reductase by dinitrogenase reductase ADP-ribosyltransferase (DRAT). The system can be reactivated when NH₄ ⁺ is exhausted, by dinitrogenase reductase activating glycohydrolase (DRAG) which removes the inactivating group. It is fascinating that some species of the genusAzospirillum possess the DRAT and DRAG systems (A. lipoferum andA. brasilense), whereasA. amazonense in the same genus lacks DRAT and DRAG.A. amazonense responds to NH₄ ⁺ but does not exhibit modification of dinitrogenase reductase characteristic of the action of DRAT. However, it has been possible to clone DRAT and DRAG and to introduce them intoA. amazonense, whereupon they become functional in this organism. The DRAT and DRAG system does not appear to function inAcetobacter diazotrophicus, an organism isolated from sugar cane, that fixes N₂ at a pH as low as 3.0.A. diazotrophicus does show a rather sluggish response to NH₄ ⁺. A level of about 10 M NH₄ ⁺ is required to switch off the system. The response to NH₄ ⁺ is influenced by the dissolved oxygen concentration (DOC) as has been reported forAzospirillum sp. A DOC in equilibrium with 0.1 to 0.2 kPa O₂ seems optimal for the response inA. diazotrophicus.     

Structural features of the plastid ribosomal RNA operons of two red algae: Antithamnion sp. and Cyanidium caldarium

The nucleotide sequences of the plastid 16S rDNA of the multicellular red alga Antithamnion sp. and the 16S rDNA/23S rDNA intergenic spacers of the plastid DNAs of the unicellular red alga Cyanidium caldarium and of Antithamnion sp. were determined. Sequence comparisons support the idea of a polyphyletic origin of the red algal and the higher-plant chloroplasts. Both spacer regions include the unsplit tRNA^Ile (GAU) and tRNA^Ala (UGC) genes and so the plastids of both algae form a homogeneous group with those of chromophytic algae and Cyanophora paradoxa characterized by small-sized rDNA spacers in contrast to green algae and higher plants. Nevertheless, remarkable sequence differences within the rRNA and the tRNA genes give the plastids of Cyanidium caldarium a rather isolated position.     

Interacting effects of increased tannin levels on leaf-mining insects

Tannin-rich extracts were painted on leaves of Q. emoryi (Fagaceae) that contained first instar larvae of a leaf miner, Cameraria sp. (Lepidoptera: Gracillariidae, Agrifoliella group), to test the role of tannins in increasing parasitism, while controlling for changes in protein content and physical damage. Miners from leaves that were treated with tannins had higher rates of parasitism by hymenopterans, but decreased mortality from other causes. The negative and positive effects of tannins balanced since overall larval survivorship was equivalent for leaf miners on tannin-enriched and control leaves. The tannin-rich extract also affected larval development because pupal weight was decreased for leaf miners in treated leaves. Tannins can have both adverse and beneficial effects on these phytophagous insects.

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Résumé Des extraits riches en tanins ont été étalés sur des feuilles de Quercus emoryi contenant des chenilles de ler stade de Cameraria sp., pour contrôler l'influence du tanin dans l'accroissement du parasitisme sans modifier la teneur en protéines ni provoquer de dommages physiques. Les mineuses de feuilles traitées avaient des taux plus élevés de parasitisme par les hyménoptères, mais une diminution des autres causes de mortalité. Les effets négatifs et positifs des tanins se sont équilibrés, puisque la survie des mineuses était la même pour les feuilles traitées que pour les témoins. Les extraits riches en tanin ont agi sur le développement larvaire puisque le poids des chrysalides des mineuses de feuilles traitées était plus faible. Les tanins peuvent ainsi avoir des effets nuisibles et bénéfiques sur ces insectes phytophages.

     

Biomanipulation and its feasibility for water quality management in shallow eutrophic water bodies in The Netherlands

Biomanipulation as a tool for lake restoration is discussed mainly using literature data. It is based on the exploitation of the interactions both within and between the trophic levels in an aquatic ecosystem. Important among the interactions are: competition for light and nutrients between aquatic macrophytes and phytoplankton and among different phytoplankton species; grazing by planktonic and benthic filter feeders; and size-selective predation by fish. In several case studies biomanipulation has proved to be successful in restorating mildly eutrophic small waterbodies. However, for long-term stability of the restored ecosystems supplementary measures like reducing the external nutrient loadings are needed. The feasibility of the different biomanipulation measures to improve the water quality in shallow Dutch lakes is discussed. Preliminary results on biomanipulation experiments in enclosures withOscillatoria agardhii and the benthic filter feederDreissena polymorpha are given.     

Comparative study of methods for the quantification of fungal spores at the leaf surfaces ofPopulus xeuramericana

Summary The removal of fungal spores, urediniospores ofMelampsora medusae and conidia ofPestalozzia sp., from the leaf surfaces ofPopulus xeuramericana (Dode) Guinier cv. I-488 was assessed using three cultural techniques conventionally employed in phylloplane studies. The method of removal and the original density of spore deposition, but not the interaction of these factors, were significant determinants of variability in spore removal. Irrespective of the original density of deposition, the leaf print method was the most, and the leaf washing technique the least, efficient means of spore removal from the leaf surface. Factors which could contribute to this difference in efficiency are discussed.