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    Tytuł:
    Toksyczne metabolity wytwarzane przez pasozytniczy grzyb Conidiobolus coronatus
    Toxic metabolites produced by parasitic fungus Conidiobolus coronatus
    Autorzy:
    Wieloch, W
    Powiązania:
    https://bibliotekanauki.pl/articles/837083.pdf
    Data publikacji:
    2007
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    metabolity toksyczne
    grzyby pasozytnicze
    parazytologia
    mikotoksyny
    Entomophthorales
    Conidiobolus coronatus
    toksycznosc
    Galleria mellonella
    Źródło:
    Annals of Parasitology; 2007, 53, 4; 355-357
    0043-5163
    Pojawia się w:
    Annals of Parasitology
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
    Tytuł:
    Toksyczne metabolity wytwarzane przez pasożytniczy grzyb Conidiobolus coronatus
    Toxic metabolites produced by parasitic fungus Conidiobolus coronatus
    Autorzy:
    Wieloch, W.
    Powiązania:
    https://bibliotekanauki.pl/articles/2144082.pdf
    Data publikacji:
    2007
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    metabolity toksyczne
    grzyby pasozytnicze
    parazytologia
    mikotoksyny
    Entomophthorales
    Conidiobolus coronatus
    toksycznosc
    Galleria mellonella
    Opis:
    Naturally occurring entomopathogens are important regulatory factors of insect populations. Among them are entomopathogenic fungi. The invasion of insects by parasitic fungi occurs through penetration of the host integument. Death of the host is a result of tissue destruction, exhaustion of nutrients or the production of toxins. Conidiobolus coronatus (Entomophthorales) is a saprophytic soil fungus that kills insects by releasing toxins inside insect body, before the invasion of host's organs and tissues by fungal hyphae. It is pathogenic to a number of insects and could be relatively easily propagated in laboratory conditions. The fungus is an interesting object to study and might be the source of new insecticidal substances as well. The main aim of the study was isolation and characterisation of active compounds produced by C. coronatus. In experimental surveys of interactions between insects and entomopathogenic fungi it is important to establish simple and reliable method of quantification of fungal pathogenicity towards insects, and to chose right insect target as well. Four methods were tested on two species — Galleria mellonella and Dendrolimus pini: (1) immersing larvae in conidial suspension; (2) deposition the conidia on the cuticle; (3) injection into hemocoel, and (4) exposure to fungal colony. Exposition of G. mellonella larvae to fungal colony was chosen, as the best method to quantify C. coronatus pathogenicity, reflecting possible contact of insects with fungal spores in nature. D. pini larvae were not chosen to further experiments. Dark colour of their body disables the estimation of fungal infection progress. The fungus produces an array of enzymes regarded as necessary in efficient penetration of insect cuticle: proteases, chitinases and lipases, which degrade the components of the integument. The activity of those enzymes was measured in mycelial homogenates and in post incubation media. In homogenates the activity of elastase, N−acetylglucosaminidase (NAGase) and lipase was denoted. The homogenate had no chymotripsin and chitinase activity. In the incubation media the activity of five examined enzymes was present. Elastase and NAGase activities were much higher than those of three other enzymes. The long term observation of four colonies in laboratory conditions from one transfer to another revealed differences in the ability to kill G. mellonella larvae. The colonies reduced pathogenicity during several transfers and then relapsed into higher level of pathogenicity again. The fluctuations were more or less regular and appeared through two−year duration of the experiment. The nature of this fluctuation is unknown and no similar phenomenon was observed elsewhere. To elicit the possible background of instability of fungal cultures towards G. mellonella, a genetic analysis was performed on colonies derived from primary conidia, containing several dozen of nuclei, and microconidia, which are formed on the surface of primary conidia and containing a maximum several nuclei. Both analyses: DNA Fingerprinting and Amplified Fragments Length Polymorphism (AFLP) revealed that colonies isolated from primary conidia or microconidia differ in the genetic profile. The genetic differences reflect the differences in the pathogenicity trait. Genetic analysis of colonies derived from microconidia proved that nuclei differ genetically, which means that C. coronatus mycelium is heterokaryotic. A chromatographic separation of fungal homogenate proteins did not succeed. Better possibility gave the separation of proteins released by fungus to minimal medium. By two step high pressure liquid chromatography: size exclusion and ion exchange four proteins were separated to homogeneity, according to SDS−PAGE. Two of them in the size 14.5 kDa and 36 kDa were moderately pathogenic to G. mellonella larvae in the dose of 1µg per larva (20% and 10% of pathogenicity, respectively), and exhibited no enzymatic activity. Third protein was a 33−34 kDa elastase with no pathogenic effect. The last protein in the size 36−37 kDa was pathogenic to the larvae (20%) and exhibited elastolytic and chitinolytic activities. Further experiments will elicit the mode of actions on cell cultures of those four isolated proteins.
    Źródło:
    Wiadomości Parazytologiczne; 2007, 53, 4; 355-357
    0043-5163
    Pojawia się w:
    Wiadomości Parazytologiczne
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
    Tytuł:
    Mechanisms of Galleria mellonella cellular immune response after infection with entomopathogenic fungus Conidiobolus coronatus
    Autorzy:
    Ligeza-Zuber, M.
    Powiązania:
    https://bibliotekanauki.pl/articles/5856.pdf
    Data publikacji:
    2012
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    mechanism
    Galleria mellonella
    cellular immune response
    fungal infection
    entomopathogenic fungi
    Conidiobolus coronatus
    host
    toxicity
    Źródło:
    Annals of Parasitology; 2012, 58, 4
    0043-5163
    Pojawia się w:
    Annals of Parasitology
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
    Tytuł:
    Activity of cuticle-degrading enzymes from entomopathogenic fungus Conidiobolus coronatus
    Autorzy:
    Czygier, M.
    Samborski, J.
    Dzik, J.M.
    Walajtys-Rode, E.
    Bogus, M.
    Powiązania:
    https://bibliotekanauki.pl/articles/839987.pdf
    Data publikacji:
    1998
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    Galleria mellonella
    lipase
    chymotrypsin
    cuticle
    enzyme
    cuticle-degrading enzyme
    larva
    Conidiobolus coronatus
    entomopathogenic fungi
    chitinase
    host
    N-acetylglucosaminidase
    Źródło:
    Annals of Parasitology; 1998, 44, 3
    0043-5163
    Pojawia się w:
    Annals of Parasitology
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
    Tytuł:
    Charakterystyka enzymow proteo-, chityno- i lipolitycznych pasozytniczego grzyba Conidiobolus coronatus
    Characterization of proteo-, chitino- and lipolytic enzymes of parasitic fungus Conidiobolus coronatus
    Autorzy:
    Wloka, E
    Powiązania:
    https://bibliotekanauki.pl/articles/836974.pdf
    Data publikacji:
    2010
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    grzyby pasozytnicze
    grzyby entomopatogenne
    Conidiobolus coronatus
    Entomophthorales
    enzymy proteolityczne
    elastaza
    N-acetyloglukozaminidaza
    enzymy chitynolityczne
    enzymy lipolityczne
    lipaza
    aktywnosc enzymatyczna
    owady
    barciak wiekszy
    Galleria mellonella
    zwalczanie szkodnikow
    metody biologiczne
    grzyby owadobojcze
    Źródło:
    Annals of Parasitology; 2010, 56, 1; 83-85
    0043-5163
    Pojawia się w:
    Annals of Parasitology
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
    Tytuł:
    Charakterystyka enzymów proteo-, chityno- i lipolitycznych pasożytniczego grzyba Conidiobolus coronatus
    Characterization of proteo-, chitino- and lipolytic enzymes of parasitic fungus Conidiobolus coronatus
    Autorzy:
    Włóka, E
    Powiązania:
    https://bibliotekanauki.pl/articles/2143413.pdf
    Data publikacji:
    2010
    Wydawca:
    Polskie Towarzystwo Parazytologiczne
    Tematy:
    grzyby pasozytnicze
    grzyby entomopatogenne
    Conidiobolus coronatus
    Entomophthorales
    enzymy proteolityczne
    elastaza
    N-acetyloglukozaminidaza
    enzymy chitynolityczne
    enzymy lipolityczne
    lipaza
    aktywnosc enzymatyczna
    owady
    barciak wiekszy
    Galleria mellonella
    zwalczanie szkodnikow
    metody biologiczne
    grzyby owadobojcze
    Opis:
    The largest problem in limitation of insect pest population is increasing resistance of them to chemical pesticides. Alternative are entomopathogens, which regulate frequency of insect pests. Among them decisive role play entomopathogenic fungi, which possess the ability to active penetration through cuticle by mechanical pressure of invasive hypha and production of proteo-, chitino- (egzo- and endochitinases) as well as lipolytic enzymes, which provide nutrients for subsequent development of fungus. Entomopathogenic soil fungus Conidiobolus coronatus (Entomophtorales) is saprophyte fungus, which demonstrates a high efficiency in the paralysis of varied insects. Although leading investigations over mechanism of insect paralysis, we still do not know, what role fungal enzymes play in insect cuticle penetration. The main aim of research was establishment of optimal conditions for elastase, N-acetylglucosaminidase (NAGase), chitobiosidase as well as lipase. Optimal reaction parameters were determined: volume of reaction mixture, volume of homogenate, working pH and the substrate concentration. Having on aim a possible use of C. coronatus in pest control, two ranges of temperatures were chosen: 20°C – optimal temperature for the fungus growing and 30°C – optimal temperature for the cultivation of the great wax moth larvae, Galleria mellonella, on which examinations were performed. Also kinetic constants Km and Vmax were determined. Activity of elastase and N-acetylglucosaminidase of C. coronatus was measured spectrophotometrically at 410 nm (towards N-Succinyl-Ala-Ala-Pro-Leu-p-Nitroanilide) and 405 nm (towards 4-Nitrophenyl-N-acetyl-b-D-glucosaminide), respectively. The following optimal conditions of elastase activity were established: the volume of reaction mixture 0.5 ml, volume of homogenate 1 ml, temperature 30°C, pH 8, substrate concentration 40 mM. Optimal conditions of NAGase assay: the volume of reaction mixture 0.5 ml, dose of homogenate 12.5 ml, temperature 30°C, pH neutral and 6 mM substrate concentration. The activities of chitobiosidase and lipase were measured spectrofluorometrically (Ex=360 nm, Em=450 nm) towards 4-Methylumbelliferyl b-D-N-N’-diacetylchitobioside and 4-Methylumbelliferyl oleate, respectively. Chitobiosidase showed the highest activity in dose of 30 ml in 1 ml volume of reaction mixture, at the temperature of 30°C, pH 7 and substrate concentration equal to 2 mM. Lipase showed the highest catalytic activity in 1 ml volume of reaction mixture, in 30°C but 50 ml of homogenate, pH 10 and 10 mM substrate concentration were needed. Higher activity investigated enzymes in 30°C than 20°C indicated that they can take part in pathogenesis. It was suggested that as first in perforation of coats of insects body elastase and lipase take part. Indicated of it, large thermoresistance of both enzymes (only 10.5% decrease of elastase activity at 20°C and 9.4% decrease of lipase activity in comparison with maximal activity at 30°C), alkalophilicity of both proteins (elastase shows the alkaline optimal pH equal to 8 at pH 9 preserves 97% activity, and at pH 10 94% activity, respectively while lipase prefers the pH 10 and at pH 8 and pH 9 enzyme keeps 57 and 60% activity, respectively) as well as lack of repression by suitable substrates. Sigmoid character of curve concerning pH influence on the activity of both enzymes, also indicates similarity between elastase and lipase. On minor part of NAGase and chitobiosidase of fungus C. coronatus in perforation of coats of host body showed high sensibility of both enzymes on hydrogen ions concentration: both enzymes prefer neutral pH, in pH 6 and 8 lose over 35% activity but subjection to substrate repression and 3–4-fold growth of activity followed only in 30°C. In the course of work it was found, that rich medium (LB) stimulates growth of mycelium and production of fungal lipases. So far nobody managed to isolate chitinolytic or lipolytic enzymes from C. coronatus homogenate. The majority of fungal enzymes were isolated from post incubation filtrates. In the literature of the subject lack of data about C. coronatus NAGase, therefore in examinations also the trial of isolation NAGase from C. coronatus homogenate was undertaken. Activity of NAGase showed only first fraction, which did not separate with none of used columns. Disappointing results of purification on cation exchanger CM, weak anion exchanger DEAE, and strong anion exchanger Q were obtained as well as after fractionation tests with the use of Microcon microcolumns. In aim of NAGase molecular mass estimation, two zymograms were made with Triton X-100 and casein and with the use of fluorescent substrate 4-Methylumbelliferyl N-acetyl-b-D-glucosaminide. Molecular mass of NAGase from C. coronatus was established on ca. 60 kDa. This is the first report describing molecular weight of NAGase from C. coronatus. Examined NAGase has different properties than known NAGases from other entomopathogenic fungi. Although its molecular weight is equal to the Metarhizium anisopliae NAGase, optimal pH for both NAGases are different: neutral in the case of C. coronatus NAGase versus acidic in the case of M. anisopliae NAGase. Knowledge of molecular mass of the C. coronatus NAGase should allow to find a new method of this enzyme isolation from C. coronatus homogenate. Thanks to developed methods of assaying activities of elastase, NAGase, chitobiosidase and lipase, real becomes the understanding of mechanism of insects paralysis through C. coronatus fungus.
    Źródło:
    Wiadomości Parazytologiczne; 2010, 56, 1; 83-85
    0043-5163
    Pojawia się w:
    Wiadomości Parazytologiczne
    Dostawca treści:
    Biblioteka Nauki
    Artykuł
      Wyświetlanie 1-6 z 6

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