Fungi in the legislation of the Republic of Serbia

Conservation and protection of fungi have lately been considered as extremely important elements of the environmental conservation, and numerous environmental, scientific, medical, economic, cultural, ethical, and other reasons for such attitude exist today. This paper presents an overview of official regulations on the protection of fungi in the Republic of Serbia from the Act of Protection of 1991 until today. The paper lists and analyses the good and bad provisions of individual legal regulations. It registers the effects of the adopted regulations on the actual efficiency of protection of endangered species of fungi (macrofungi, mushrooms), and considers the impact of chronological development of legislation on the population of fungi in nature, and presents general measures to improve protection of mushrooms in the future. These measures primarily include reliable information and study of fungi as a basis for their effective protection based on scientific knowledge. [Projekat Ministarstva nauke Republike Srbije, br. OI-179079

Effect of cultivation conditions on ligninolytic enzyme production by Ganoderma carnosum

Ganoderma carnosum has been found in Europe only, at coniferous trees and it is difficult to distinguish it morphologically from G. lucidum. Since G. carnosum has not been studied biochemically yet, the aim of this study was to analyse its ability to produce Mn-dependent peroxidase (MnP), versatile peroxidase (VP) and laccase (Lac) under conditions of solid-state fermentation of selected plant raw materials as carbon sources, in the presence of different nitrogen concentrations in the medium. Wheat straw, corn stem, oak and grapevine sawdust were the analysed plant raw materials. Nitrogen source in synthetic medium was NH4NO3 and its concentrations were: 10mM N and 20 mM N. Enzyme activity was determined spectrophotometrically, using ABTS and phenol red, as the substrates for Lac and Mn-oxidizing peroxidases, respectively. Maximum level of MnP activity (56.82 U/l) was obtained in the medium with wheat straw and nitrogen concentration of 10 mM. Best carbon source for VP production was grapevine sawdust at nitrogen concentration of 10 mM (80.80 U/l). The obtained Lac activity was very low in the medium with wheat straw (1.80 U/l), while it was not detected in the presence of other three analyzed plant raw materials. Maximum of total protein content (0.06 mgml-1) was noted in the medium where oak sawdust was carbon source and nitrogen concentration was 20 mM.Vrste roda Ganoderma su producenti mnogih biološki aktivnih supstanci pa su objekti proučavanja mnogih medicinskih i farmaceutskih studija. U novije vreme se sve više pažnje poklanja upoznavanju njihovog ligninolitičkog enzimskog sistema, u cilju uspešne primene u različitim biotehnološkim procesima. Za razliku od G. lucidum, čiji je ligninolitički sistem intenzivno proučavan, ostale vrste ovog roda su retko bile objekti istraživanja. G. carnosum se teško morfološki razlikuje od G. lucidum. i još uvek nije biohemijski proučena, pa je cilj našeg istraživanja bio analiza produkcije Mn-oksidujućih peroksidaza i lakaza u uslovima čvrste kultivacije, na različitim biljnim ostacima (pšenična slama, stabljike kukuruza, piljevina hrasta i vinove loze) u prisustvu azota u obliku NH4NO3 i u koncentracijama od 10 mM, odnosno 20 mM. Maksimum produkcije Mn-zavisne peroksidaze dobijen je na pšeničnoj slami pri koncentraciji azota od 10 mM (56.82 U/l). Piljevina vinove loze i koncentracija azota od 10 mM bili su optimalni za sintezu verzatil peroksidaze (80.80 U/l). Dobijena aktivnost lakaza je bila izuzetno niska u medijumu sa pšeničnom slamom (1.80 U/l), dok na ostalim biljnim ostacima nije zabeležena. Maksimalni sadržaj ukupnih proteina je bio najveći u medijumu sa piljevinom hrasta kao izvorom ugljenika i koncentracijom azota od 10 mM.Projekat ministarstva br. 14304

Mold attack on frescoes and stone walls of Gradac monastery

Microfungi can colonize stone surfaces and form sub-aerial biofilms which can lead to biodeterioration of historic monuments. In this investigation samples for mycological analyses were collected from stone material with visible alteration on stone walls of Gradac monastery exterior. The prevailing fungi found on stone walls were dematiaceous hyphomycetes with melanized hyphae and reproductive structures (Alternaria, Aureobasidium, Cladosporium and Epicoccum species). The frescoes inside the monastery building were also analyzed for the presence of mycobiota. The predominant fungi found on frescoes were osmophilic species from genera Aspergillus and Penicillium. The significant result is identification of human pathogen species Aspergillus fumigatus on frescoes

Comparison of anti-Aspergillus activity of Origanum vulgare L. essential oil and commercial biocide based on silver ions and hydrogen peroxide

The antifungal activities of Origanum vulgare essential oil (EO) and of a biocide based on silver and hydrogen peroxide (Sanosil S003) against seven Aspergillus species isolated from different substrata (stone, brick, silk and paper) of cultural heritage objects in Serbia were evaluated. Microdilution, agar dilution and microatmosphere methods were used to determine minimal fungistatic and minimal fungicidal concentrations (MIC and MFC), and light microscopy to determine structural abnormalities. MIC and MFC values for O. vulgare EO ranged from 0.2 to 5 mg mL−1 and for Sanosil S003 from 5 to 250 mg mL−1. Aspergillus sp. sect. fumigati was the most susceptible isolate, where MIC and MFC values were achieved at 0.5 mg mL−1 for O. vulgare EO, while MIC and MFC values for Sanosil S003 were achieved at 5 and 10 mg mL−1, respectively. Morpho-physiological changes were documented in all isolates, including lack of sporulation, depigmentation of conidiogenous apparatus and conidia, and presence of aberrant fungal structures. O. vulgare EO exhibited stronger anti-Aspergillus activity than Sanosil S003, as demonstrated by the higher MIC and MFC values and fewer morpho-physiological changes observed in the tested Sanosil S003 concentrations. O. vulgare EO could be an excellent alternative to commercial biocides, with high potential in the field of cultural heritage conservation

The effect of Nepeta rtanjensis essential oil on test micromycetes mycelia growth

The antifungal activity of Nepeta rtanjensis Diklić et Milojević essential oil on mycelia growth has been performed by macrodilution method. The most efficient impact of N. rtanjensis essential oil on mycelia growth in vitro was found in Alternaria species with the same value of minimal inhibitory quantity (MIQ) of 0.6 μl/ml. Bipolaris spicifera and Cladosporium cladosporoioides had MIQ values of 1.0 μl/ml whereas Trichoderma viride with MIQ value of 1.6 μl/ml showed the most efficient defense against the essential oil examined. The values of minimal fungicidal quantity (MFQ) in Alternaria sp. 2, B. spicifera and C. cladosporioides match the MIQ values whereas MFQ values in Alternaria sp. 1 is 0.8 μl/ml, and in T. viride 1.8 μl/ml.Makrodilucionom metodom testiran je uticaj etarskog ulja endemične biljke Nepeta rtanjensis (Lamiaceae) na micelijalni rast mikromiceta: Alternaria sp. 1, Alternaria sp. 2, Bipolaris spicifera, Cladosporium cladosporioides i Trichoderma viride. Hemijska analiza etarskog ulja N. rtanjensis pokazala je apsolutnu dominaciju 4aα, 7α, 7aβ nepetalaktona (79.89%). Najefikasniji uticaj na rast micelije in vitro zabeležen je kod vrsta roda Alternaria sa istom vrednošću minimalne inhibitorne količine (MIK) od 0.6 μl/ml. Bipolaris spicifera i Cladosporium cladosporioides su imali MIK 1.0 μl/ml, dok je Trichoderma viride, sa vrednošću MIK od 1.6 μl/ml, pokazala najveću otpornost na dejstvo ispitivanog ulja. Vrednosti minimalne fungicidne količine (MFK) se kod Alternaria sp. 2 (0.6 μl/ml), B. spicifera (1.0 μl/ml) i C. cladosporioides (1.0 μl/ml) poklapaju sa vrednostima MIK, dok je kod Alternaria sp. 1 MFK 0.8 μl/ml, a kod T. viride 1.8 μl/ml. Etarsko ulje N. rtanjensis pokazalo je jako antifungalno dejstvo na rast micelije testiranih mikromiceta.nul

WHEAT STRAW CONVERSION BY ENZYMATIC SYSTEM OF GANODERMA LUCIDUM

The purpose of this study was to resolve the question of whether various nitrogen sources and concentrations affect characteristics of selected G. lucidum ligninolytic enzymes participating in wheat straw fermentation. This is the first study reporting the presence of versatile peroxidase activity in crude extract of G. lucidum culture, as well as isoforms profile of Mn-oxidizing peroxidases. NH4NO3 was the optimum nitrogen source for laccase and Mn-dependent peroxidase activity, while peptone was the optimum one for versatile peroxidase activity. Four bands with laccase activity were obtained by native PAGE and IEF separations from medium enriched with inorganic nitrogen source, and only two bands from medium containing organic source. Medium composition was not shown to affect isoenzyme patterns of Mn-oxidizing peroxidases. Four isoforms of Mn-dependent peroxidase and three of versatile peroxidase were obtained on native PAGE. By IEF separation, five isoforms of Mn-dependent peroxidase and only two of versatile peroxidase were observed. The results demonstrated that G. lucidum has potential for mineralization and transformation of various agricultural residues and should take more significant participation in large-scale biotechnological processes

Antifungal activity of essential oil Hyssopus officinalis L. against micopathogen Mycogone perniciosa (Mang)

The most commonly cultivated mushroom species is the Agaricus bisporus Lange (Imb). One of the major pathogenic diseases of the cultivated mushroom in Serbia is Mycogone perniciosa (Mang). Biological control systems are not much used in mushroom cultivation. Medical and aromatic plants have been placed in the focus of intense studies. Pure culture of the M. perniciosa was isolated from infected A. bisporus. The essential oil of Hyssopus officinalis L. is used as a potential antifungal agent. The most abundant components in oil are isopinocamphone (43.29%), pinocamphone (16.79%) and b-pinene (16.31%). Antifungal activity of Hyssop was investigated by the modified microatmosphere method. The minimal inhibitory quantity was 5 μL/mL and a minimal fungicidal quantity was 15-20 μL/mL. There is no report on the use of Hyssop essential oil in mushroom disease.Agaricus bisporus Lange (Imb) je najčešće komercijalno gajena jestiva gljiva Različiti mikroorganizmi gljive, bakterije i virusi su izazivači bolesti u gajilištima šampinjona. Mycogone perniciosa (Mang) je izazivač bolesti poznate pod nazivom vlažni mehur i najčešći uzročnik gubitaka u gajilištima u Srbiji. Biološka kontrola, koja je uspešno primenjivana na nekim poljoprivrednim kulturama, nije korišćena prilikom uzgoja gljiva. Jedna od mogućnosti je primena biljnih sprejova. Lekovite i aromatične vrste biljaka se intenzivno istražuju kao mogući antifungalni agensi. Uzorci obolelih šampinjona su sakupljani u gajilištima u Srbiji. Kulture M. perniciosa su izolovane sa obolelih plodonosnih tela A. bisporus. Korišćeno je etarsko ulje Hyssopus officinalis. Najzastupljenije komponente ulja su izopinokamfon (43.29%) trans-pinokamfon (16.79%) i b-pinen (16.31%). Antifungalna aktivnost etarskog ulja izopa ispitivana je modifikovanom "mikroatmosfera"- metodom. Minimalna inhibitorna količina je bila 5 μL/mL, a minimalna fungicidna količina 15-20 μL/mL. Velik broj preparata je napravljen i primenjen za kontrolisanje oboljenja pečuraka: fungicidi, primena mikrotalasa ili dejstvo nekih antagonističkih bakterija. Dosad nije bilo saopštenja o primeni etarskog ulja izopa protiv izazivača bolesti gajenih gljiva.nul

Effect of cultivation conditions on ligninolytic enzyme production by Ganoderma carnosum

Ganoderma carnosum has been found in Europe only, at coniferous trees and it is difficult to distinguish it morphologically from G. lucidum. Since G. carnosum has not been studied biochemically yet, the aim of this study was to analyse its ability to produce Mn-dependent peroxidase (MnP), versatile peroxidase (VP) and laccase (Lac) under conditions of solid-state fermentation of selected plant raw materials as carbon sources, in the presence of different nitrogen concentrations in the medium. Wheat straw, corn stem, oak and grapevine sawdust were the analysed plant raw materials. Nitrogen source in synthetic medium was NH4NO3 and its concentrations were: 10mM N and 20 mM N. Enzyme activity was determined spectrophotometrically, using ABTS and phenol red, as the substrates for Lac and Mn-oxidizing peroxidases, respectively. Maximum level of MnP activity (56.82 U/l) was obtained in the medium with wheat straw and nitrogen concentration of 10 mM. Best carbon source for VP production was grapevine sawdust at nitrogen concentration of 10 mM (80.80 U/l). The obtained Lac activity was very low in the medium with wheat straw (1.80 U/l), while it was not detected in the presence of other three analyzed plant raw materials. Maximum of total protein content (0.06 mgml-1) was noted in the medium where oak sawdust was carbon source and nitrogen concentration was 20 mM.Vrste roda Ganoderma su producenti mnogih biološki aktivnih supstanci pa su objekti proučavanja mnogih medicinskih i farmaceutskih studija. U novije vreme se sve više pažnje poklanja upoznavanju njihovog ligninolitičkog enzimskog sistema, u cilju uspešne primene u različitim biotehnološkim procesima. Za razliku od G. lucidum, čiji je ligninolitički sistem intenzivno proučavan, ostale vrste ovog roda su retko bile objekti istraživanja. G. carnosum se teško morfološki razlikuje od G. lucidum. i još uvek nije biohemijski proučena, pa je cilj našeg istraživanja bio analiza produkcije Mn-oksidujućih peroksidaza i lakaza u uslovima čvrste kultivacije, na različitim biljnim ostacima (pšenična slama, stabljike kukuruza, piljevina hrasta i vinove loze) u prisustvu azota u obliku NH4NO3 i u koncentracijama od 10 mM, odnosno 20 mM. Maksimum produkcije Mn-zavisne peroksidaze dobijen je na pšeničnoj slami pri koncentraciji azota od 10 mM (56.82 U/l). Piljevina vinove loze i koncentracija azota od 10 mM bili su optimalni za sintezu verzatil peroksidaze (80.80 U/l). Dobijena aktivnost lakaza je bila izuzetno niska u medijumu sa pšeničnom slamom (1.80 U/l), dok na ostalim biljnim ostacima nije zabeležena. Maksimalni sadržaj ukupnih proteina je bio najveći u medijumu sa piljevinom hrasta kao izvorom ugljenika i koncentracijom azota od 10 mM.Projekat ministarstva br. 14304

Morpho-physiological characteristics and interactions of isolates of Mycogone perniciosa (Magnus) Delacr

Mycogone perniciosa (Magnus) Delacr., which causes wet bubble disease of Agaricus bisporus Lange (Imb), results in a considerable crop loss on mushroom farms in Serbia. The isolation and identification of five isolates of M. perniciosa from diseased fruit bodies of white button mushroom from mushroom units in Serbia, Bosnia and Herzegovina and Holland were made. Morpho-physiological characteristics and inter-relationships of the obtained isolates were studied. Macroscopic and microscopic investigations of different zones between colonies of the isolates of M. perniciosa revealed the phenomenon of the hyphal interference between different isolates. The obtained results suggest that hyphal interference could serve as an additional parameter for a more reliable determination of fungal specifity.Mycogone perniciosa (Magnus) Delacr., izazivač oboljenja mokre truleži najčešći je uzročnik gubitaka u gajilištima Agaricus bisporus Lange (Imb) u Srbiji. Izvršena je izolacija i identifikacija 5 izolata M. perniciosa sa obolelih plodonosnih tela šampinjona iz gajilišta u Srbiji, Bosni i Hercegovini i Holandiji. Ispitivane su morfo-fiziološke karakteristike kao i stepen srodnosti proučavanih izolata na osnovu analize međusobnog delovanja kolonija, odnosno korišćenjem fenomena hifalne interferencije. Makroskopska i mikroskopska istraživanja odnosa izolata i dobijeni rezultati ukazuju da hifalna interferencija može predstavljati dodatni parametar u razlikovanju izolata M. perniciosa. Izolati dobijeni iz gajilišta u Srbiji slični su međusobno, kao i izolati iz Holandije koji su pokazali međusobnu sličnost ali se razlikuju od prethodnih. Izolati iz Bosne i Hercegovine razlikovali su se i od srpskih i od holandskih.Projekat ministarstva br. 14304

The effect of Nepeta rtanjensis essential oil on test micromycetes mycelia growth

The antifungal activity of Nepeta rtanjensis Diklić et Milojević essential oil on mycelia growth has been performed by macrodilution method. The most efficient impact of N. rtanjensis essential oil on mycelia growth in vitro was found in Alternaria species with the same value of minimal inhibitory quantity (MIQ) of 0.6 μl/ml. Bipolaris spicifera and Cladosporium cladosporoioides had MIQ values of 1.0 μl/ml whereas Trichoderma viride with MIQ value of 1.6 μl/ml showed the most efficient defense against the essential oil examined. The values of minimal fungicidal quantity (MFQ) in Alternaria sp. 2, B. spicifera and C. cladosporioides match the MIQ values whereas MFQ values in Alternaria sp. 1 is 0.8 μl/ml, and in T. viride 1.8 μl/ml.Makrodilucionom metodom testiran je uticaj etarskog ulja endemične biljke Nepeta rtanjensis (Lamiaceae) na micelijalni rast mikromiceta: Alternaria sp. 1, Alternaria sp. 2, Bipolaris spicifera, Cladosporium cladosporioides i Trichoderma viride. Hemijska analiza etarskog ulja N. rtanjensis pokazala je apsolutnu dominaciju 4aα, 7α, 7aβ nepetalaktona (79.89%). Najefikasniji uticaj na rast micelije in vitro zabeležen je kod vrsta roda Alternaria sa istom vrednošću minimalne inhibitorne količine (MIK) od 0.6 μl/ml. Bipolaris spicifera i Cladosporium cladosporioides su imali MIK 1.0 μl/ml, dok je Trichoderma viride, sa vrednošću MIK od 1.6 μl/ml, pokazala najveću otpornost na dejstvo ispitivanog ulja. Vrednosti minimalne fungicidne količine (MFK) se kod Alternaria sp. 2 (0.6 μl/ml), B. spicifera (1.0 μl/ml) i C. cladosporioides (1.0 μl/ml) poklapaju sa vrednostima MIK, dok je kod Alternaria sp. 1 MFK 0.8 μl/ml, a kod T. viride 1.8 μl/ml. Etarsko ulje N. rtanjensis pokazalo je jako antifungalno dejstvo na rast micelije testiranih mikromiceta.nul