Extracellular enzymatic activity ofMicrosporum canis isolates

The enzymatic activity of 70 feline and canineMicrosporum canis isolates was determined by the Api-Zym^® test. The liquid phase of cultures, inoculated into Tryptic Soy Broth, was used to examine 19 enzymes. Considerable differences were observed among the extracellular enzymatic patterns. All the isolates produced alkaline phosphatase and beta-glucosidase, while lipase (C14), trypsin, chymotrypsin, beta-glucuronidase, and alpha-fucosidase activity was never revealed. Esterase (C4) activity was present in 57 samples (81%), esterase lipase (C8) in 31 (44%), leucine arylamidase in 35 (50%), valine arylamidase and cystine arylamidase in 7 (10%), acid phosphatase in 64 (91%), naphthol-AS-BI-phosphohydrolase in 60 (86%), alpha-galactosidase in 5 (7%), beta-galactosidase in 6 (8%), alpha-glucosidase in 25 (36%), N-acetyl-beta-glucosaminidase in 41 (58%), and alpha-mannosidase in 51 (73%). The beta-galactosidase activity ofM. canis has not been reported previously. Remarkable variations of intensity for each enzymatic activity were also detected. It is believed that these results could provide basic data for further investigations on the pathogenic role of enzymes secreted byM. canis.     

Extracellular proteinases of filamentous fungi as potential markers of phytopathogenesis

he presence of proteins in the culture liquid of filamentous fungi under study was found to induce the secretion of proteinases. The inhibitory analysis of the major extracellular proteinases of the saprotrophic fungus Trichoderma harzianum and the phytopathogenic fungus Alternaria alternata showed that they both belong to the group of serine proteinases. The substrate specificity of these proteinases and their sensitivity to inhibitors suggest that the enzyme of T. harzianum is a subtilisin-like proteinase and the enzyme of A. alternata is a trypsin-like proteinase. This difference between the proteinases may reflect the physiological difference between their producers (saprotroph and phytopathogen).     

Seasonal periodicity of aquatic fungi in tanks at Kurukshetra,India

The occurrence, distribution and seasonal periodicity of 49 species of aquatic fungi belonging to the orders Chytridiales, Lagenidiales, Blastocladiales, Saprolegniales and Peronosporales from two tanks of Kurukshetra, India, is described. Seasonal periodicity was found to be governed by temperature.     

Increased heat tolerance afforded by oil-based conidial formulations of Metarhizium anisopliae and Metarhizium robertsii

The thermotolerance of oil-based conidial formulations of Metarhizium anisopliae s.l. (IP 46) and Metarhizium robertsii (ARSEF 2575) were investigated. Conidia of IP 46 or ARSEF 2575 were suspended in different adjuvants and exposed to 45?±?0.2°C for 4, 6, 8 or 24?h; their viability was then assessed after 48?h incubation at 27?±?1°C. Conidia heated in pure mineral or vegetable oil exhibited mean relative viability exceeding 70% after 8?h of heat exposure, whereas low germination (≤20%) was observed when conidia were heated in water (Tween 80^® 0.01%), carboxymethyl cellulose gel or emulsifiable oils (Graxol^® or Assist^®) and exposed to heat for 6 or 8?h. In addition, conidia of IP 46 suspended in either pure mineral or canola oil and exposed to heat for 48?h had moderate viability, 57% or 41%, respectively. Unstable oil-in-water emulsions showed a higher percentage of conidia incorporated into oil micellae, while the stable emulsions had higher percentage of conidia outside the oil micellae. The thermotolerance of conidia formulated in stable emulsions, however, did not differ from that of conidia formulated in unstable emulsions. The present study highlights possibilities to alleviate the deleterious effects of heat stress towards Metarhizium spp. conidia applied for controlling arthropod pests and vectors through oil-based formulations.     

Diversity and activity of aquatic fungi under low oxygen conditions

1. The objective was to test whether a decrease in oxygen concentration in streams affects the diversity and activity of aquatic hyphomycetes and consequently leaf litter decomposition. 2. Senescent leaves of Alnus glutinosa were immersed for 7 days in a reference stream, for fungal colonization, and then incubated for 18 days in microcosms at five oxygen concentrations (4%, 26%, 54%, 76% and 94% saturation). Leaf decomposition (as loss of leaf toughness), fungal diversity, reproduction (as spore production) and biomass (ergosterol content) were determined. 3. Leaf toughness decreased by 70% in leaves exposed to the highest O₂ concentration, whereas the decrease was substantially less (from 25% to 45%) in treatments with lower O₂. Fungal biomass decreased from 99 to 12 mg fungi g⁻¹ ash‐free dry mass on exposure to 94% and 4% O₂ respectively. Sporulation was strongly inhibited by reduction of dissolved O₂ in water (3.1 × 10⁴ versus 1.3 × 10³ spores per microcosms) for 94% and 4% saturation respectively. 4. A total of 20 species of aquatic hyphomycetes were identified on leaves exposed to 94% O₂, whereas only 12 species were found in the treatment with 4% O₂ saturation. Multidimensional scaling revealed that fungal assemblages exposed to 4% O₂ were separated from all the others. Articulospora tetracladia, Cylindrocarpon sp. and Flagellospora curta were the dominant species in microcosms with 4% O₂, while Flagellospora curvula and Anguillospora filiformis were dominant at higher O₂ concentrations. 5. Overall results suggest that the functional role of aquatic hyphomycetes as decomposers of leaf litter is limited when the concentration of dissolved oxygen in streams is low.     

Extracellular and cellular mechanisms sustaining metal tolerance in ectomycorrhizal fungi

This review focuses on recent evidence that identifies potential extracellular and cellular mechanisms that may be involved in the tolerance of ectomycorrhizal fungi to excess metals in their environment. It appears likely that mechanisms described in the nonmycorrhizal fungal species are used in the ectomycorrhizal fungi as well. These include mechanisms that reduce uptake of metals into the cytosol by extracellular chelation through extruded ligands and binding onto cell-wall components. Intracellular chelation of metals in the cytosol by a range of ligands (glutathione, metallothioneins), or increased efflux from the cytosol out of the cell or into sequestering compartments are also key mechanisms conferring tolerance. Free-radical scavenging capacities through the activity of superoxide dismutase or production of glutathione add another line of defence against the toxic effect of metals.     

Conidial water affinity is an important characteristic for thermotolerance in entomopathogenic fungi

The interspecific thermotolerance of several species of entomopathogenic fungi was evaluated based on the conidial water affinity. The species were divided between hydrophilic and hydrophobic conidia. The species with hydrophobic conidia were Beauveria bassiana (ARSEF 252), Metarhizium brunneum (ARSEF 1187), Metarhizium robertsii (ARSEF 2575), Isaria fumosorosea (ARSEF 3889) and Metarhizium anisopliae s.l. (ARSEF 5749). The species with hydrophilic conidia were Tolypocladium cylindrosporum (ARSEF 3392), Tolypocladium inflatum (ARSEF 4877), Simplicillium lanosoniveum (ARSEF 6430), Lecanicillium aphanocladii (ARSEF 6433), S. lanosoniveum (ARSEF 6651), Aschersonia placenta (ARSEF 7637) and Aschersonia aleyrodis (ARSEF 10276). The conidial surface tension of each isolate was also studied. Conidial suspensions were exposed to 38, 41 or 45 °C. After exposure, the suspensions were inoculated on media and conidial germination was evaluated. Considerable differences in thermotolerance were found among the 12 entomopathogenic fungal species. Species with hydrophobic conidia were generally more thermotolerant than species with hydrophilic conidia. All isolates with hydrophobic conidia showed higher conidial surface tension than the isolates with hydrophilic conidia.     

Higher temperature reduces the effects of litter quality on decomposition by aquatic fungi

1. We investigated the effects of riparian plant diversity (species number and identity) and temperature on microbially mediated leaf decomposition by assessing fungal biodiversity, fungal reproduction and leaf mass loss. 2. Leaves of five riparian plant species were first immersed in a stream to allow microbial colonisation and were then exposed, alone or in all possible combinations, at 16 or 24 °C in laboratory microcosms. 3. Fungal biodiversity was reduced by temperature but was not affected by litter diversity. Temperature altered fungal community composition with species of warmer climate, such as Lunulospora curvula, becoming dominant. 4. Fungal reproduction was affected by litter diversity, but not by temperature. Fungal reproduction in leaf mixtures did not differ or was lower than that expected from the weighted sum of fungal sporulation on individual leaf species. At the higher temperature, the negative effect of litter diversity on fungal reproduction decreased with the number of leaf species. 5. Leaf mass loss was affected by the identity of leaf mixtures (i.e. litter quality), but not by leaf species number. This was mainly explained by the negative correlation between leaf decomposition and initial lignin concentration of leaves. 6. At 24 °C, the negative effects of lignin on microbially mediated leaf decomposition diminished, suggesting that higher temperatures may weaken the effects of litter quality on plant litter decomposition in streams. 7. The reduction in the negative effects of lignin at the higher temperature resulted in an increased microbially mediated litter decomposition, which may favour invertebrate‐mediated litter decomposition leading to a depletion of litter stocks in streams.     

Biochemical properties of different entomopathogenic fungi and their virulence against Chilo suppressalis (Lepidoptera: Crambidae) larvae

We determined the virulence of Beauveria bassiana, Metarhizium anisopliae, Isaria fumosoroseus and Lecanicilium lecanii against larvae of Chilo suppressalis Walker by bioassay and evaluated several enzymatic and non-enzymatic components. LC₅₀ values of the entomopathogenic fungi revealed 90, 32, 45,000, 4600, 42,000 and 1,540,000 spores/larva for isolates BB1–BB3 of B. bassiana, I. fumosoroseus, M. anisopliae and L. lecanii, respectively. Isolate BB3 and I. fumosoroseus had the highest amounts of total protein and hydrophobin and isolates BB3 and M. anisopliae showed the highest activities of lipases and chitinases. In case of proteases, the highest activities were observed for Pr1 of BB1 and Pr2 of L. lecanii. The highest general esterase activities were obtained in I. fumosoroseus and BB1 when 1-naphtyl acetate and 2-naphtyl acetate were used as substrates. The highest activity of glutathione S-transferase (GST) was observed in I. fumosoroseus by using both reagents but BB1 demonstrated the highest activities of alkaline phosphatase and acid phosphatase. Clustering of the fungi using biochemical enzymes revealed BB2 and BB3 as a separate group of entomopathogenic fungi. In another group, I. fumosoroseus and L. lecanii had the most similarity and were separated from BB1 and M. anisopliae. The fungi exhibited different virulence on larvae of C. suppressalis by producing adhering protein and extracellular enzymes. Overall, results of the bioassays and clustering based on enzymatic activities revealed that isolate BB2 was the most effective fungus against larvae of C. suppressalis.     

Extracellular enzymatic activity profiles in fungi isolated from oil-rich environments

About 34 wild fungal species associated with edible oil mill wastes were isolated by the serial dilution technique. Methods for rapid screening of fungal species against production of extracellular enzymes such as amylase, protease, cellulase, and lipase are reported. Among all the species, Aspergillus versicolor exhibited high amlylolytic and gelatinolytic activity, whereas Penicillium citrinum showed only high amylolytic activity. Maximum cellulolytic activity was recorded for Absidia corymbifera, As. niger, Cunninghamella echinulata, Curvularia lunata, Fusarium solani, Mucor racemosus, Paecilomyces variotii, and Syncephalastrum racemosum. The fungal species Ab. corymbifera, As. fumigatus, As. japonicus, As. nidulans, As. terreus, Cun. verticillata, Cur. pallescens, F. oxysporum, Geotrichum candidum, M. racemosus, Pe. citrinum, Pe. frequentans, Rhizopus stolonifer, and Trichoderma viride exhibited maximum lipase activity. This study confirms the isolated fungi present on a wide range of substrates in the ambient environment, and these results could provide basic data for further investigations on fungal extracellular enzymes.First two authors equally contributed to this work     

Notes on Algerian aquatic fungi: Pythium pythioides

Pythium pythioides (Roze & Cornu) Ramsbottom is recorded from Algeria. Since its original discovery in 1869 this species had not been found back. A brief history is given and morphological and reproductive characters are discussed.     

澜沧江两条支流捕食线虫真菌水陆分布差异及关联性

为探究微生物在流域中的水、陆分布差异和相互关系,对澜沧江两条支流捕食线虫真菌多样性水陆分布进行了调查研究。在枯水期,以澜沧江一级支流沘江和黑惠江为研究区域,系统布设12个采样点(水流交汇点),每个采样点采集水、陆对称样品各5份,共采集土样120份。结合传统分离纯化、形态学及分子生物学方法筛选和鉴定菌株,按照《Nematode-Trapping Fungi》进行分类,共获得2属13种88株捕食线虫真菌;其中,陆地样品中共分离到2属11种45株,水体底泥中分离出1属10种43株,检出率分别为41.67%和53.33%。结果表明,澜沧江两条支流捕食线虫真菌在物种、属、群落结构3个层面上均存在水陆差异,也相互联系;陆地土壤可能是流域内捕食线虫真菌多样性的源,水流是其重要的传播因子。在流域生态系统内,水陆间的扩散限制和水流的连通性都是维持微生物物种多样性的重要机制。     

Impact of sixteen chemical pesticides on conidial germination of two entomophthoralean fungi: Conidiobolus thromboides and Pandora nouryi

Effects of sixteen chemical pesticides on conidial germination of C. thromboides and P. nouryi were investigated at two concentrations (R and 0.2R, where R=the lowest manufacture's recommended concentration for field use). Of the fungicides tested, propamocarb hydrochloride did not significantly affect conidial germination of C. thromboides, but at R it completely inhibited conidial germination of P. nouryi. At 0.2R of procymidone, iprodione, haloxyfop-methyl and fenoxaprop-p-ethyl, conidia of C. thromboides had some germination after 24 h, although they entirely restricted conidial germination of the fungus at the concentration of R. Tribenuron-methyl, one herbicide, had no adverse effect on conidial germination of C. thromboides and P. nouryi after 6 h at the two concentrations, whereas the other seven herbicides were strongly antagonistic to P. nouryi, except for carfentrazone-ethyl at 0.2R.     

亚热带森林根系和菌根真菌调控凋落叶分解和主场效应

根系向凋落物层生长是森林生态系统中存在的普遍现象,研究根系和菌根真菌如何调控凋落物分解对理解森林生态系统的养分循环具有重要意义。通过在亚热带楠木(丛枝菌根树种)和格氏栲(外生菌根树种)人工林中进行了2年的凋落物互换和菌根排除实验,研究根系及其共生菌根真菌如何影响森林凋落物的分解速率和主场效应。结果发现:经过720 d的分解,在楠木人工林中,根系的存在促使楠木和格氏栲凋落物的分解速度分别提高了29%和47%,而在格氏栲人工林中,根系存在对格氏栲凋落物分解无显著影响,但增加了楠木凋落物的分解速率。此外,楠木人工林中,有根和无根处理下凋落物分解均无主场效应,而格氏栲人工林凋落物仅在根系存在的情况下显示出负的主场效应。进一步发现格氏栲凋落物的负主场效应归因于楠木人工林凋落物层更大的腐生真菌丰度和更强的磷降解酶活性。结果表明,在亚热带不同菌根类型树种中,菌根真菌与腐生真菌的相互作用(拮抗或促进),很有可能通过影响磷降解酶的活性,进而影响凋落物的分解率和主场效应。     

Possible relation between phytoplankton numbers and Saprolegnioid fungi in Shatt Al-Arab near Basrah,Iraq

This study, carried out in Shatt Al-Arab river near Basrah, Iraq, reports a close negative (inverse) relation between phytoplankton numbers and decomposing fungi belonging to the family Saprolegniaceae (Oomycetes). This relation seems to depend on substrate relationship and antagonism. In addition the role of other physico-chemical factors, pH, temperature and salinity has been discussed.     

Anaerobic fungi and their cellulolytic and xylanolytic enzymes

Anaerobic fungi are the inhabitants of the digestive tract of herbivorous mammals, ruminants as well as non-ruminants. One of the major characteristics of all anaerobic fungi examined thus far, is their production and secretion of a range of polysaccharide-degrading enzymes, including cellulases, xylanases and glucoside-hydrolases. The cellulolytic enzymes of the anaerobic fungusNeocallimastix frontalis have been shown to possess a high activity. Therefore anaerobic fungi and/or their enzymes could be interesting for many biotechnological applications including saccharafication of lignocellulosic residues, production of polysacchari-dehydrolysing enzymes. This review summarizes the present knowledge of anaerobic fungi with special emphasis on their cellulolytic and xylanolytic enzymes. Further, a comparison with aerobic fungi is made.Abbreviations G₂ cellobiose - G₃ cellotriose - G₄ cellotetraose - G₅ cellopentaose - HMM-complex high molecular mass complex - PNPF p-nitrophenyl--fucopyranoside - PNPG p-nitrophenyl--glucopyranoside     

Extracellular phosphatase activity in sediments of the Breitenbach,a Central European mountain stream

Activity of extracellular phosphatases (phosphomonoesterases) was measured in sandy streambed sediments of the Breitenbach, a small unpolluted upland stream in Central Germany. Fluorigenic 4-methylumbelliferyl phosphate served as a model substrate. Experiments were conducted using sediment cores in a laboratory simulation of diffuse groundwater discharge through the stream bed, a natural process occurring in the Breitenbach as well as many other streams.Streambed sediments contained high levels of particulate phosphorus, but concentrations of dissolved phosphorus in the interstitial water were 3 to 4 orders of magnitude lower. These interstitial concentrations were similar to those in the stream and groundwater. Extracellular phosphatase activity was high in the streambed sediments. These enzymes probably contribute significantly to the flux of phosphorus in sediment by hydrolyzing phosphomonoesters, making free phosphate available to the sediment microorganisms.Factors influencing the kinetic parameters V _max (maximum activity) and apparent K _m (enzyme affinity) of phosphatase were discharge rates of water through the sediment, water quality (ground- or stream water), and substrate (phosphomonoesters) as well as dissolved ortho-phosphate concentrations. Enzymes are supposed to be effective at limiting substrate concentrations, where, in this study, changes in discharge rates had little influence on rates of hydrolysis. Higher V _max and lower K _m values were found during percolation of groundwater through the sediment cores, compared with stream water. This indicates that rates of hydrolysis were higher with groundwater, both at substrate limitation and at substrate saturation. This was probably a consequence of the lower levels of dissolved ortho-phosphate in the groundwater.