Growth and Characteristics of Two Different Epichloë sinensis Strains Under Different Cultures

In the present study, two Epichloë sinensis endophyte strains isolated from different Festuca sinensis ecotypes were inoculated on potato dextrose agar (PDA) and potato dextrose broth (PDB) media with or without (control) exogenous additives. After 4weeks of growth, the growth (colony diameter, hyphal diameter, and mycelial biomass) and other characteristics (pH and antioxidant capacity of culture filtrate, mycelial ion contents, and hormone contents) were measured. The results showed that the culture conditions had significant effects (p<0.05) on the hyphal diameter, mycelial biomass, and hormone content of the two strains. The mycelial biomass of the two strains in PDB was significantly higher (p<0.05) than that on PDA. Except for strain 1 with indole-3-acetic acid (IAA) treatment and strain 84F with control and VB1 treatments, the hyphal diameter of the two strains in PDB under the other treatments was significantly higher (p<0.05) than that on PDA. In most cases, the IAA, cytokinins (CTK), abscisic acid (ABA), and gibberlic acid (GA) contents in the mycelia on PDA of the two strains were significantly higher (p<0.05) than those in PDB. The two E. sinensis strains exhibited significantly different performances (p<0.05) under the five treatments. The indices, including colony diameter, mycelial biomass, scavenging ability of superoxide anion radicals and hydroxyl radicals, pH of culture filtrate, ion contents, hyphal diameter, and IAA, CTK, GA, and ABA contents were significantly different (p<0.05) between the two strains, although the performance was inconsistent. Exogenous additives had significant effects (p<0.05) on the performance of the two E. sinensis strains. Indole-3-acetic acid and VB1 treatments significantly promoted (p<0.05) the growth of the two strains on both PDA and PDB. Indole-3-acetic acid treatment also significantly increased the hyphal diameters of the two strains in PDB (p<0.05). Indole-3-acetic acid and VB1 treatments significantly reduced (p<0.05) the antioxidant ability of these two strains in PDB. NaCl and ZnCl2 treatments had significant inhibitory effects (p<0.05) on fungal growth and promotion effects on the antioxidant ability of the two strains. The treatments also had significant effects (p<0.05) on hyphal diameters and ion and hormone contents, although the effects varied with different indices.

Morpho-molecular characterization of Ceratobasidium sp.: A mycorrhizal fungi isolated from a rare epiphytic orchid Gastrochilus calceolaris (J. E. Sm.) D. Don

A mycorrhizal fungus, Ceratobasidium sp. GC (NCBI Gene Bank Accession no GQ369961) associated with the roots of an epiphytic orchid Gastrochilus calceolaris was investigated by cultural morphology, microscopic features and molecular analysis of Internal Transcribed Spacer (ITS) regions sequences of nuclear ribosomal DNA. The colony appearance of the fungal endophyte was fluffy growth pattern and the colour of the young colony was milky white on both surfaces that turned in to brown at maturity on the upper surface and deep brown on reverse surface. The microscopic features of the fungus i.e. hyphal diameter, multi-nucleate vegetative cells, right angle branching pattern with slight constriction at branching point and a dolipore septum near the branching point were observed. All the characters corroborated the identity with anamorphic Rhizoctonia like fungi. The ITS regions sequences of nrDNA and phylogenetic analysis based on the neighbor-joining method showed clustered with Rhizoctonia like fungi, and the maximum identity (98.28%) with Ceratobasidium RR and Ceratobasidium FPUB isolated from Rhynchostylis retusa and Dactylorrhiza hetagera, respectively. Thus, the ITS of nrDNA sequences validated the morphological data. This is the first report of orchid mycorrhizal fungi from Bangladesh.

Tradeoffs in hyphal traits determine mycelium architecture in saprobic fungi

The fungal mycelium represents the essence of the fungal lifestyle, and understanding how a mycelium is constructed is of fundamental importance in fungal biology and ecology. Previous studies have examined initial developmental patterns or focused on a few strains, often mutants of model species, and frequently grown under non-harmonized growth conditions; these factors currently collectively hamper systematic insights into rules of mycelium architecture. To address this, we here use a broader suite of fungi (31 species including members of the Ascomycota, Basidiomycota and Mucoromycota), all isolated from the same soil, and tested for ten architectural traits under standardized laboratory conditions. We find great variability in traits among the saprobic fungal species, and detect several clear tradeoffs in mycelial architecture, for example between internodal length and hyphal diameter. Within the constraints so identified, we document otherwise great versatility in mycelium architecture in this set of fungi, and there was no evidence of trait ‘syndromes’ as might be expected. Our results point to an important dimension of fungal properties with likely consequences for coexistence within local communities, as well as for functional complementarity (e.g. decomposition, soil aggregation).

Opportunistic Fungal Infections in Small Animals

ABSTRACT Opportunistic fungal infections have long been recognized as rare causes of disease in immunocompetent dogs and cats. Recently, the escalating use of multiagent immunosuppression protocols (especially those that include cyclosporine) has resulted in an increased number of patients with opportunistic fungal infection encountered by small animal practitioners and has altered the typical case phenotype. Based on histologic and cytologic features such as pigmentation, hyphal diameter, and distribution in tissue, these opportunistic mycoses can be placed into categories such as phaeohyphomycosis, hyalohyphomycosis, and eumycotic mycetoma. This review aims to summarize the clinical presentations, methods for diagnosis, treatment recommendations, and prognosis for both immunocompetent and immunosuppressed patients with opportunistic fungal infections. An example case description is included to illustrate the most common current clinical presentation.

How to build a mycelium: tradeoffs in fungal architectural traits

The fungal mycelium represents the essence of the fungal lifestyle, and understanding how a mycelium is constructed is of fundamental importance in fungal biology and ecology. Previous studies have examined initial developmental patterns or focused on a few strains, often mutants of model species, and frequently grown under non-harmonized growth conditions; these factors currently collectively hamper systematic insights into rules of mycelium architecture. To address this, we here use a broader suite of fungi (31 species including members of the Ascomycota, Basidiomycota and Mucoromycotina), all isolated from the same soil, and test for ten architectural traits under standardized laboratory conditions.We find great variability in traits among the saprobic fungal species, and detect several clear tradeoffs in mycelial architecture, for example between internodal length and hyphal diameter. Within the constraints so identified, we document otherwise great versatility in mycelium architecture in this set of fungi, and there was no evidence of trait ‘syndromes’ as might be expected.Our results point to an important dimension of fungal properties with likely consequences for coexistence within local communities, as well as for functional complementarity (e.g. decomposition, soil aggregation).

First Report of Web Blight on Rose Campion (Lychnis coronaria) Caused by Rhizoctonia solani AG 1-IB in Italy

Lychnis coronaria (syn. Silene coronaria), rose campion, is a perennial in the Caryophyllaceae used in gardens. In the summer of 2014, a web blight was observed in a private garden located near Biella (northern Italy), approximately 45°39′N 8°00′E, on 40% of 100 5-month-old plants grown in sandy soil. In the days preceding the outbreak of the disease, daytime temperatures ranged from 18 to 24°C and relative humidity from 45 to 83%. Affected plants showed pale brown discoloration of stems, starting from the base, and eventually collapsed. Under conditions of high relative humidity, a scant amount of whitish mycelium developed on leaves of about 50% of diseased plants. Eventually, infected plants died about 10 days after symptoms appeared. Symptomatic tissues of stems and leaves were disinfected for 10 s in 1% NaOCl, rinsed in sterile water, and plated on potato dextrose agar (PDA). A fungus with the morphological characters of Rhizoctonia solani (3) was consistently recovered. Three representative isolates were paired with tester strains of R. solani (AG 1, AG 2-2-IIIB, AG 4, AG 7, and AG 11) (2) and examined microscopically. The Lychnis isolates anastomosed only with the AG 1 tester strain, with low fusion frequency. The anastomosis point was obvious: the hyphal diameter at the point of anastomosis was reduced and death of adjacent cells was observed, indicating an anastomosis reaction (1). Mycelium maintained on PDA at 23 ± 1°C was coarse and reddish brown. After 5 days of growth, mycelium started differentiating numerous sclerotia, often aggregated. Mature sclerotia were dark, spheroidal, with diameters ranging from 0.2 to 1.6 (average 0.6) mm. The internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS1/ITS4 and sequenced. BLASTn analysis of the 609-bp amplicon (GenBank Accession No. KM596899) showed a 98% homology with the sequence of the R. solani isolate FJ746937 obtained from Zoysiagrass. On the basis of molecular and cultural characteristics and anastomosis tests, the isolates from L. coronaria were identified as R. solani AG 1-IB (4). For pathogenicity tests performed in August, mycelial disks (8 mm diam.) from 10-day-old PDA cultures of an isolate of the fungus were placed on four healthy 6-month-old L. coronaria plants (four stem and six leaf disks per plant). Four plants inoculated with disks of PDA served as controls. Plants were covered with plastic bags for 4 days and maintained in a garden located in the same area in which the disease appeared, at field temperatures ranging from 15 to 28°C. The first symptoms developed 4 days after inoculation, and 15 days after the artificial inoculation, all inoculated plants were dead. R. solani was re-isolated from the stem of symptomatic plants, whereas no colonies developed from controls, which all remained healthy. This is the first report of blight of L. coronaria caused by R. solani in Italy or anywhere else in the world. The impact of this disease may become a significant problem for L. coronaria, a very common species in Italian gardens. References: (1) D. E. Carling. Page 37 in: Rhizoctonia Species: Taxonomy, Molecular Biology, Ecology, Pathology and Disease Control. Kluwer Academic Publishers, The Netherlands, 1996. (2) A. Ogoshi. Ann. Rev. Phytopathol. 25:125, 1987. (3) B. Sneh et al. Identification of Rhizoctonia species. APS Press, St. Paul, MN, 1991. (4) R. T. Sherwood. Phytopathology 59:1924, 1969.

Alternaria alternata transcription factor CmrA controls melanization and spore development

Melanin is a black pigment widely distributed across the kingdoms, from bacterial to human. The filamentous fungus Alternaria alternata is a typical ‘black fungus’, which produces melanin in its hyphal and especially its asexual spore cell walls. Its biosynthesis follows the dihydroxynaphthalene (DHN) pathway with 1,8-DHN as an intermediate. Two genes, encoding a polyketide synthase (pksA) and a 1,3,8-trihydroxynaphthalene (THN) reductase (brm2), along with a putative transcription factor, CmrA, comprise a small gene cluster. Here we show that CmrA controls the expression of pksA and brm2, but that it also controls the expression of a scytalone dehydratase encoding gene (brm1) located elsewhere in the genome. The regulatory function of CmrA was shown in a reporter assay system. Al. alternata CmrA was expressed in the filamentous fungus Aspergillus nidulans where it was able to induce the expression of a reporter construct under the control of the putative pksA promoter. This suggests direct binding of CmrA to the promoter of pksA in the heterologous system. Likewise, silencing of cmrA in Al. alternata led to white colonies due to the lack of melanin. In addition, hyphal diameter and spore morphology were changed in the mutant and the number of spores reduced. Silencing of brm2 and inhibition of melanin biosynthesis by tricyclazole largely phenocopied the effects of cmrA silencing, suggesting a novel regulatory function of melanin in morphogenetic pathways.

Rhizoctonia cerealis anastomosis group GAG-1, the common pathogen of wheat, barley and sugar beet

Isuluies of <i>Rhizoctonia cerealis</i> anastomosis group GAG-1 were obtained from sharp eyespot lesions on wheat and on barley culms and from diseased sugar beet seedlings. Isolates of <i>R. cerealis</i> were collected from a fields with crop rotation experiments: sugar beet-spring wheat-winter barley. In pathogenicity tests isolates of <i>R. cerealis</i> from sugar beet seedlings and from sharp eyespot lesions on wheat and barley were pathogenic to these crops. Isolates of <i>R. cerealis</i> from sharp eyespot lesions on wheat and barley caused severe damping-ofTof sugar beet. Isolates of <i>R. cerealis</i> from sugar beet seedlings also caused symptoms of sharp eyespot on wheat and barley. None of the wheat and barley isolates of <i>R. cerealis</i> tested caused root-rot on wheat or barley seedlings. Isolates of <i>R. cerealis</i> obtained from diseased plants of wheat, barley and sugar beet were similar in morphology of cultures and anastomosed with GAG-1 tester isolate. The relatinoship between anastomosis. colony characters, growth rate, hyphal diameter and pathogenicity of AG-4. AG-2-2 and AG-5 isolates obtained together with <i>R. cerealis</i> from diseased plants were also investigated.

First Report of Web Blight on Oregano (Origanum vulgare L.) Caused by Rhizoctonia solani AG-1-IB in Italy

Origanum vulgare L., common name oregano, family Labiatae, is grown for its aromatic and medicinal properties and as ornamental. In the fall of 2012, a blight was observed in a farm located near Albenga (northern Italy) on 6% of 30,000 50-day-old plants, grown in trays in a peat/perlite mix. Semicircular, water soaked lesions appeared on leaves and stems, starting from the basal ones. As the disease progressed, blighted leaves turned brown, withered, clung to the shoots, and matted on the surrounding foliage. Eventually, infected plants died. Leaf and stem fragments taken from the margin of the diseased tissues belonging to 10 plants were disinfected for 10 s in 1% NaOCl, rinsed with sterile water, and plated on potato dextrose agar (PDA). A fungus with the morphological characters of Rhizoctonia solani was consistently recovered. Three isolates of R. solani obtained from affected plants were successfully anastomosed with R. solani isolate AG 1 (ATCC 58946). Three pairings were made for each tester strain. The hyphal diameter at the point of anastomosis was reduced, the anastomosis point was obvious, and death of adjacent cells was observed. Results were consistent with other reports on anastomosis reactions (2). Isolates from oregano were paired with R. solani isolates AG 2, 3, 4, 6, 7, or 11 and examined microscopically. Anastomosis was not observed in any of the pairings. Tests were conducted twice. Mycelium of 10-day-old isolates from oregano appeared reddish brown, coarse, and radiate. Numerous dark brown sclerotia, 0.3 to 1.0 mm diameter (average 0.7) developed within 10 days after transfer of mycelia to PDA in 90 mm diameter petri dishes at 21 to 24°C. The descriptions of mycelium and sclerotia were typical for subgroup IB Type 1 (4). The internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS1/ITS4 and sequenced. BLASTn analysis (1) of the 538 bp showed a 99% homology with the sequence of R. solani FJ746937, confirming the morphological identification of the species. The nucleotide sequence has been assigned the GenBank Accession KC493638. For pathogenicity tests, one of the isolates assigned to the anastomosis group AG-1-IB was tested by placing 9 mm diameter mycelial disks removed from PDA 10-day-old cultures of the fungus on leaves of 90-day-old oregano plants (n = 35). Thirty-five plants inoculated with non-inoculated PDA disks served as controls. Plants were covered with plastic bags and maintained in a growth chamber at 25 ± 1°C with 12 h light/dark. The first symptoms, similar to those observed in the farm, developed 3 days after inoculation. Nine days after the artificial inoculation, 50% of plants were dead. About 10 colonies of R. solani were reisolated from infected leaves of inoculated plants. Control plants remained healthy. The pathogenicity test was carried out twice with similar results. Symptoms caused by R. solani have been recently observed on O. vulgare in Greece (3). This is, to our knowledge, the first report of blight of O. vulgare caused by R. solani in Italy. References: (1) S. F. Altschul et al. Nucleic Acids Res., 25:3389, 1997. (2) D. E. Carling. Grouping in Rhizoctonia solani by hyphal anastomosis reactions. In: Rhizoctonia Species: Taxonomy, Molecular Biology, Ecology, Pathology and Disease control. Kluwer Academic Publishers, The Netherlands, pp. 37-47, 1996. (3) C. D. Holevas et al. Benaki Phytopathol. Inst., Kiphissia, Athens, 19:1-96, 2000. (4) R. T. Sherwood. Phytopathology 59:1924, 1969.