Isolation and identification of mycorrhizal fungi associating with an achlorophyllous plant, Epipogium roseum (Orchidaceae)

The identity of mycorrhizal fungi associated with the achlorophyllous orchid Epipogium roseum was investigated by DNA analysis. The fungi were isolated from each coiled hypha (peloton), and the ITS region of nuclear rDNA was sequenced. Phylogenetic analysis based on the neighbor-joining method showed that all the isolates clustered with fungi belonging to Psathyrella or Coprinus in Coprinaceae. Those fungi are known as saprobes, using dead organic materials for a nutritive source. Large colonies of this orchid were frequently found around tree stumps or fallen logs. In such colonies, these decaying wood materials would be used as a large and persistent carbon source for the growth of this orchid. This is the first report of Coprinaceae as an orchid mycorrhizal fungi.     

A study of fungal spore poupulations in the atmosphere of Kuwait

The fungal air-spora of Kuwait was investigated using the Petri-dish trapping technique. During the period from 1st April, 1974 to 30th June, 1975, a total of 3685 colonies were recorded from 2 % malt agar plates. Fifty-five genera and 116 species were identified. Alternaria occupied the first place in the order of percentage incidence, being represented by 18.3% of the entire catch, followed by Aspergillus (17.1%), Penicillium (14%), Cladosporium (13.6%), Drechslera (13.3%), and Ulocladium (7.1%). The widest spectrum of species was displayed by Aspergillus (possessed 19 species) followed by Penicillium (17 species), Alternaria (6 species), Drechslera (5 species), and Ulocladium (4 species). The monthly total number of fungi showed marked seasonal periodicity with the greatest number of colonies occurring in March–April and November. The lowest counts were recorded in mid-summer (July) and mid-winter (January). Only 220 colonies and 17 species of thermophilous fungi were reported from plates incubated at 45 °C. The most common species were Aspergillus fumigatus and A. terreus.     

The fungal Flora of the air near the ground in San Antonio,Texas

Summary Daily exposure of petri dishes containing Littman's oxgall medium were used to survey the incidence of air-borne fungi in San Antonio. A total of 7026 colonies was obtained. Hormodendrum andAlternaria were found to be the most common fungi present. About 80 genera were isolated, including the yeast-like fungi.No direct correlation was found between the seasonal pattern of theHormodendrum, Alternaria or the total counts, and any single environmental condition in the area. It seems as if all factors combine including the long summer months of the area to control the number of viable spores in the air.This study was supported by U.S.P.H.S. grant 5-ROL-HE-3834-07.     

Fungal Epiphytes and Endophytes of Coffee Leaves (Coffea arabica)

Plants harbor diverse communities of fungi and other microorganisms. Fungi are known to occur both on plant surfaces (epiphytes) and inside plant tissues (endophytes), but the two communities have rarely been compared. We compared epiphytic and endophytic fungal communities associated with leaves of coffee (Coffea arabica) in Puerto Rico. We asked whether the dominant fungi are the same in both communities, whether endophyte and epiphyte communities are equally diverse, and whether epiphytes and endophytes exhibit similar patterns of spatial heterogeneity among sites. Leaves of naturalized coffee plants were collected from six sites in Puerto Rico. Epiphytic and endophytic fungi were isolated by placing leaf pieces on potato dextrose agar without and with surface sterilization, respectively. A total of 821 colonies were isolated and grouped into 131 morphospecies. The taxonomic affinities of the four most common nonsporulating fungi were determined by sequencing the nuclear ribosomal internal transcribed spacer (ITS) region: two grouped with Xylaria and one each with Botryosphaeria and Guignardia. Of the most common genera, Pestalotia and Botryosphaeria were significantly more common as epiphytes; Colletotrichum, Xylaria, and Guignardia were significantly more common as endophytes. Suprisingly, more morphospecies occurred as endophytes than as epiphytes. Differences among sites in number of fungi per plant were significant. Thus epiphytic and endophytic communities differed greatly on a single leaf, despite living only millimeters apart, and both communities differed from site to site. Significant correlations between occurrence of fungal morphospecies suggested that fungi may have positive or negative effects on their neighbors. This is the first quantitative comparison of epiphytic and endophytic fungal floras in any plant, and the first to examine endophytic fungi or epiphytic fungi in leaves of coffee, one of the world’s most valuable crops.     

Comparison of four media for the isolation ofAspergillus flavus group fungi

Four agar media used to isolate aflatoxin producing fungi were compared for utility in isolating fungi in theAspergillus flavus group from agricultural soils collected in 15 fields and four states in the southern United States. The four media wereAspergillus flavus andparasiticus Agar (AFPA, 14), the rose bengal agar described by Bell and Crawford (BCRB; 3), a modified rose bengal agar (M-RB), and Czapek's-Dox Agar supplemented with the antibiotics in BC-RB (CZ-RB). M-RB was the most useful for studying the population biology of this group because it permitted both identification of the greatest number ofA. flavus group strains and growth of the fewest competing fungi. M-RB supported an average of 12% moreA. flavus group colonies than the original rose bengal medium while reducing the number of mucorales colonies and the number of total fungi by 99% and 70%, respectively. M-RB was successfully employed to isolate all three aflatoxin producing species,A. flavus, A. parasiticus andA. nomius, and both the S and L strains ofA. flavus. M-RB is a defined medium without complex nitrogen and carbon sources (e.g. peptone and yeast extract) present in BC-RB. M-RB should be useful for studies on the population biology of theA. flavus group.Abbreviations M-RB Modified Rose Bengal Agar - CZ-RB Czapeks Rose Bengal Agar - BC-RB Bell and Crawford's Rose Bengal Agar - AFPA Aspergillus flavus andparasiticus agar     

The effect of freezing and the influence of isolation medium on the recovery of pathogenic fungi from sputum

The primary objective of this study was to determine whether freezing sputa in dry ice had any effect on the recovery of pathogenic fungi. Sputa seeded with each of five fungi (Histoplasma capsulatum, Blastomyces dermatitidis, Cryptococcus neoformans, Coccidioides immitis, and Aspergillus fumigatus) were frozen and stored for 24, 48, and 72 hours on dry ice. H. capsulatum was killed, and only a few colonies of B. dermatitidis and C. neoformans were isolated from these sputa. However, A. fumigatus and C. immitis withstood the effects of freezing. A second objective was to compare the recovery of all five fungi from seeded sputa stored at room temperature for 24, 48, and 72 hours, on yeast extract-phosphate agar with NH₄OH and on Sabhi agar. The yeast extract-phosphate agar with NH₄OH was superior to Sabhi agar, for the isolation of all fungi studied, except A. fumigatus.     

Indoor airbornePenicillium species in Taiwan

Penicillium species, well-known indoor airborne fungi, are considered to be important causative agents of extrinsic bronchial asthma. For selecting the proper species for clinical investigations, it is important to know which are found indoors. In the present study, 176 plates of modified Mehrlich's Medium were exposed for 20 min in living rooms and bedrooms of 88 homes in Taipei, Taiwan. A total of 5897 colonies of fungi were isolated, varying from one to 329 per house. Of the total, 910 colonies (15.4%) were identified asPenicillium spp. The three most frequently encountered species wereP. citrinum, P. crustosum andP. implicatum. Penicillium citrinum accounted for 40.5% (369 from 28 houses);P. implicatum, for 5.2% (47 from three houses); andP. crustosum, for 2.1% (19 from seven houses).Penicillium citrinum was judged to be the most prevalent airborne fungus in Taiwan. The allergenic significance of this mold is under further investigation.     

Evaluation of the Effects of Biological Preparations on Phytopathogenic Fungi Didymella applanata and Botrytis cinerea

Fungicidal and fungistatic effects of biological preparations involving bacteria of the genera Pseudomonas and Bacillusand fungi of the genusChaetomium on phytopathogenic fungi Didymella applanata and Botrytis cinereawere evaluated. All the biological preparations under study inhibited the growth of colonies of the fungi; however, the degree of the inhibition depended on the nature of each particular microorganism and the concentration of each particular preparation. A preparation containing Bacillus subtilis at a concentration of 0.2% effected maximum suppression ofB. cinerea (the diameter of the colonies decreased sevenfold). Preparations containing bacteria of the genus Pseudomonas and fungi of the genus Chaetomium were most efficient in suppressing D. applanata.Preparations containingB. subtilis and Chaetomium spp. showed promise as agents against simultaneous development of spur blight and Botrytis blight.     

Preliminary surveys of outdoor and indoor aeromycobiota in Uganda

This investigation was conducted during the period of March through June 1998 to trap, enumerate and identify the different airborne fungi in a variety of microhabitats of outdoor and indoor environments in different localities of Uganda. The settle plate method was used and Czapek-Dox agar was the isolation medium. A total of 47 genera and 61 species in addition to some other unidentified airborne fungi were trapped from all exposures at outdoor (39 genera and 52 species) and indoor (35 and 49) environments. The total fungal catches of outdoor airspora obtained from all exposures (and even in most individual exposures) were more than twice (5222 colonies) of that of the indoor ones (4361) when the exposure periods are taken into consideration. It is worth mentioning that the most highly polluted sites were either parks, forests or river banks for outdoor exposures, or teaching laboratory, library, laterines or bathrooms for indoor exposures. The most prevalent fungi from both outdoor and indoor microhabitats being species of Mycosphaerella, Yeasts, Penicillium, Fusarium, Aspergillus,Cochliobolus and Alternaria. However, several others were trapped frequently from either outdoor or indoor environments. On the other hand, several others were trapped only, but not frequently (in low or rare instances) from either outdoor or indoor microhabitats. The implications of these airborne spores are also discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Reduction of selenium oxyanions by unicellular,polymorphic and filamentous fungi: Cellular location of reduced selenium and implications for tolerance

Summary The ability of several filamentous, polymorphic and unicellular fungi to reduce selenite to elemental selenium on solid medium was examined.Fusarium sp. andTrichoderma reeii were the only filamentous fungi, of those tested, which reduced selenite to elemental selenium on Czapek-Dox agar resulting in a red colouration of colonies. Other organisms (Aspergillus niger, Coriolus versicolor, Mucor SK, andRhizopus arrhizus) were able to reduce selenite only on malt extract agar. Several fungi were able to grow in the presence of sodium selenite but were apparently unable to reduce selenite to elemental selenium, indicating that other mechanisms of selenite tolerance were employed, such as reduced uptake and/or biomethylation to less toxic, volatile derivatives. Sodium selenate was more toxic toFusarium sp. than selenite, and the toxicity of both oxyanions was increased in sulphur-free medium, with this effect being more marked for selenate. Scanning electron microscopy ofAspergillus funiculosus andFusarium sp. incubated with sodium selenite showed the presence of needle-like crystals of elemental selenium on the surfaces of hyphae and conidia, while transmission electron microscopy ofA. funiculosus revealed the deposition of electron-dense granules in vacuoles of selenite-treated fungi. Several yeasts were able to grow on MYGP agar containing sodium selenate or sodium selenite at millimolar concentrations. Sone, notablyRhodotorula rubra andCandida lipolytica, and the polymorphic fungusAureobasidium pullulans were also effective at reducing selenite to elemental selenium, resulting in red-coloured colonies.Schizosaccharomyces pombe was able to grow at selenite concentrations up to 5 mmol L^–1 without any evidence of reduction, again indicating the operation of other tolerance mechanisms.     

Isolation of Beauveria bassiana and Metarhizium anisopliae var. anisopliae from Boophilus microplus tick (Canestrini, 1887), in Rio de Janeiro State,Brazil

The objective of this work were to isolate and identify strains of entomopathogenic fungi from ingurgitated female Boophilus microplus ticks, collected from the soil in the municipality of Paracambi, Rio de Janeiro State, Brazil. The ingurgitated females were inoculated in the selective medium oat dodine agar (oda), where 49 colonies of Beauveria bassiana (71%) and 20 of Metarhizium anisopliae var. anisopliae (29%) were isolated. These isolated strains characterize for the first time in Brazil the natural occurrence of these species of fungi in this tick, and will be used to conduct bioassays to evaluate the pathogenicity and virulence of these strains for ticks of the genus Boophilus microplus. This revised version was published online in June 2006 with corrections to the Cover Date.     

Fertile fungal spores collected on different faced surfaces in the atmosphere of Giza,Egypt

A quantitative and qualitative survey was carried out for airborne fungus spores coming into contact with horizontally and vertically gravitation sampling oriented surfaces in the atmosphere of Giza city. Czapek Dox agar, malt extract agar, potato dextrose agar and Sabouraud dextrose agar Petri dishes were exposed monthly to the five oriented surfaces of a polystyrene cube, throughout a one-year period. Significant differences (P < 0.01) were observed between the total counts of caught airborne fungi contacting with the horizontal compared to other vertically oriented surfaces. Conversely, there were no significant differences observed between the total catch of airborne fungi using the various sampling media. The results revealed that vertical sampling provides valuable information that may be lost from horizontal sampling alone. A total of 5,053 colonies belonging to 40 fungal organisms were identified. Alternaria (24.26%), Aspergillus (19.2%), Cladosporium (14.5%) and Penicillium (11.43%) were the most predominant fungal genera. Collected fungi were grouped into high, medium, low and rare components depending upon their frequency in the studied atmosphere. Aspergillus niger, Aspergillus parasiticus, Alternaria, Cladosporium and Penicillium were regularly found on all oriented surfaces. However, Arthrobotrys, Biospora, Chaetomium, Pleospora, Trichothecium and Verticillium were rarely found in the air. Positive and/or negative correlations were observed between the total fungal counts and the predominant fungal types with meteorological parameters during sampling days.     

Regular dorsal dimples and damaged mites of <Emphasis Type="Italic">Varroa destructor</Emphasis> in some Iranian honey bees (<Emphasis Type="Italic">Apis mellifera</Emphasis>)

The frequency of damaged Varroa destructor Anderson and Trueman (Mesostigmata: Varroidae) found on the bottom board of hives of the honey bee, Apis mellifera L. (Hymenoptera: Apidae) has been used as an indicator of the degree of tolerance or resistance of honey bee colonies against mites. However, it is not clear that this measure is adequate. These injuries should be separated from regular dorsal dimples that have a developmental origin. To investigate damage to Varroa mites and regular dorsal dimples, 32 honey bee (A. mellifera) colonies were selected from four Iranian provinces: Isfahan, Markazi, Qazvin, and Tehran. These colonies were part of the National Honey bee Breeding Program that resulted in province-specific races. In April, Varroa mites were collected from heavily infested colonies and used to infest the 32 experimental colonies. In August, 20 of these colonies were selected (five colonies from each province). Adult bees from these colonies were placed in cages and after introducing mites, damaged mites were collected from each cage every day. The average percentage of injured mites ranged from 0.6 to 3.0% in four provinces. The results did not show any statistical differences between the colonies within provinces for injuries to mites, but there were some differences among province-specific lines. Two kinds of injuries to the mites were observed: injuries to legs and pedipalps, and injuries to other parts of the body. There were also some regular dorsal dimples on dorsal idiosoma of the mites that were placed in categories separate from mites damaged by bees. This type of classification helps identifying damage to mites and comparing them with developmental origin symptoms, and may provide criteria for selecting bees tolerant or resistant to this mite.     

Endophytic Bacterial Communities in Ginseng and their Antifungal Activity Against Pathogens

Plant roots are associated with diverse communities of endophytic bacteria which do not exert adverse effects. The diversity of bacterial endophytes associated with ginseng roots cultivated in three different areas in Korea was investigated. Sixty-three colonies were isolated from the interior of ginseng roots. Phylogenetic analysis based on 16S rRNA gene sequences showed that the isolates belonged to three major phylogenetic groups: the high G+C Gram-positive bacteria (HGCGPB), low G+C Gram-positive bacteria (LGCGPB), and the Proteobacteria. The dominant species at the three different ginseng growing areas were: HGCGPB at Ganghwa (55.0%), LGCGPB at Geumsan (45.5%), and Proteobacteria at Jinan (61.9%). Most cellulase-, xylanase-, and pectinase-producing colonies among the isolates belong to the LGCGPB group, except for Pectobacterium carotovora which belonged to the Proteobacteria. The 13 isolates belonging to LGCGPB and Proteobacteria were assessed for their antifungal activity against phytopathogenic fungi such as Rhizoctonia solani. Among them, Paenibacillus polymyxa GS01, Bacillus sp. GS07, and Pseudomonas poae JA01 show potential activity as biocontrol agents against phytopathogenic fungi. Finally, most of the low G+C Gram-positive bacteria with antifungal activity against phytopathogenic microorganisms showed cellulolytic enzyme activity while some Proteobacteria with the antifungal activity and the high G+C Gram-positive bacteria did not show any cellulolytic activity.     

Composition and variability of airborne fungi in an enclosed rabbit house in China

Sampling was conducted from June 2007 to May 2008 in an enclosed rabbit house to investigate composition and variability of airborne fungi. Samples were collected using an Andersen-6 sampler, with Sabouraud culture medium as sampling medium. The results showed that monthly mean concentration was 2.79–5.46 × 10³ colony forming unit/m³ air (CFU/m³ air), with the maximum level in October, and the minimum level in January. Within a day, the maximum level occurred at 09:00, followed by 17:00 and then 13:00. A total of 6,523 fungal colonies, belonging to 17 genera and 36 species, were obtained. The predominant genera included Cladosporium, Penicillium, Aspergillus and Altemaria, comprising 71.45% of the colony count. The obtained fungi of the year were mainly centralized in the stage D of the sampler (2.0–3.0 μm), accounting for 37.8% of the colonies. The minimum value occurred at stage F (<0.65 μm), accounting for 1.10% of the colonies.     

Fungi in healthy and diseased sea fans (<Emphasis Type="Italic">Gorgonia ventalina)</Emphasis>: is <Emphasis Type="Italic">Aspergillus sydowii</Emphasis> always the pathogen?

Caribbean corals, including sea fans (Gorgonia spp.), are being affected by severe and apparently new diseases. In the case of sea fans, the pathogen is reported to be the fungus Aspergillus sydowii, and the disease is named aspergillosis. In order to understand coral diseases and pathogens, knowledge of the microbes associated with healthy corals is also necessary. In this study the fungal community of healthy Gorgonia ventalina colonies was contrasted with that of diseased colonies. In addition, the fungal community of healthy and diseased tissue within colonies with aspergillosis was contrasted. Fungi were isolated from healthy and diseased fans from 15 reefs around Puerto Rico, and identified by sequencing the nuclear ribosomal ITS region and by morphology. Thirty fungal species belonging to 15 genera were isolated from 203 G. ventalina colonies. Penicillum and Aspergillus were the most common genera isolated from both healthy and diseased fans. However, the fungal community of healthy fans was distinct and more diverse than that of diseased ones. Within diseased fans, fungal communities from diseased tissues were distinct and more diverse than from healthy tissue. The reduction of fungi in diseased colonies may occur prior to infection due to environmental changes affecting the host, or after infection due to increase in dominance of the pathogen, or because of host responses to infection. Data also indicate that the fungal community of an entire sea fan colony is affected even when only a small portion of the colony suffers from aspergillosis. An unexpected result was that A. sydowii was found in healthy sea fans but never in diseased ones. This result suggests that A. sydowii is not the pathogen causing aspergillosis in the studied colonies, and suggests several fungi common to healthy and diseased colonies as opportunistic pathogens. Given that it is not clear that Aspergillus is the sole pathogen, calling this disease aspergillosis is an oversimplification at best. Communicated by Biology Editor Dr Michael Lesser     

The mycoflora of groundnut rhizosphere,soil and geocarposphere on light,medium and heavy soils and its relations to aspergillus flavus

Summary A comparative study was carried out in groundnut fields in Israel of the mycoflora of rhizosphere (R) and soil adhering to geocarps (G) and of the soil (S).Of a total of 157 fungal species isolated and listed, 133 occurred in R, 96 in S, and 86 in G. Ten species considered rare or worthy of special note are briefly discussed.Species ofAspergillus were most numerous in heavy soil, species ofPenicillium in light soil, and species ofFusarium in medium soil.Penicillium funiculosum andP. rubrum were the species occurring with greatest frequency, and these were equally common in R, S, and G of light and medium soil. Soil inoculation withA. flavus depressed the number of species in the mycoflora of R and S.Quantitative comparison of R, S, and G on the three soils showed that numbers for R exceeded those for S and G, forP. funiculosum andP. rubrum on all soils, and for several Aspergilli on heavy soil. Among fungi more prevalent in S than in R and G wasA. flavus on medium and heavy soil. No special affinity of any one of the 3 major groups (Aspergillus, Penicillium, Fusarium) for the mycoflorae of R or S or G has been found. The relative number of colonies of all fungi in R and S (R/S ratio) was in almost all cases in excess of 1 on all soil types.This research is supported by grant number FG-161 of the United States Department of Agriculture, to whom the author is indebted.     

New records of microcyclic conidiogenesis in some powdery mildew fungi

Microcyclic conidiogenesis (MC) was recently described in several species of powdery mildew fungi. This process, defined as the production of conidia on a fungal spore without any, or only a minimal, involvement of hyphal growth, was observed on powdery mildew conidia that have already germinated on host plant surfaces and have been attached to the epidermal cells. Most probably, MC contributes to a quick propagation of young powdery mildew colonies because new conidia are sometimes produced in a shorter time on microcyclic conidiophores than on the hyphae of the young mycelium. This article reports MC in Erysiphe necator ex grapevine, Podosphaera leucotricha ex apple, Golovinomyces orontii ex tobacco, and Neoerysiphe galeopsidis ex Lamium purpureum based on light and low-temperature scanning electron microscopic studies.     

Isolation and toxigenicity of Aspergillus fumigatus from moldy silage

Thirty-nine silage samples were collected from various siloson Terceira Island in the Azores. Samples were examined for the presence of total fungi, and isolates of Aspergillus fumigatus were analyzed for their ability to produce fumitremorgens B and C, fumigaclavines B and C, and gliotoxin. Thirty-four silage samples (87%) were contaminated with fungi, and A. fumigatus was isolated from 27 samples (69%). Samples that were taken from the surface of silos had significantly higher populations of both total fungi and A. fumigatus than did samples taken from the middle of silos. Analysis of 27 A. fumigatus isolates (one representing each positive sample) showed that 59.3% produced fumitremorgen B; 33.3% produced fumitremorgen C; 29.6% produced fumigaclavine B; 7.4% produced fumigaclavine C; and 11.1% produced gliotoxin. Fifty-two percent of the isolates produced multiple toxins, and 25.9% did not produce any of these toxins. Gliotoxin and fumigaclavine C were always produced in combination with other toxins. Because of the demonstrated potential of these A. fumigatus isolates to producemycotoxins, it is important to properly construct and manage silos to prevent their contamination with A. fumigatus.This revised version was published online in October 2005 with corrections to the Cover Date.     

Acquisition of symbiotic dinoflagellates (Symbiodinium) by juveniles of the coral Acropora longicyathus

Scleractinian corals may acquire Symbiodinium from their parents (vertically) or from the environment (horizontally). In the present study, adult colonies of the coral Acropora longicyathus from One Tree Island (OTI) on the southern Great Barrier Reef (Australia) acquired two distinct varieties of symbiotic dinoflagellates (Symbiodinium) from the environment. Adult colonies had either Symbiodinium from clade C (86.7%) or clade A (5.3%), or a mixture of both clades A and C (8.0% of all colonies). In contrast, all 10-day-old juveniles were associated with Symbiodinium from clade A, while 83-day-old colonies contained clades A, C and D even though they were growing at the same location. Symbiodinium from clade A were dominant in both 10- and 83-day-old juveniles (99 and 97% of all recruits, respectively), while clade D was also found in 31% of 83-day-old juveniles. Experimental manipulation also revealed that parental association (with clade A or C), or the location within the OTI reef, did not influence which clade of symbiont was acquired by juvenile corals. The differences between the genetic identity of populations of Symbiodinium resident in juveniles and adult A. longicyathus suggest that ontogenetic changes in the symbiosis may occur during the development of scleractinian corals. Whether or not these changes are due to host selective processes or differences in the physical environment associated with juvenile versus adult colonies remains to be determined.