Characterization of fungi from hypersaline environments of solar salterns using morphological and molecular techniques

The Cabo Rojo Solar Salterns located on the southwest coast of Puerto Rico are composed of two main ecosystems (i.e., salt ponds and microbial mats). Even though these locations are characterized by high solar radiation (mean light intensity of 39 mol photons m⁻² d⁻¹) they harbour a diverse microscopic life. We used morphological and molecular techniques to identify a series of halotolerant fungi. A total of 183 isolates and 36 species were cultured in this study. From the water from the salt ponds, 86 isolates of 26 species were cultured. The halotolerant fungi isolated from water were: Cladosporium cladosporioides, nine Aspergillus sp., five Penicillium sp. and the black yeast Hortaea werneckii. A distinctive isolate with a blue mycelium was cultured from the salt ponds, representing a new species of Periconia based on morphology and rDNA analysis. Forty-four isolates from eight species were cultured from the sediments around the salt ponds. Most of the sediment isolates formed only sterile mycelium, while several were Chaetomium globosum. A total of 53 isolates from 16 species were isolated from the three layers of the microbial mats, of which Aspergillus niger was the most frequent isolate. Phospholipid fatty acid profiles generated from the different layers of the microbial mats indicated that the uppermost layers of the mats contained fungal biomarker, 18:2w6. This fatty acid decreased with depth, the highest concentration was observed in the green upper layer and it disappeared in the black bottom anoxic layer. This correlates with the isolation of fungi using the serial dilution technique. This is the first study that documents the presence of fungi in microbial mats.     

Dark-pigmented biodeteriogenic fungi in etruscan hypogeal tombs: New data on their culture-dependent diversity,favouring conditions,and resistance to biocidal treatments

Subterranean Cultural Heritage sites are frequently subject to biological colonization due to the high levels of humidity, even in conditions of low irradiance and oligotrophy. Here microorganisms form complex communities that may be dangerous through mineral precipitation, through the softening of materials or causing frequent surface discolorations. A reduction of contamination’s sources along with the control of microclimatic conditions and biocide treatments (overall performed with benzalkonium chloride) are necessary to reduce microbial growths. Dark discolorations have been recorded in the painted Etruscan tombs of Tarquinia, two of which have been analyzed to collect taxonomical, physiological, and ecological information. Eighteen dark-pigmented fungi were isolated among a wider culturable fraction: nine from blackening areas and nine from door sealings, a possible route of contamination. Isolates belonged to three major groups: Chaetothyriales, Capnodiales (Family Cladosporiaceae), and Acremonium-like fungi. Exophiala angulospora and Cyphellophora olivacea, a novelty for hypogea, were identified, while others need further investigations as possible new taxa. The metabolic skills of the detected species showed their potential dangerousness for the materials. Their tolerance to benzalkonium chloride-based products suggested a certain favouring effect through the decreasing competitiveness of less resistant species. The type of covering of the dromos may influence the risk of outer contamination. Fungal occurrence can be favoured by root penetration.     

Selective isolation of agents of chromoblastomycosis from insect-associated environmental sources

Chromoblastomycosis is a neglected disease characterized by cutaneous, subcutaneous or disseminated lesions. It is considered an occupational infectious disease that affects mostly rural workers exposed to contaminated soil and vegetal matter. Lesions mostly arise after a traumatic inoculation of herpotrichiellaceous fungi from the Chaetothyriales order. However, the environmental niche of the agents of the disease remains obscure. Its association with insects has been predicted in a few studies. Therefore, the present work aimed to analyze if social insects, specifically ants, bees, and termites, provide a suitable habitat for the fungi concerned. The mineral oil flotation method was used to isolate the microorganisms. Nine isolates were recovered and phylogenetic analysis identified two strains as potential agents of chromoblastomycosis, i.e., Fonsecaea pedrosoi CMRP 3076, obtained from a termite nest (n = 1) and Rhinocladiella similis CMRP 3079 from an ant exoskeleton (n = 1). In addition, we also identified Fonsecaea brasiliensis CMRP 3445 from termites (n = 1), Exophiala xenobiotica CMRP 3077 from ant exoskeleton (n = 1), Cyphellophoraceae CMRP 3103 from bees (n = 1), Cladosporium sp. CMRP 3119 from bees (n = 1), Hawksworthiomyces sp. CMRP 3102 from termites (n = 1), and Cryptendoxyla sp. from termites (n = 2). The environmental isolate of F. pedrosoi CMRP 3076 was tested in two animal models, Tenebrio molitor and Wistar rat, for its pathogenic potential with fungal retention in T. molitor tissue. In the Wistar rat, the cells resembling muriform cells were observed 30 d after inoculation.     

Exophiala sp. LHL08 reprograms Cucumis sativus to higher growth under abiotic stresses

Endophytic fungi are potential sources of secondary metabolites; however, they are little known for phytohormones secretion and amelioration of plant growth under abiotic stresses. We isolated a novel endophyte from the roots of Cucumis sativus and identified it as a strain of Exophiala sp. by sequencing internal transcribed spacer/large subunit rDNA and phylogenetic analysis. Prior to identification, culture filtrate (CF) of Exophiala sp. has shown significant growth promotion of Waito‐C [a gibberellins (GAs)‐deficient mutant cultivar] and Dongjin‐byeo (normal GAs biosynthesis cultivar) rice seedlings. CF analysis of Exophiala sp. showed the presence of physiologically active GAs (GA₁, GA₃, GA₄ and GA₇) and inactive GAs (GA₅, GA₈, GA₉, GA₁₂ and GA₂₀). Exophiala sp. had higher GAs in its CF than wild‐type strain of Gibberella fujikuroi except GA₃. Influence of Exophiala sp. was assessed on cucumber plant's growth and endogenous abscisic acid (ABA), salicylic acid (SA) and bioactive GAs under salinity and drought stresses. Exophiala sp.‐treated plants have shown significantly higher growth and rescued the host plants from stress promulgated water deficit, osmotic and cellular damage. The altered levels of stress‐responsive ABA showed low level of stress confined to endophyte‐applied plants than control. Elevated levels of SA and bioactive GAs (GA₃ and GA₄) in endophyte‐associated plants suggest stress‐modulating response toward salinity and drought. In conclusion, symbiotic relations between Exophiala and cucumber have reprogrammed the host plant growth under abiotic stresses, thus indicating a possible threshold role of endophytic fungi in stress alleviation. This study could be extended for improving agricultural productivity under extreme environmental conditions.     

Detection and identification of opportunistic Exophiala species using the rolling circle amplification of ribosomal internal transcribed spacers

Deep infections by melanized fungi deserve special attention because of a potentially fatal, cerebral or disseminated course of disease in otherwise healthy patients. Timely diagnostics are a major problem with these infections. Rolling circle amplification (RCA) is a sensitive, specific and reproducible isothermal DNA amplification technique for rapid molecular identification of microorganisms. RCA-based diagnostics are characterized by good reproducibility, with few amplification errors compared to PCR. The method is applied here to species of Exophiala known to cause systemic infections in humans. The ITS rDNA region of five Exophiala species (E. dermatitidis, E. oligosperma, E. spinifera, E. xenobiotica, and E. jeanselmei) was sequenced and aligned in view of designing specific padlock probes to be used for the detection of single nucleotide polymorphisms (SNPs) of the Exophiala species concerned. The assay proved to successfully amplify DNA of the target fungi at the level of species; while no cross-reactivity was observed. Amplification products were visualized on 1% agarose gels to verify the specificity of probe-template binding. Amounts of reagents were minimized to avoid the generation of false positive results. The sensitivity of RCA may help to improve early diagnostics of these difficult to diagnose infections.     

Isolation and Screening of Black Fungi as Degraders of Volatile Aromatic Hydrocarbons

Black fungi reported as degraders of volatile aromatic compounds were isolated from hydrocarbon-polluted sites and indoor environments. Several of the species encountered are known opportunistic pathogens or are closely related to pathogenic species causing severe mycoses, among which are neurological infections in immunocompetent individuals. Given the scale of the problem of environmental pollution and the phylogenetic relation of aromate-degrading black fungi with pathogenic siblings, it is of great interest to select strains able to mineralize these substrates efficiently without any risk for public health. Fifty-six black strains were obtained from human-made environments rich in hydrocarbons (gasoline car tanks, washing machine soap dispensers) after enrichment with some phenolic intermediates of toluene and styrene fungal metabolism. Based on ITS sequencing identification, the majority of the obtained isolates were members of the genus Exophiala. Exophiala xenobiotica was found to be the dominant black yeast present in the car gasoline tanks. A higher biodiversity, with three Exophiala species, was found in soap dispensers of washing machines. Strains obtained were screened using a 2,6-dichlorophenol-indophenol (DCPIP) assay, optimized for black fungi, to assess their potential ability to degrade toluene. Seven out of twenty strains tested were able to use toluene as carbon source.     

The antifungal activity of fatty acids of all stages of Sarcophaga carnaria L. (Diptera: Sarcophagidae)

Fatty acids as components of cuticular lipids of insects play a significant role in antifungal in protection against fungal infection. The chemical composition of cuticular and internal extracts obtained from all developmental stages of flesh flies Sarcophaga carnaria was identified. The fatty acids were detected using gas chromatography coupled with mass spectrometry and the most abundant for all examined stages were: 18:1 > 16:0 > 16:1 > 18:0 > 18:2. Polyunsaturated fatty acids (PUFA) C20 were found in both, cuticular and internal extracts. GC–MS analysis showed higher relative content of PUFA in adults than in preimaginal stages.Fatty acids alone as well as their cuticular and internal extracts obtained from larvae, pupae male and female of S. carnaria were tested according to their potential antimicrobial activity against entomopathogenic fungi: Paecilomyces lilacinus, Paecilomyces fumosoroseus, Lecanicillium lecanii, Metarhizium anisopliae, Beauveria bassiana (Tve-N39) and B. bassiana (Dv-1/07). FA presented diverse antimicrobial activity depending on the length of the chain and the presence of unsaturated bonds. Short chain and unsaturated FA (6:0, 11:0, 13:0) have shown significantly stronger activity against fungi but they were detected in lower concentrations. PUFA inhibit fungal growth more effectively than unsaturated long chain fatty acids. Cuticular and internal extracts of all living forms of S. carnaria exhibited approximately equal activity against tested entomopathogenic fungi. We presumed that the most abundant saturated long chain FA and additionally PUFA founded in our analysis are involved in protecting the flies against fungal infection.     

海水来源的548株真菌的近海及远海分布模式及其抑菌活性分析

【目的】海洋真菌是新颖天然产物的理想来源。本研究旨在加深对可培养海洋真菌多样性的了解,以及挖掘具有应用潜力的海洋真菌。【方法】采用膜过滤法分离近海和远海海水样品中的真菌,通过菌株分离纯化、ITS基因序列测序,分析浅海带、半深海带和深海带海水样品中可培养真菌的多样性。在此基础上,采用固体平板法筛选具有抑菌活性的菌株。【结果】本研究共获得548株纯培养真菌,它们分属于2个门6个纲11个目19个科24个属44个种,其中包含海洋真菌种15个和潜在新分类单元3个。在纲水平上,座囊菌纲(Dothideomycetes)包含种的数量最多(13个),其次为粪壳菌纲(Sordariomycetes)(11个)和散囊菌纲(Eurotiomycetes)(11个)。在属水平上,青霉属(Penicillium)、枝孢属(Cladosporium)、外瓶霉属(Exophiala)和拟青霉属(Simplicillium)的分离频率较高。44个真菌种呈现出5种不同的地理分布特征,包括在浅海带、半深海带和深海带均有分布(15个种),仅分布于浅海带(5个种),不存在于浅海带(12个种),不存在于半深海带(5个种),以及...     

Fungal Parasites of the Potato Cyst Nematode Globodera rostochiensis: Isolation and Reinfection

Fungal parasitism of eggs of the potato cyst nematode Globodera rostochiensis was < 1, 3, and 17% at three sites in Sweden. The fungi isolated most frequently from infected eggs were a Septocylindrium-like fungus ( 19 %), Exophiala spp. (17 %), and Cylindrocarpon spp. (13 %). Verticillium suchtasporium was isolated from infected eggs at a low frequency (4%). In laboratory experiments V. suchlasporium infected 93% of the eggs within cysts after 10 days on dilute corn meal agar. This species showed chitinase and protease activity. Infection of eggs by the Septocylindrium-like fungus was moderate, whereas Cylindrocarpon destructans and Cladosporium cladosporoides did not infect eggs. No chitinase activity was found in these fungi, but protease activity was recorded in all. Growth of the fungi in cysts did not influence the number of physiologically disordered eggs.     

Black yeast habitat choices and species spectrum on high altitude creosote-treated railway ties

Polyextremotolerant black yeast-like fungi thrive in moderately hostile environments where they are concomitantly subjected to several types of stress, such as toxicity, scarce nutrient availability, and high or low temperature extremes. Their ability to assimilate alkylbenzenes (toxic environmental pollutants) enhances their growth in harsh conditions, including on railway ties. Samples were collected using cotton swabs, premoistened with physiological saline, from 658 oak and concrete railway ties at six train stations in Turkey at altitudes ranging between 1026 and 1427 m. The samples were inoculated on malt extract agar supplemented with chloramphenicol, and incubated at 26 °C for 4 weeks. Twenty-four samples (3.6 %), 17 from oak and 7 from concrete (5.6 % vs. 2 %; P = 0.02), tested positive for fungi. Exophiala crusticola was found to be the most common species (n = 13), followed by Exophiala phaeomuriformis (n = 7) and Exophiala heteromorpha (n = 4). These results suggest that hydrocarbons, particularly creosote-treated oak woods, support the growth of black yeasts, some of which are opportunists in humans.     

Massive Contamination of Exophiala dermatitidis and E. phaeomuriformis in Railway Stations in Subtropical Turkey

In order to reveal the source of contamination of opportunistic fungi, their natural habitat has to be understood. Black yeast-like fungi are abundant in man-made environments, particularly in those that are rich in toxic hydrocarbons such as railway ties. In this study, we investigated the presence of black fungi on creosote-treated oak railway ties and concrete sleepers stained with petroleum oil. Samples were collected at two central stations in Turkish cities, Mersin and Adana, and from Tarsus town station located between these two. The sample locations had subtropical climates. A total of 570 railway samples, including 320 from oak and 250 from concrete, were collected. Cotton swabs moistened with sterile physiological saline were applied to the ties and inoculated onto malt extract agar followed by incubation at 37 °C. Overall, we recovered 97 black yeast-like fungi (17.0 % positive). Sixty-three fungi (19.7 %) were collected from creosote-treated oak, whereas 34 isolates (13.6 %) were derived from concrete; the difference was significant (P = 0.05). Identification using rDNA internal transcribed spacer revealed Exophiala dermatitidis (57.7 %) and Exophiala phaeomuriformis (42.3 %). This study suggested that hydrocarbons enrich these opportunistic black yeasts. An eventual health risk is discussed.     

The effects of laurel (Laurus nobilis L.) on development of two mycorrhizal fungi

Hundreds of aromatic plant species are growing naturally around Mediterranean. Plant essential oils are incorporated in aromatic plant material and follow the litter fall. During litter degradation, the presence of essential oils can affect soil microorganisms. Mycorrhizal fungi have never been investigated so far under the presence of volatile oils. The aim of this study was to explore the effect of aromatic Laurus nobilis L. on development of two mycorrhizal species Glomus deserticola and Glomus intraradices. The response of fungi colonization and host growth were monitored under different concentrations of L. nobilis leaves and essential oil. The major compounds of L. nobilis essential oil were 1,8-cineole (49.6%), sabinene (7.8%), ??-pinene (6.0%), eugenole (5.6%), ??-terpinyl acetate (5.2%) and ??-pinene (5.1%). Both mycorrhizal fungi colonized successfully the host plants whose growth was positively influenced by mycorrhizal fungi. G. deserticola presented higher infection level than G. intraradices. The addition of L. nobilis leaves in the soil resulted in mycorrhiza inhibition. The level of inhibition was positively correlated with the added amount of aromatic leaves in the soil. The essential oil presented a little higher inhibition than the leaves. The presence of this aromatic plant in many different ecosystems could contribute in mycorrhiza inhibition and it is suggested, when it’s possible, reduction of laurel litter before reforestation programs.     

Fungal infection of sea turtle eggs in the sea turtle hatcheries in Peninsular Malaysia

Fungal infection in sea turtle nests has become a potential threat to sea turtle embryos. We screened the hatchery nest sand, eggshells of failed eggs, and stillbirths of green turtles and hawksbills collected from hatcheries in Malacca, Pahang, Perak and Terengganu for the presence of fungi. The DNA sequence of the ITS region of the three highest occurring isolated fungi confirmed that these species were Pseudallescheria ellipsoidea (35.4%), Scedosporium aurantiacum (27.2%), and Fusarium solani (22.0%). Morphological characteristics of these fungi were recorded. Although the total fungi abundance had no significant effect on hatching success (p > 0.05), the abundance of P. ellipsoidea significantly increased mortality in the nests (r = 0.70, p < 0.05). Future research should focus on understanding the biological aspects of this species to establish a more effective mitigation technique for the prevention of fungal infection of sea turtle eggs and hatchery employees.     

Free living amoebae could enhance Fusarium oxysporum growth

Free living amoebae and Fusarium oxysporum can be recovered in the same environment and may potentially interact. The presence of these protozoa could lead to an increased development of this filamentous fungus. To assess this potential risk, the interactions between two free living amoebae, Acanthamoeba castellanii and Hartmanella vermiformis, and F. oxysporum, which can be isolated from soil and water, were studied. After 48 hr of coincubation in tap water, culturable fungi were quantified. In addition, the interactions between the free living amoebae and the fungus were investigated using electron microscopy. We show that the presence of amoeba trophozoites increased the growth of F. oxysporum without fungal influence on amoebae viability. In the same way, incubation of the fungus with culture supernatants of the two amoebae induced fungal germination and increased fungal growth. The results of this study confirm that the presence of amoebae should be taken into consideration in the different environments where they may be in contact with Fusarium.     

The Diversity and Distribution of Fungi on Residential Surfaces

The predominant hypothesis regarding the composition of microbial assemblages in indoor environments is that fungal assemblages are structured by outdoor air with a moderate contribution by surface growth, whereas indoor bacterial assemblages represent a mixture of bacteria entered from outdoor air, shed by building inhabitants, and grown on surfaces. To test the fungal aspect of this hypothesis, we sampled fungi from three surface types likely to support growth and therefore possible contributors of fungi to indoor air: drains in kitchens and bathrooms, sills beneath condensation-prone windows, and skin of human inhabitants. Sampling was done in replicated units of a university-housing complex without reported mold problems, and sequences were analyzed using both QIIME and the new UPARSE approach to OTU-binning, to the same result. Surfaces demonstrated a mycological profile similar to that of outdoor air from the same locality, and assemblages clustered by surface type. “Weedy” genera typical of indoor air, such as Cladosporium and Cryptococcus, were abundant on sills, as were a diverse set of fungi of likely outdoor origin. Drains supported more depauperate assemblages than the other surfaces and contained thermotolerant genera such as Exophiala, Candida, and Fusarium. Most surprising was the composition detected on residents’ foreheads. In addition to harboring Malassezia, a known human commensal, skin also possessed a surprising richness of non-resident fungi, including plant pathogens such as ergot (Claviceps purperea). Overall, fungal richness across indoor surfaces was high, but based on known autecologies, most of these fungi were unlikely to be growing on surfaces. We conclude that while some endogenous fungal growth on typical household surfaces does occur, particularly on drains and skin, all residential surfaces appear – to varying degrees – to be passive collectors of airborne fungi of putative outdoor origin, a view of the origins of the indoor microbiome quite different from bacteria.     

Molecular cloning, characterization and differential expression of Cdc42 in Fonsecaea monophora

The cell divisions cycle 42 (Cdc42) gene has been characterized in the fungi, such as Candida albicans, Penicillium marneffei, and Wangiella (Exophiala) dermatitidis, which plays important roles during growth and development. The partial cDNA sequence of Cdc42 of Fonsecaea monophora (F. monophora), designated FmCdc42, was obtained using degenerate primers based on the conserved domain of the other fungi Cdc42. Then the complete cDNA sequence of FmCdc42 was obtained by 5′ and 3′ RACE. The full-length cDNA is 1,510 bp in size which had an open reading frame (ORF) of 582 bp, encoding 193 amino acid residues. The predicted molecular mass of FmCdc42 is 21.5 kDa with an estimated theoretical isoelectric point of 5.67. The deduced amino acid sequence of FmCdc42 shows 99% identity to that of Wangiella (Exophiala) dermatitidis. 5 exons and 4 introns are identified within the 1,617 bp FmCdc42 genomic DNA sequence of F. monophora. The ORF could be subcloned into the pCDNA6/myc-His B expression vector. The recombinant protein about 27.5 kD infusion protein had high expression level in Vero cells with SDS-PAGE and Western blot analysis. Quantitative real time RT-PCR revealed that FmCdc42 was the highest expression in the sclerotic bodies’ stage compared with that in the mycelia and conidia stages, which indicated that the FmCdc42 may be involved in formation of F. monophora sclerotic bodies.     

Arbuscular mycorrhizal fungal abundance in elevation belts of the hyperarid Atacama Desert

Arbuscular mycorrhizal (AM) fungi affect ecosystem processes improving plant tolerance in (hyper)arid/saline environments. However, there are no previous studies on the presence of AM fungi in the Atacama Desert (Northern Chile), the driest desert in the world. Here, we studied the root and rhizosphere in 111 samples of representative plants from three elevation gradients: (i) hyperarid desert (700–2000 m a.s.l.), (ii) pre-Puna (2000–3100 m a.s.l.) and (iii) Puna (3100–4500 m a.s.l.) elevation belts. Soil pH, electrical conductivity (EC), organic matter, cations and the AM colonization and fungal structures were determined. All plants showed colonization and fungal structures. Root colonization ranged from 3.5 to 87%, hyphae showed densities from 0.13 to 204 m g⁻¹, and spore densities between 20 and 45,500 per 100 g of soil. The highest fungal structure abundances were found in Prosopis tamarugo, Baccharis scandens, Werneria pinnatifida, Deyeuxia curvula and Festuca deserticola rhizospheres. In general, EC and cations showed strong relationship with fungal structure abundance. Here, we reported for the first time the widespread presence of AM symbiosis in all the elevation belts of the Atacama Desert, constituting a first step to understand the ecological role that AM fungi play under extreme aridity and salinity conditions.     

Black Yeast Biota in the Mangrove, in Search of the Origin of the Lethargic Crab Disease (LCD)

Knowledge of natural ecology is essential for a better understanding of pathogenicity and opportunism in black yeast-like fungi. Although etiological agents of diseases caused by these fungi are supposed to originate from the environment, their isolation from nature is difficult. This is probably due to their oligotrophic nature, low competitive ability, and, overall, insufficient data on their natural habitat. We obtained environmental samples from mangrove areas where mortalities by lethargic crab disease (LCD) are reported and areas without disease recorded. Isolation of chaetothyrialean black yeasts and relatives was performed using a highly selective protocol. Species-specific primers were used to determine if these isolates represented Exophiala cancerae or Fonsecaea brasiliensis, two proven agents of LCD, in order to test hypotheses about the origin of the disease. Isolates, identified by morphology as Fonsecaea- or Exophiala-like, were tested specific diagnostic markers for the fungi associated with LCD. Although several black fungi were isolated, the main causative agent of the LCD, E. cancerae, was not found. Molecular markers for F. brasiliensis revealed 10 positive bands for isolates from biofilms on mangrove leaves, branches, and aerial roots, of which four were confirmed by ITS sequencing. The absence of E. cancerae in environmental samples suggests that the species is dependent on the crab, as a genuine pathogen, different from F. brasiliensis, which is probably not dependent on the host species, U. cordatus. However, we did not attempt isolation from the marine water, which may represent the pathway of dispersion of the black yeast species between neighbor mangroves.     

Antimicrobial activity of invertebrate-pathogenic fungi in the genera Akanthomyces and Gibellula

Infectious disease caused by antibiotic resistant microorganisms is a global public health problem. There is a need to search for new bioactive compounds from new sources. In this study, we focused on invertebrate-pathogenic fungi infecting spiders. One hundred and sixty-five crude extracts from Akanthomyces (n = 45) and Gibellula (n = 10) were screened for their antimicrobial activity against nine human pathogens. Twenty-one extracts out of 165 (12.73%) from 16 (29.09%) isolates exhibited antimicrobial activity against at least one test strain. The most activity was against Staphylococcus aureus American Type Culture Collection (ATCC 25923) (8.48%) followed by Cryptococcus neoformans ATCC 90112 (3.03%), C. neoformans ATCC 90113 (2.42%), methicillin-resistant Staphylococcus aureus (MRSA) SK-1 (2.42%), Penicillium marneffei (2.42%), Microsporum gypseum (1.21%), Candida albicans ATCC 90028 (1.21%), Pseudomonas aeruginosa ATCC 27853 (0.61%) and Escherichia coli ATCC 25922 (0.61%), respectively. The ethyl acetate extract of mycelia from Gibellula pulchra EPF083 had the strongest broad spectrum antimicrobial activity with a minimum inhibitory concentration (MIC) value of 16 μg/ml against S. aureus ATCC 25923, MRSA SK-1, C. neoformans (ATCC 90112 and ATCC 90113) and P. marneffei and exhibited fungicidal activity against C. neoformans ATCC 90112 and P. marneffei with minimum fungicidal concentration (MFC) values of 16 and 32 μg/ml, respectively. These preliminary data show that invertebrate-pathogenic fungi could be a potential source of antimicrobial agents.