New species in Aspergillus section Usti and an overview of Aspergillus section Cavernicolarum.

Aspergillus sections Usti and Cavernicolarum are accommodated in the subgenus Nidulantes. In the present study, a polyphasic approach using morphology and multi-gene phylogeny was applied to investigate the taxonomy of these two sections. Based on the phylogenetic analysis, Aspergillus section Usti includes 25 species, which can be assigned to four series: Calidousti, Deflecti, Monodiorum and Usti. Aspergillus sigarelli is newly described in this section and this species was isolated from a cigarette from PR China and belongs to series Calidousti. It is clearly distinct from other members in this series based on ITS, BenA, CaM and RPB2 sequences. Aspergillus section Usti members like A. calidoustus and A. granulosus are important opportunistic pathogens, it is speculative that more pathogenetic species will be found by using polyphasic taxonomy approaches. Aspergillus section Cavernicolarum includes five species, the growth rates on agar media and size and ornamentation of conidia are important characters for differentiating species in section Cavernicolarum.

cyp51A Mutations, Extrolite Profiles, and Antifungal Susceptibility in Clinical and Environmental Isolates of the Aspergillus viridinutans Species Complex.

The past decade has seen an increase in aspergillosis in humans and animals due to Aspergillus viridinutans species complex members. Azole resistance is common to these infections, carrying a poor prognosis. cyp51A gene mutations are the main cause of acquired azole resistance in Aspergillus fumigatus This study aimed to determine if the azole-resistant phenotype in A. viridinutans complex members is associated with cyp51A mutations or extrolite profiles. The cyp51A gene of clinical and environmental isolates was amplified using novel primers, antifungal susceptibility was tested using the Clinical and Laboratory Standards Institute methodology, and extrolite profiling was performed using agar plug extraction. Very high azole MICs were detected in 84% of the isolates (31/37). The MICs of the newer antifungals luliconazole and olorofim (F901318) were low for all isolates. cyp51A sequences revealed 113 nonsynonymous mutations compared to the sequence of wild-type A. fumigatus M172A/V and D255G, previously associated with A. fumigatus azole resistance, were common among all isolates but were not correlated with azole MICs. Two environmental isolates with nonsusceptibility to itraconazole and high MICs of voriconazole and isavuconazole harbored G138C, previously associated with azole-resistant A. fumigatus Some novel mutations were identified only among isolates with high azole MICs. However, cyp51A homology modeling did not cause a significant protein structure change for these mutations. There was no correlation between extrolite patterns and susceptibility. For A. viridinutans complex isolates, cyp51A mutations and the extrolites that they produced were not major causes of antifungal resistance. Luliconazole and olorofim show promise for treating azole-resistant infections caused by these cryptic species.

Mould spoilage of foods and beverages: Using the right methodology.

Fungal spoilage of products manufactured by the food and beverage industry imposes significant annual global revenue losses. Mould spoilage can also be a food safety issue due to the production of mycotoxins by these moulds. To prevent mould spoilage, it is essential that the associated mycobiota be adequately isolated and accurately identified. The main fungal groups associated with spoilage are the xerophilic, heat-resistant, preservative-resistant, anaerobic and psychrophilic fungi. To assess mould spoilage, the appropriate methodology and media must be used. While classic mycological detection methods can detect a broad range of fungi using well validated protocols, they are time consuming and results can take days or even weeks. New molecular detection methods are faster but require good DNA isolation techniques, expensive equipment and may detect viable and non-viable fungi that probably will not spoil a specific product. Although there is no complete and easy method for the detection of fungi in food it is important to be aware of the limitation of the methodology. More research is needed on the development of methods of detection and identification that are both faster and highly sensitive.

New Penicillium and Talaromyces species from honey, pollen and nests of stingless bees.

Penicillium and Talaromyces species have a worldwide distribution and are isolated from various materials and hosts, including insects and their substrates. The aim of this study was to characterize the Penicillium and Talaromyces species obtained during a survey of honey, pollen and the inside of nests of Melipona scutellaris. A total of 100 isolates were obtained during the survey and 82% of those strains belonged to Penicillium and 18% to Talaromyces. Identification of these isolates was performed based on phenotypic characters and ß-tubulin and ITS sequencing. Twenty-one species were identified in Penicillium and six in Talaromyces, including seven new species. These new species were studied in detail using a polyphasic approach combining phenotypic, molecular and extrolite data. The four new Penicillium species belong to sections Sclerotiora (Penicillium fernandesiae sp. nov., Penicillium mellis sp. nov., Penicillium meliponae sp. nov.) and Gracilenta (Penicillium apimei sp. nov.) and the three new Talaromyces species to sections Helici (Talaromyces pigmentosus sp. nov.), Talaromyces (Talaromyces mycothecae sp. nov.) and Trachyspermi (Talaromyces brasiliensis sp. nov.). The invalidly described species Penicillium echinulonalgiovense sp. nov. was also isolated during the survey and this species is validated here.

Indoor airborne fungal pollution in newborn units in Turkey.

Pathogenic and/or opportunistic fungal species are major causes of nosocomial infections, especially in controlled environments where immunocompromised patients are hospitalized. Indoor fungal contamination in hospital air is associated with a wide range of adverse health effects. Regular determination of fungal spore counts in controlled hospital environments may help reduce the risk of fungal infections. Because infants have inchoate immune systems, they are given immunocompromised patient status. The aim of the present study was to evaluate culturable airborne fungi in the air of hospital newborn units in the Thrace, Marmara, Aegean, and Central Anatolia regions of Turkey. A total of 108 air samples were collected seasonally from newborn units in July 2012, October 2012, January 2013, and April 2013 by using an air sampler and dichloran 18% glycerol agar (DG18) as isolation media. We obtained 2593 fungal colonies comprising 370 fungal isolates representing 109 species of 28 genera, which were identified through multi-loci gene sequencing. Penicillium, Aspergillus, Cladosporium, Talaromyces, and Alternaria were the most abundant genera identified (35.14, 25.40, 17.57, 2.70, and 6.22% of the total, respectively).

Aureobasidium melanogenum: a native of dark biofinishes on oil treated wood.

The genus Aureobasidium, which is known as a wood staining mould, has been detected on oil treated woods in the specific stain formation called biofinish. This biofinish is used to develop a new protective, self-healing and decorative biotreatment for wood. In order to understand and control biofinish formation on oil treated wood, the occurrence of different Aureobasidium species on various wood surfaces was studied. Phenotypic variability within Aureobasidium strains presented limitations of morphological identification of Aureobasidium species. PCR amplification and Sanger sequencing of ITS and RPB2 were used to identify the culturable Aureobasidium species composition in mould stained wood surfaces with and without a biofinish. The analysed isolates showed that several Aureobasidium species were present and that Aureobasidium melanogenum was predominantly detected, regardless of the presence of a biofinish and the type of substrate. A. melanogenum was detected on wood samples exposed in the Netherlands, Cameroon, South Africa, Australia and Norway. ITS-specific PCR amplification, cloning and sequencing of DNA extracted from biofinish samples confirmed results of the culturing based method: A. melanogenum is predominant within the Aureobasidium population of biofinishes on pine sapwood treated with raw linseed oil and the outdoor placement in the Netherlands.

Discovery of a sexual cycle in Talaromyces amestolkiae.

Talaromyces amestolkiae is a common cosmopolitan species that has been cultured from indoor house dust, sputum and lungs from cystic fibrosis patients, indoor air, wheat, soil, pineapple, sculptures and manure. It was described as an asexual Talaromyces species and was reported to produce black sclerotia. In this study we report on the induction of sexual reproductive structures in T. amestolkiae. The mating type of 18 T. amestolkiae strains was determined with MAT-specific primers. Subsequently opposite mating types were inoculated on oatmeal agar and malt-extract agar and incubated 6-20 wk at 25 and 30 C in darkness. After incubation single ascospore isolations were made and evidence of recombination in the offspring was examined by amplified fragment length polymorphism and pairwise homoplasy index test, which is implemented in Splitstree4. The offspring displayed clear evidence of recombination on a genetic level as shown in the variations observed between banding patterns in the amplified fragment length polymorphism. Also a net-like and reticulated NeighborNet was observed and the pairwise homoplasy index test for recombination supported the presence of recombination (P = 0.003372). The distribution of MAT1-1 and MAT1-2 genes in the progeny showed a close to 1:1 ratio. Talaromyces amestolkiae is only the second heterothallic Talaromyces species to produce ascomata and ascospores under laboratory conditions.

Understanding fungal functional biodiversity during the mitigation of environmentally dispersed pentachlorophenol in cork oak forest soils.

Pentachlorophenol (PCP) is globally dispersed and contamination of soil with this biocide adversely affects its functional biodiversity, particularly of fungi - key colonizers. Their functional role as a community is poorly understood, although a few pathways have been already elucidated in pure cultures. This constitutes here our main challenge - elucidate how fungi influence the pollutant mitigation processes in forest soils. Circumstantial evidence exists that cork oak forests in N. W. Tunisia - economically critical managed forests are likely to be contaminated with PCP, but the scientific evidence has previously been lacking. Our data illustrate significant forest contamination through the detection of undefined active sources of PCP. By solving the taxonomic diversity and the PCP-derived metabolomes of both the cultivable fungi and the fungal community, we demonstrate here that most strains (predominantly penicillia) participate in the pollutant biotic degradation. They form an array of degradation intermediates and by-products, including several hydroquinone, resorcinol and catechol derivatives, either chlorinated or not. The degradation pathway of the fungal community includes uncharacterized derivatives, e.g. tetrachloroguaiacol isomers. Our study highlights fungi key role in the mineralization and short lifetime of PCP in forest soils and provide novel tools to monitor its degradation in other fungi dominated food webs.

Name changes in medically important fungi and their implications for clinical practice.

Recent changes in the Fungal Code of Nomenclature and developments in molecular phylogeny are about to lead to dramatic changes in the naming of medically important molds and yeasts. In this article, we present a widely supported and simple proposal to prevent unnecessary nomenclatural instability.

Genetic relatedness versus biological compatibility between Aspergillus fumigatus and related species.

Aspergillus section Fumigati contains 12 clinically relevant species. Among these Aspergillus species, A. fumigatus is the most frequent agent of invasive aspergillosis, followed by A. lentulus and A. viridinutans. Genealogical concordance and mating experiments were performed to examine the relationship between phylogenetic distance and mating success in these three heterothallic species. Analyses of 19 isolates from section Fumigati revealed the presence of three previously unrecognized species within the broadly circumscribed species A. viridinutans. A single mating type was found in the new species Aspergillus pseudofelis and Aspergillus pseudoviridinutans, but in Aspergillus parafelis, both mating types were present. Reciprocal interspecific pairings of all species in the study showed that the only successful crosses occurred with the MAT1-2 isolates of both A. parafelis and A. pseudofelis. The MAT1-2 isolate of A. parafelis was fertile when paired with the MAT1-1 isolates of A. fumigatus, A. viridinutans, A. felis, A. pseudoviridinutans, and A. wyomingensis but was not fertile with the MAT1-1 isolate of A. lentulus. The MAT1-2 isolates of A. pseudofelis were fertile when paired with the MAT1-1 isolate of A. felis but not with any of the other species. The general infertility in the interspecies crossings suggests that genetically unrelated species are also biologically incompatible, with the MAT1-2 isolates of A. parafelis and A. pseudofelis being the exception. Our findings underscore the importance of genealogical concordance analysis for species circumscription, as well as for accurate species identification, since misidentification of morphologically similar pathogens with differences in innate drug resistance may be of grave consequences for disease management.

Modern taxonomy of biotechnologically important Aspergillus and Penicillium species.

Taxonomy is a dynamic discipline and name changes of fungi with biotechnological, industrial, or medical importance are often difficult to understand for researchers in the applied field. Species belonging to the genera Aspergillus and Penicillium are commonly used or isolated, and inadequate taxonomy or uncertain nomenclature of these genera can therefore lead to tremendous confusion. Misidentification of strains used in biotechnology can be traced back to (1) recent changes in nomenclature, (2) new taxonomic insights, including description of new species, and/or (3) incorrect identifications. Changes in the recent published International Code of Nomenclature for Algae, Fungi and Plants will lead to numerous name changes of existing Aspergillus and Penicillium species and an overview of the current names of biotechnological important species is given. Furthermore, in (biotechnological) literature old and invalid names are still used, such as Aspergillus awamori, A. foetidus, A. kawachii, Talaromyces emersonii, Acremonium cellulolyticus, and Penicillium funiculosum. An overview of these and other species with their correct names is presented. Furthermore, the biotechnologically important species Talaromyces thermophilus is here combined in Thermomyces as Th. dupontii. The importance of Aspergillus, Penicillium, and related genera is also illustrated by the high number of undertaken genome sequencing projects. A number of these strains are incorrectly identified or atypical strains are selected for these projects. Recommendations for correct strain selection are given here. Phylogenetic analysis shows a close relationship between the genome-sequenced strains of Aspergillus, Penicillium, and Monascus. Talaromyces stipitatus and T. marneffei (syn. Penicillium marneffei) are closely related to Thermomyces lanuginosus and Th. dupontii (syn. Talaromyces thermophilus), and these species appear to be distantly related to Aspergillus and Penicillium. In the last part of this review, an overview of heterothallic reproduction in Aspergillus and Penicillium is given. The new insights in the taxonomy of Aspergillus, Penicillium, and related genera will help to interpret the results generated with comparative genomics studies or other studies dealing with evolution of, for example, enzymes, mating-type loci, virulence genes, and secondary metabolite biosynthetic gene clusters.

Taxonomic re-evaluation of black koji molds.

Black koji molds including its albino mutant, the white koji mold, have been widely used for making the distilled spirit shochu in Northeast Asia because they produce citric acid which prevents undesirable contamination from bacteria. Since Inui reported Aspergillus luchuensis from black koji in Okinawa in 1901, many fungal names associated with black koji molds were reported. However, some species are similar and differentiation between species is difficult. Fungal taxonomists tried to arrange a taxonomic system for black koji molds, but the results were not clear. Recently, multi-locus sequence typing has been successfully used to taxonomy of black Aspergillus. According to ß-tubulin and calmodulin gene sequences, black koji molds can be subdivided in three species, A. luchuensis, Aspergillus niger, and Aspergillus tubingensis. Aspergillus awamori, Aspergillus kawachii, Aspergillus inuii, Aspergillus nakazawai, and Aspergillus coreanus are synonyms of A. luchuensis, Aspergillus batatae, Aspergillus aureus (or Aspergillus foetidus), Aspergillus miyakoensis, and Aspergillus usamii (including A. usamii mut. shirousamii) are synonyms of A. niger and Aspergillus saitoi and A. saitoi var. kagoshimaensis are synonyms of A. tubingensis. A. luchuensis mut. kawachii was suggested particular names for A. kawachii because of their industrial importance. The history and modern taxonomy of black koji molds is further discussed.

Diversity of Penicillium section Citrina within the fynbos biome of South Africa, including a new species from a Protea repens infructescence.

During a survey of the fynbos biome in the Western Cape of South Africa, 61 Penicillium species were isolated and nine belong to Penicillium section Citrina. Based on morphology and multigene phylogenies, section Citrina species were identified as P. cairnsense, P. citrinum, P. pancosmium, P. pasqualense, P. sanguifluum, P. sizovae, P. sumatrense and P. ubiquetum. One of the species displayed unique phenotypic characters and DNA sequences and is described here as P. sucrivorum. Multigene phylogenies consistently resolved the new species in a clade with P. aurantiacobrunneum, P. cairnsense, P. miczynksii, P. neomiczynskii and P. quebecense. However, ITS, ß-tubulin and calmodulin gene sequences are unique for P. sucrivorum and growth rates on various media, the ability to grow at 30 C, a positive Ehrlich reaction and the absence of sclerotia on all media examined, distinguish P. sucrivorum from all of its close relatives.

Complex microbiota of a Chinese "Fen" liquor fermentation starter (Fen-Daqu), revealed by culture-dependent and culture-independent methods.

Daqu is a traditional fermentation starter that is used for Chinese liquor production. Although partly mechanized, its manufacturing process has remained traditional. We investigated the microbial diversity of Fen-Daqu, a starter for light-flavour liquor, using combined culture-dependent and culture-independent approaches (PCR-DGGE). A total of 190 microbial strains, comprising 109 bacteria and 81 yeasts and moulds, were isolated and identified on the basis of the sequences of their 16S rDNA (bacteria) and 26S rDNA and ITS regions (fungi). DGGE of DNA extracted from Daqu was used to complement the culture-dependent method in order to include non-culturable microbes. Both approaches revealed that Bacillus licheniformis was an abundant bacterial species, and Saccharomycopsis fibuligera, Wickerhamomyces anomalus, and Pichia kudriavzevii were the most common yeasts encountered in Fen-Daqu. Six genera of moulds (Absidia, Aspergillus, Mucor, Rhizopus, Rhizomucor and Penicillium) were found. The potential function of these microorganisms in starters for alcoholic fermentation is discussed. In general the culture-based findings overlapped with those obtained by DGGE by a large extent. However, Weissella cibaria, Weissella confusa, Staphylococcus saprophyticus, Enterobacter aerogenes, Lactobacillus sanfranciscensis, Lactobacillus lactis, and Bacillus megaterium were only revealed by DGGE.

Penicillium araracuarense sp. nov., Penicillium elleniae sp. nov., Penicillium penarojense sp. nov., Penicillium vanderhammenii sp. nov. and Penicillium wotroi sp. nov., isolated from leaf litter.

Several species of the genus Penicillium were isolated during a survey of the mycobiota of leaf litter and soil in Colombian Amazon forest. Five species, Penicillium penarojense sp. nov. (type strain CBS 113178(T) = IBT 23262(T)), Penicillium wotroi sp. nov. (type strain CBS 118171(T) = IBT 23253(T)), Penicillium araracuarense sp. nov. (type strain CBS 113149(T) = IBT 23247(T)), Penicillium elleniae sp. nov. (type strain CBS 118135(T) = IBT 23229(T)) and Penicillium vanderhammenii sp. nov. (type strain CBS 126216(T) = IBT 23203(T)) are described here as novel species. Their taxonomic novelty was determined using a polyphasic approach, combining phenotypic, molecular (ITS and partial ß-tubulin sequences) and extrolite data. Phylogenetic analyses showed that each novel species formed a unique clade for both loci analysed and that they were most closely related to Penicillium simplicissimum, Penicillium janthinellum, Penicillium daleae and Penicillium brasilianum. An overview of the phylogeny of this taxonomically difficult group is presented, and 33 species are accepted. Each of the five novel species had a unique extrolite profile of known and uncharacterized metabolites and various compounds, such as penicillic acid, andrastin A, pulvilloric acid, paxillin, paspaline and janthitrem, were commonly produced by these phylogenetically related species. The novel species had a high growth rate on agar media, but could be distinguished from each other by several macro- and microscopical characteristics.

Alternaria hungarica sp. nov., a minor foliar pathogen of wheat in Hungary.

During routine wheat disease surveys in Hungary in 2007 Alternaria was isolated from leaf samples collected in Debrecen. Macro- and micro-morphological examinations and ITS sequence analyses indicated that the isolates represented a new Alternaria species, which we described as A. hungarica. The usually solitary conidia of A. hungarica resemble those of A. mouchaccae and A. molesta. However growth and sporulation pattern are more like those of A. geniostomatis and A. soliaridae. Phylogenetic analysis of ITS sequences indicated that this new species can be distinguished from all other examined Alternaria and Embellisia species. Pathogenicity tests indicated that A. hungarica can be considered a minor pathogen of wheat.

Identification of Paecilomyces variotii in clinical samples and settings.

Paecilomyces variotii is a commonly occurring species in air and food, but it is also associated with many types of human infections and is among the emerging causative agents of opportunistic mycoses in immunocompromised hosts. Paecilomyces can cause hyalohyphomycosis, and two species, Paecilomyces lilacinus and P. variotii, are the most frequently encountered organisms. In the present study, a set of 34 clinical isolates morphologically identified as P. variotii or P. lilacinus were formally identified by sequencing intergenic transcribed spacer regions 1 and 2 (including 5.8S rDNA) and a part of the beta-tubulin gene. Three isolates were identified as P. lilacinus, and five of the presumptive P. variotii isolates did not belong to the genus Paecilomyces but were identified as Talaromyces eburneus (anamorph, Geosmithia argillacea) or Hamigera avellanea (anamorph, Merimbla ingelheimense). Applying the most recent taxonomy, we found that the clinical P. variotii isolates could be identified as P. variotii sensu stricto (14 strains), P. formosus (11 strains), and P. dactylethromorphus (1 strain). These data indicate that P. formosus occurs in clinical samples as commonly as P. variotii. Susceptibility tests showed that the antifungal susceptibility profiles of P. variotii, P. formosus, and P. dactylethromorphus are similar and that all strains tested were susceptible to amphotericin B in vitro. P. lilanicus, T. eburneus, and H. avellanea had different susceptibility profiles; and flucytosine and voriconazole were the least active of the antifungal drugs tested against these species. Our results indicate that correct species identification is important to help guide appropriate antifungal therapy.

Aspergillus alabamensis, a new clinically relevant species in the section Terrei.

Phylogenetic analyses of sequences generated from portions of three genes coding for the proteins enolase (enoA), beta-tubulin (benA), and calmodulin (calM) of a large number of isolates within the section Terrei, genus Aspergillus, revealed the presence of a new cryptic species within this section, Aspergillus alabamensis. Most members of this new cryptic species were recovered as colonizing isolates from immunocompetent patient populations, had decreased in vitro susceptibilities to the antifungal drug amphotericin B, and were morphologically similar to but genetically distinct from Aspergillus terreus isolates.

New section and species in Talaromyces.

Talaromyces is a monophyletic genus containing seven sections. The number of species in Talaromyces grows rapidly due to reliable and complete sequence data contributed from all over the world. In this study agricultural soil samples from Fujiang, Guangdong, Jiangxi, Shandong, Tibet and Zhejiang provinces of China were collected and analyzed for fungal diversity. Based on a polyphasic approach including phylogenetic analysis of partial ITS, BenA, CaM and RPB2 gene sequences, macro- and micro-morphological analyses, six of them could not be assigned to any described species, and one cannot be assigned to any known sections. Morphological characters as well as their phylogenetic relationship with other Talaromyces species are presented for these putative new species. Penicillium resedanum is combined in Talaromyces section Subinflati as T. resedanus.

Mycotic keratitis due to Aspergillus nomius.

We report the first known case of fungal keratitis caused by Aspergillus nomius. Ocular injury was known as a predisposing factor. The patient was treated with natamycin and econazole eye drops, itraconazole eye ointment, and oral ketoconazole. A therapeutic penetrating keratoplasty was performed 16 days after presentation. A sequence-based approach was used to assign the isolate to a species.