Publications by authors named "Katja Sterflinger"

59 Publications

Effects of Simulated Microgravity on the Proteome and Secretome of the Polyextremotolerant Black Fungus .

Front Genet 2021 18;12:638708. Epub 2021 Mar 18.

Biotechnology and Planetary Protection Group, Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA, United States.

Black fungi are a group of melanotic microfungi characterized by remarkable polyextremotolerance. Due to a broad ecological plasticity and adaptations at the cellular level, it is predicted that they may survive in a variety of extreme environments, including harsh niches on Earth and Mars, and in outer space. However, the molecular mechanisms aiding survival, especially in space, are yet to be fully elucidated. Based on these premises, the rock-inhabiting black fungus (Wt) and its non-melanized mutant (Mut) were exposed to simulated microgravity-one of the prevalent features characterizing space conditions-by growing the cultures in high-aspect-ratio vessels (HARVs). Qualitative and quantitative proteomic analyses were performed on the mycelia and supernatant of culture medium (secretome) to assess alterations in cell physiology in response to low-shear simulated microgravity (LSSMG) and to ultimately evaluate the role of cell-wall melanization in stress survival. Differential expression was observed for proteins involved in carbohydrate and lipid metabolic processes, transport, and ribosome biogenesis and translation via ribosomal translational machinery. However, no evidence of significant activation of stress components or starvation response was detected, except for the scytalone dehydratase, enzyme involved in the synthesis of dihydroxynaphthalene (DNH) melanin, which was found to be upregulated in the secretome of the wild type and downregulated in the mutant. Differences in protein modulation were observed between Wt and Mut, with several proteins being downregulated under LSSMG in the Mut when compared to the Wt. Lastly, no major morphological alterations were observed following exposure to LSSMG. Similarly, the strains' survivability was not negatively affected. This study is the first to characterize the response to simulated microgravity in black fungi, which might have implications on future astrobiological missions.
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http://dx.doi.org/10.3389/fgene.2021.638708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8012687PMC
March 2021

Natural sciences at the service of art and cultural heritage: an interdisciplinary area in development and important challenges.

Microb Biotechnol 2021 05 10;14(3):806-809. Epub 2021 Feb 10.

Institute of Natural Sciences and Technology in the Arts (INTK), Academy of Fine Arts Vienna, Vienna, Austria.

Our cultural heritage is a common asset that tells the story of our shared past, is part of our origin and identity and has wide social relevance. Our works of art and our heritage must be enjoyed, appreciated and preserved for future generations. To this end, a wide and varied group of professionals, including conservators, restorers, curators, bibliographers, historians, archivists, but also scientists, such as biologists, chemists, physicists and bioinformaticians, work side by side to preserve our cultural heritage. Working together in this wide range of disciplines included in the so-called 'heritage sciences' is the only plausible way to contribute to the sustainable preservation of our heritage. The great progress made in recent years in conservation and restoration work, but also in the natural sciences considered within heritage science, has provided powerful tools and strategies for analytical and experimental research into historical and cultural objects that open up new frontiers for their diagnosis, monitoring and protection. Here we highlight some of the advances and challenges faced by the natural sciences at the service of art.
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http://dx.doi.org/10.1111/1751-7915.13766DOI Listing
May 2021

Shed Light in the DaRk LineagES of the Fungal Tree of Life-STRES.

Life (Basel) 2020 Dec 19;10(12). Epub 2020 Dec 19.

Institute of Natural Sciences and Technology in the Arts, Academy of Fine Arts Vienna, Vienna 22180, Austria.

The polyphyletic group of black fungi within the Ascomycota (Arthoniomycetes, Dothideomycetes, and Eurotiomycetes) is ubiquitous in natural and anthropogenic habitats. Partly because of their dark, melanin-based pigmentation, black fungi are resistant to stresses including UV- and ionizing-radiation, heat and desiccation, toxic metals, and organic pollutants. Consequently, they are amongst the most stunning extremophiles and poly-extreme-tolerant organisms on Earth. Even though ca. 60 black fungal genomes have been sequenced to date, [mostly in the family Herpotrichiellaceae (Eurotiomycetes)], the class Dothideomycetes that hosts the largest majority of extremophiles has only been sparsely sampled. By sequencing up to 92 species that will become reference genomes, the "Shed light in The daRk lineagES of the fungal tree of life" (STRES) project will cover a broad collection of black fungal diversity spread throughout the Fungal Tree of Life. Interestingly, the STRES project will focus on mostly unsampled genera that display different ecologies and life-styles (e.g., ant- and lichen-associated fungi, rock-inhabiting fungi, etc.). With a resequencing strategy of 10- to 15-fold depth coverage of up to ~550 strains, numerous new reference genomes will be established. To identify metabolites and functional processes, these new genomic resources will be enriched with metabolomics analyses coupled with transcriptomics experiments on selected species under various stress conditions (salinity, dryness, UV radiation, oligotrophy). The data acquired will serve as a reference and foundation for establishing an encyclopedic database for fungal metagenomics as well as the biology, evolution, and ecology of the fungi in extreme environments.
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http://dx.doi.org/10.3390/life10120362DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7767062PMC
December 2020

An Overview of Genomics, Phylogenomics and Proteomics Approaches in Ascomycota.

Life (Basel) 2020 Dec 17;10(12). Epub 2020 Dec 17.

Department of Biotechnology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria.

Fungi are among the most successful eukaryotes on Earth: they have evolved strategies to survive in the most diverse environments and stressful conditions and have been selected and exploited for multiple aims by humans. The characteristic features intrinsic of Fungi have required evolutionary changes and adaptations at deep molecular levels. Omics approaches, nowadays including genomics, metagenomics, phylogenomics, transcriptomics, metabolomics, and proteomics have enormously advanced the way to understand fungal diversity at diverse taxonomic levels, under changeable conditions and in still under-investigated environments. These approaches can be applied both on environmental communities and on individual organisms, either in nature or in axenic culture and have led the traditional morphology-based fungal systematic to increasingly implement molecular-based approaches. The advent of next-generation sequencing technologies was key to boost advances in fungal genomics and proteomics research. Much effort has also been directed towards the development of methodologies for optimal genomic DNA and protein extraction and separation. To date, the amount of proteomics investigations in Ascomycetes exceeds those carried out in any other fungal group. This is primarily due to the preponderance of their involvement in plant and animal diseases and multiple industrial applications, and therefore the need to understand the biological basis of the infectious process to develop mechanisms for biologic control, as well as to detect key proteins with roles in stress survival. Here we chose to present an overview as much comprehensive as possible of the major advances, mainly of the past decade, in the fields of genomics (including phylogenomics) and proteomics of Ascomycota, focusing particularly on those reporting on opportunistic pathogenic, extremophilic, polyextremotolerant and lichenized fungi. We also present a review of the mostly used genome sequencing technologies and methods for DNA sequence and protein analyses applied so far for fungi.
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http://dx.doi.org/10.3390/life10120356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765829PMC
December 2020

The Microbiome of Leonardo da Vinci's Drawings: A Bio-Archive of Their History.

Front Microbiol 2020 20;11:593401. Epub 2020 Nov 20.

Department of Biotechnology, Institute of Microbiology and Microbial Biotechnology, University of Natural Resources and Life Sciences (BOKU), Vienna, Austria.

Seven emblematic Leonardo da Vinci's drawings were investigated through third generation sequencing technology (Nanopore). In addition, SEM analyses were carried out to acquire photographic documentation and to infer the nature of the micro-objects removed from the surface of the drawings. The Nanopore generated microbiomes can be used as a "bio-archive" of the drawings, offering a kind of fingerprint for current and future biological comparisons. This information might help to create a biological catalog of the drawings (cataloging), a microbiome-fingerprint for each single analyzed drawing, as a reference dataset for future studies (monitoring) and last but not least a bio-archive of the history of each single object (added value). Results showed a relatively high contamination with human DNA and a surprising dominance of bacteria over fungi. However, it was possible to identify typical bacteria of the human microbiome, which are mere contaminants introduced by handling of the drawings as well as other microorganisms that seem to have been introduced through vectors, such as insects and their droppings, visible through the SEM analyses. All drawings showed very specific bio-archives, but a core microbiome of bacteria and fungi that are repeatedly found in this type of material as true degraders were identified, such as members of the phyla , , and among bacteria, and fungi belonging to the classes and . In addition, some similarities were observed that could be influenced by their geographical location (Rome or Turin), indicating the influence of this factor and denoting the importance of environmental and storage conditions on the specific microbiomes.
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http://dx.doi.org/10.3389/fmicb.2020.593401DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7718017PMC
November 2020

Author Correction: Shotgun proteomics reveals putative polyesterases in the secretome of the rock-inhabiting fungus Knufia chersonesos.

Sci Rep 2020 Oct 2;10(1):16713. Epub 2020 Oct 2.

Institute of Microbiology and Microbial Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190, Vienna, Austria.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41598-020-70778-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7532174PMC
October 2020

Contamination of wounds with fecal bacteria in immuno-suppressed mice.

Sci Rep 2020 07 13;10(1):11494. Epub 2020 Jul 13.

Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Donaueschingenstraße 13, 1200, Vienna, Austria.

Immunocompromised patients are predisposed to chronically infected wounds. Especially ulcers in the dorsal region often experience secondary polymicrobial infections. However, current wound infection models mostly use single-strain bacteria. To mimic clinically occurring infections caused by fecal contamination in immunocompromised/immobile patients, which differ significantly from single-strain infections, the present study aimed at the establishment of a new mouse model using infection by fecal bacteria. Dorsal circular excision wounds in immunosuppressed mice were infected with fecal slurry solution in several dilutions up to 1:8,000. Impact of immunosuppressor, bacterial load and timing on development of wound infections was investigated. Wounds were analyzed by scoring, 3D imaging and swab analyses. Autofluorescence imaging was not successful. Dose-finding of cyclophosphamide-induced immunosuppression was necessary for establishment of bacterial wound infections. Infection with fecal slurry diluted 1:166 to 1:400 induced significantly delayed wound healing (p < 0.05) without systemic reactions. Swab analyses post-infection matched the initial polymicrobial suspension. The customized wound score confirmed significant differences between the groups (p < 0.05). Here we report the establishment of a simple, new mouse model for clinically occurring wound infections by fecal bacteria and the evaluation of appropriate wound analysis methods. In the future, this model will provide a suitable tool for the investigation of complex microbiological interactions and evaluation of new therapeutic approaches.
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http://dx.doi.org/10.1038/s41598-020-68323-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7359036PMC
July 2020

Shotgun proteomics reveals putative polyesterases in the secretome of the rock-inhabiting fungus Knufia chersonesos.

Sci Rep 2020 06 17;10(1):9770. Epub 2020 Jun 17.

Institute of Microbiology and Microbial Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190, Vienna, Austria.

Knufia chersonesos is an ascomycotal representative of black fungi, a morphological group of polyextremotolerant melanotic fungi, whose ability to resort to recalcitrant carbon sources makes it an interesting candidate for degradation purposes. A secretome screening towards polyesterases was carried out for the fungus and its non-melanized mutant, grown in presence of the synthetic copolyester Polybutylene adipate terephthalate (PBAT) as additional or sole carbon source, and resulted in the identification of 37 esterolytic and lipolytic enzymes across the established cultivation conditions. Quantitative proteomics allowed to unveil 9 proteins being constitutively expressed at all conditions and 7 which were instead detected as up-regulated by PBAT exposure. Protein functional analysis and structure prediction indicated similarity of these enzymes to microbial polyesterases of known biotechnological use such as MHETase from Ideonella sakaiensis and CalA from Candida albicans. For both strains, PBAT hydrolysis was recorded at all cultivation conditions and primarily the corresponding monomers were released, which suggests degradation to the polymer's smallest building block. The work presented here aims to demonstrate how investigations of the secretome can provide new insights into the eco-physiology of polymer degrading fungi and ultimately aid the identification of novel enzymes with potential application in polymer processing, recycling and degradation.
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http://dx.doi.org/10.1038/s41598-020-66256-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7299934PMC
June 2020

Decoding the biological information contained in two ancient Slavonic parchment codices: an added historical value.

Environ Microbiol 2020 08 29;22(8):3218-3233. Epub 2020 May 29.

Institute of Microbiology and Microbial Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 11, A-1190, Vienna, Austria.

This study provides an example in the emerging field of biocodicology showing how metagenomics can help answer relevant questions that may contribute to a better understanding of the history of ancient manuscripts. To this end, two Slavonic codices dating from the 11th century were investigated through shotgun metagenomics. Endogenous DNA enabled to infer the animal origin of the skins used in the manufacture of the two codices, while nucleic sequences recovered from viruses were investigated for the first time in this material, opening up new possibilities in the field of biocodicology. In addition, the microbiomes colonizing the surface of the parchments served to determine their conservation status and their latent risk of deterioration. The saline environment provided by the parchments selected halophilic and halotolerant microorganisms, which are known to be responsible for the biodegradation of parchment. Species of Nocardiopsis, Gracilibacillus and Saccharopolyspora, but also members of the Aspergillaceae family were detected in this study, all possessing enzymatic capabilities for the biodeterioration of this material. Finally, a relative abundance of microorganisms originating from the human skin microbiome were identified, most probably related to the intensive manipulation of the manuscripts throughout the centuries, which should be taken with caution as they can be potential pathogens.
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http://dx.doi.org/10.1111/1462-2920.15064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7687136PMC
August 2020

Peculiar genomic traits in the stress-adapted cryptoendolithic Antarctic fungus Friedmanniomyces endolithicus.

Fungal Biol 2020 05 31;124(5):458-467. Epub 2020 Jan 31.

Department of Microbiology and Plant Pathology, Institute for Integrative Genome Biology, University of California, Riverside, CA, USA. Electronic address:

Friedmanniomyces endolithicus is a highly melanized fungus endemic to the Antarctic, occurring exclusively in endolithic communities of the ice-free areas of the Victoria Land, including the McMurdo Dry Valleys, the coldest and most hyper-arid desert on Earth and accounted as the Martian analog on our planet. F. endolithicus is highly successful in these inhospitable environments, the most widespread and commonly isolated species from these peculiar niches, indicating a high degree of adaptation. The nature of its extremo tolerance has not been previously studied. To investigate this, we sequenced genome of F. endolithicus CCFEE 5311 to explore gene content and genomic patterns that could be attributed to its specialization. The predicted functional potential of the genes was assigned by similarity to InterPro and CAZy domains. The genome was compared to phylogenetically close relatives which are also melanized fungi occurring in extreme environments including Friedmanniomyces simplex, Baudoinia panamericana, Acidomyces acidophilus, Hortaea thailandica and Hortaea werneckii. We tested if shared genomic traits existed among these species and hyper-extremotolerant fungus F. endolithicus. We found that some characters for stress tolerance such as meristematic growth and cold tolerance are enriched in F. endolithicus that may be triggered by the exposure to Antarctic prohibitive conditions.
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http://dx.doi.org/10.1016/j.funbio.2020.01.005DOI Listing
May 2020

Back to the Salt Mines: Genome and Transcriptome Comparisons of the Halophilic Fungus and Its Halotolerant Relative .

Genes (Basel) 2019 05 20;10(5). Epub 2019 May 20.

VIBT EQ Extremophile Center, Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.

Salt mines are among the most extreme environments as they combine darkness, low nutrient availability, and hypersaline conditions. Based on comparative genomics and transcriptomics, we describe in this work the adaptive strategies of the true halophilic fungus , found in a salt mine in Austria, and compare this strain to the ex-type halotolerant fungal strain . On a genomic level, exhibits a reduced genome size compared to , as well as a contraction of genes involved in transport processes. The proteome of exhibits an increased proportion of alanine, glycine, and proline compared to the proteome of non-halophilic species. Transcriptome analyses of both strains growing at 5% and 20% NaCl show that regulates three-times fewer genes than in order to adapt to the higher salt concentration. In , the increased osmotic stress impacted processes related to translation, transcription, transport, and energy. In contrast, membrane-related and lignolytic proteins were significantly affected in .
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http://dx.doi.org/10.3390/genes10050381DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6563132PMC
May 2019

Transcriptome Study of an Mutant on an Skin Model: Is Melanin Important for Infection?

Front Microbiol 2018 3;9:1457. Epub 2018 Jul 3.

Department of Biotechnology, VIBT EQ Extremophile Center, University of Natural Resources and Life Sciences, Vienna, Austria.

The black yeast is a polyextremophilic human pathogen, especially known for growing in man-made extreme environments. Reported diseases caused by this fungus range from benign cutaneous to systemic infections with 40% fatality rate. While the number of cases steadily increases in both immunocompromised and immunocompetent people, detailed knowledge about infection mechanisms, virulence factors and host response are scarce. To understand the impact of the putative virulence factor melanin on the infection, we generated a polyketide synthase () mutant using CRISPR/Cas9 resulting in a melanin deficient strain. The mutant and the wild-type fungus were inoculated onto skin explants using an skin organ culture model to simulate cutaneous infection. The difference between the mutant and wild-type transcriptional landscapes, as assessed by whole RNA-sequencing, were small and were observed in pathways related to the copper homeostasis, cell wall genes and proteases. Seven days after inoculation the wild-type fungus completely colonized the stratum corneum, invaded the skin and infected keratinocytes while the mutant had only partially covered the skin and showed no invasiveness. Our results suggest that melanin dramatically improves the invasiveness and virulence of during the first days of the skin infection.
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http://dx.doi.org/10.3389/fmicb.2018.01457DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6037837PMC
July 2018

Big Sound and Extreme Fungi-Xerophilic, Halotolerant Aspergilli and Penicillia with Low Optimal Temperature as Invaders of Historic Pipe Organs.

Life (Basel) 2018 Jun 14;8(2). Epub 2018 Jun 14.

VIBT-Extremophile Center, University of Natural Resources and Life Sciences Vienna, Muthgasse 18, A-1190 Vienna, Austria.

Recent investigations have shown that xerophilic fungi may pose a biodeterioration risk by threatening objects of cultural heritage including many types of materials, including wood, paint layers, organic glues or leather and even metal. Historic—and also new built—pipe organs combine all those materials. In this study, halotolerant aspergilli and penicillia with low optimal temperatures were shown to be the most frequent invaders of pipe organs. The fungi form white mycelia on the organic components of the organs with a clear preference for the bolus paint of the wooden pipes, the leather-made hinges of the stop actions and all parts fixed by organic glue. Physiological tests showed that the strains isolated from the instruments all show a halotolerant behavior, although none was halophilic. The optimum growth temperature is below 20 °C, thus the fungi are perfectly adapted to the cool and relatively dry conditions in the churches and organs respectively. The genome sequences analyses of the strains are currently ongoing and will reveal the genomic basis for the halotolerant behavior of the fungi.
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http://dx.doi.org/10.3390/life8020022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6027336PMC
June 2018

Draft Genome Sequence of the Saccharomyces cerevisiae Saccharomyces kudriavzevii HA1836 Interspecies Hybrid Yeast.

Genome Announc 2018 May 17;6(20). Epub 2018 May 17.

VIBT-Extremophile Center, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.

× interspecies hybrid yeasts have frequently been isolated from alcoholic fermentation environments. Here, we report the draft genome sequence of the × HA1836 strain isolated from grapes from an Austrian vineyard.
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http://dx.doi.org/10.1128/genomeA.00343-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5958256PMC
May 2018

Draft Genome Sequence of the Interspecies Hybrid Saccharomyces pastorianus Strain HA2560, Isolated from a Municipal Wastewater Treatment Plant.

Genome Announc 2018 Apr 26;6(17). Epub 2018 Apr 26.

VIBT-Extremophile Center, University of Natural Resources and Life Sciences Vienna, Vienna, Austria

is an industrially relevant yeast frequently isolated from brewing environments. Here, we report the draft genome sequence of the HA2560 strain isolated from a municipal wastewater treatment plant.
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http://dx.doi.org/10.1128/genomeA.00341-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5920194PMC
April 2018

Draft Genome Sequences of the Black Rock Fungus and Its Spontaneous Nonmelanized Mutant.

Genome Announc 2017 Nov 2;5(44). Epub 2017 Nov 2.

VIBT-Extremophile Center, University of Natural Resources and Life Sciences, Vienna, Austria.

The fungal genus mostly comprises extremotolerant species from environmental sources, especially rock surfaces. The draft genome sequence of the rock fungus presented here is the first whole-genome sequence of the only species among black fungi known to have a nonmelanized spontaneous mutant.
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http://dx.doi.org/10.1128/genomeA.01242-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5668551PMC
November 2017

Genomic and transcriptomic analysis of the toluene degrading black yeast Cladophialophora immunda.

Sci Rep 2017 09 12;7(1):11436. Epub 2017 Sep 12.

Department of Biotechnology, VIBT-EQ Extremophile Center, University of Natural Resources and Life Sciences, 1190, Vienna, Austria.

Cladophialophora immunda is an ascomycotal species belonging to the group of the black yeasts. These fungi have a thick and melanized cell wall and other physiological adaptations that allows them to cope with several extreme physical and chemical conditions. Member of the group can colonize some of the most extremophilic environments on Earth. Cladophialophora immunda together with a few other species of the order Chaetothyriales show a special association with hydrocarbon polluted environments. The finding that the fungus is able to completely mineralize toluene makes it an interesting candidate for bioremediation purposes. The present study is the first transcriptomic investigation of a fungus grown in presence of toluene as sole carbon and energy source. We could observe the activation of genes involved in toluene degradatation and several stress response mechanisms which allowed the fungus to survive the toluene exposure. The thorough comparative genomics analysis allowed us to identify several events of horizontal gene transfer between bacteria and Cladophialophora immunda and unveil toluene degradation steps that were previously reported in bacteria. The work presented here aims to give new insights into the ecology of Cladophialophora immunda and its adaptation strategies to hydrocarbon polluted environments.
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http://dx.doi.org/10.1038/s41598-017-11807-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595782PMC
September 2017

Aspergillus atacamensis and A. salisburgensis: two new halophilic species from hypersaline/arid habitats with a phialosimplex-like morphology.

Extremophiles 2017 Jul 12;21(4):755-773. Epub 2017 May 12.

Department of Biotechnology, Vienna Institute of Biotechnology (VIBT), University of Natural Resources and Life Sciences, Muthgasse 11, 1190, Vienna, Austria.

Halophilic fungal strains isolated from historical wooden staircase in a salt mine in Austria, and from wall biofilm and soil of a cave in the Coastal Range of the hyperarid Atacama Desert in Chile were characterised and described newly as Aspergillus salisburgensis and Aspergillus atacamensis. Morphological characters including solitary phialides producing solitary conidia and conidia in chains and/or heads suggested affinity to Aspergillus subgenus Polypaecilum. Strains required salt for growth, grew optimally on media with 10-25% NaCl and at 15-28 °C. These values are similar to those observed for Aspergillus salinarus comb. nov. (Phialosimplex salinarum), while the ex-type strains of Aspergillus sclerotialis, Aspergillus chlamydosporus and Aspergillus caninus (all belonging to Aspergillus subgen. Polypaecilum) grew optimally at 0-5% NaCl and showed fastest growth at 28-37 °C. Phylogenetic analyses on the basis of rDNA sequences, RAPD-PCR fingerprint patterns, and cellobiohydrolase gene (cbh-I) polymorphism clustered the strains into three groups and supported their taxonomic recognition as A. salinarus, A. atacamensis and A. salisburgensis. On the basis of phylogenetic inferences, also Sagenomella keratitidis is newly combined as Aspergillus keratitidis and inferred as a species of Aspergillus subgenus Polypaecilum.
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http://dx.doi.org/10.1007/s00792-017-0941-3DOI Listing
July 2017

The Transcriptome of during Skin Model Infection.

Front Cell Infect Microbiol 2016 24;6:136. Epub 2016 Oct 24.

Department of Biotechnology, VIBT EQ Extremophile Center, University of Natural Resources and Life Sciences Vienna, Austria.

The black yeast is a widespread polyextremophile and human pathogen, that is found in extreme natural habitats and man-made environments such as dishwashers. It can cause various diseases ranging from phaeohyphomycosis and systemic infections, with fatality rates reaching 40%. While the number of cases in immunocompromised patients are increasing, knowledge of the infections, virulence factors and host response is still scarce. In this study, for the first time, an artificial infection of an skin model with was monitored microscopically and transcriptomically. Results show that is able to actively grow and penetrate the skin. The analysis of the genomic and RNA-sequencing data delivers a rich and complex transcriptome where circular RNAs, fusion transcripts, long non-coding RNAs and antisense transcripts are found. Changes in transcription strongly affect pathways related to nutrients acquisition, energy metabolism, cell wall, morphological switch, and known virulence factors. The L-Tyrosine melanin pathway is specifically upregulated during infection. Moreover the production of secondary metabolites, especially alkaloids, is increased. Our study is the first that gives an insight into the complexity of the transcriptome of during artificial skin infections and reveals new virulence factors.
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http://dx.doi.org/10.3389/fcimb.2016.00136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5075926PMC
September 2017

Specific Antibodies for the Detection of Alternaria Allergens and the Identification of Cross-Reactive Antigens in Other Fungi.

Int Arch Allergy Immunol 2016 26;170(4):269-278. Epub 2016 Oct 26.

Christian Doppler Laboratory for Allergy Research, Division of Immunopathology, Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria.

Background: The mould Alternaria alternata is an important source of respiratory allergens. A. alternata extracts show great variations regarding allergenic potency. The aim of this study was to generate antibody probes specific for important Alternaria allergens and to use them to study allergen expression, depending on different culture conditions, as well as to search for cross-reactive allergens in other mould species.

Methods: Synthetic peptides from antigenic regions of A. alternata allergens (Alt a 1, Alt a 2, Alt a 3, Alt a 6 and Alt a 8) were used to raise highly specific rabbit antibodies. These antibodies and IgE from allergic patients were used to detect allergens by immunoblotting in extracts of 4 A. alternata strains grown under varying culturing conditions, in commercial skin-prick extracts and in closely (Cladosporium herbarum and Aureobasidium pullulans) or distantly related (Aspergillus niger and Penicillium chrysogenum) mould species.

Results: There was a wide variation of expression of the individual A. Alternata allergens, depending on the strain and culture conditions, but the antibody probes allowed us to distinguish strains and culture conditions with low and high allergen expression. In the commercial skin-prick solutions, varying levels of Alt a 1 were found, but no other allergens were detectable. Alt a 1 was identified as species-specific A. Alternata allergen, whereas Alt a 3, 6- and Alt a 8-cross-reactive antigens were found in C.herbarum and/or A. pullulans.

Conclusions And Clinical Relevance: Peptide-specific antibodies are useful to analyze diagnostic and therapeutic mould extracts, to study the presence of A. Alternata allergens in biological samples and to search for cross-reactive allergens in other mould species.
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http://dx.doi.org/10.1159/000449415DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5321516PMC
February 2017

Characterization of Yeasts and Filamentous Fungi using MALDI Lipid Phenotyping.

J Microbiol Methods 2016 11 18;130:27-37. Epub 2016 Aug 18.

Shimadzu, Wharfside, Trafford Wharf Road, Manchester, M17 1GP, UK.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) becomes the method of choice for the rapid identification of microorganisms (i.e. protein biotyping). Although bacterial identification is already quite advanced, biotyping of other microbes including yeasts and fungi are still under development. In this context, lipids (e.g. membrane phospholipids) represent a very important group of molecules with essential functions for cell survival and adaptation to specific environments and habitats of the microorganisms. Therefore, lipids show the potential to serve as additional molecular parameters to be used for biotyping purposes. In this paper we present a molecular characterisation of yeasts and filamentous fungi based on the analysis of lipid composition by MALDI-MS (i.e. MALDI lipid phenotyping). Using a combination of Principal Component Analysis (PCA) and Hierarchical Clustering we could demonstrate that this approach allowed a classification and differentiation of several groups of yeasts (e.g. Saccharomyces) and filamentous fungi (e.g. Aspergillus, Penicillium, Trichoderma) at the species/strain level. By analysing the MALDI lipid profiles we were able to differentiate 26 closely related yeast strains, for which discrimination via genotypic methods like AFLP in this case are relatively more elaborate. Moreover, employing statistical analysis we could identify those lipid parameters (e.g. PCs and LPCs), which were responsible for the differentiation of the strains, thus providing insights into the molecular basis of our results. In summary, MALDI lipid phenotyping represents a suitable method for fungal characterization and shows the potential to be used as companion tool to genotyping and/or protein biotyping for the characterization and identification of yeasts and fungi in diverse areas (e.g. environmental, pharmaceutical, clinical applications, etc.).
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http://dx.doi.org/10.1016/j.mimet.2016.08.010DOI Listing
November 2016

Genotypic and phenotypic evolution of yeast interspecies hybrids during high-sugar fermentation.

Appl Microbiol Biotechnol 2016 Jul 13;100(14):6331-6343. Epub 2016 Apr 13.

Department of Biotechnology, University of Natural Resources and Applied Life Sciences, Muthgasse 11, A-1190, Vienna, Austria.

The yeasts of the Saccharomyces genus exhibit a low pre-zygotic barrier and readily form interspecies hybrids. Following the hybridization event, the parental genomes undergo gross chromosomal rearrangements and genome modifications that may markedly influence the metabolic activity of descendants. In the present study, two artificially constructed hybrid yeasts (Saccharomyces cerevisiae x Saccharomyces uvarum and S. cerevisiae x Saccharomyces kudriavzevii) were used in order to evaluate the influence of high-sugar wine fermentation on the evolution of their genotypic and phenotypic properties. It was demonstrated that the extent of genomic modifications differs among the hybrids and their progeny, but that stress should not always be a generator of large genomic disturbances. The major genome changes were observed after meiosis in the F1 segregants in the form of the loss of different non-S. cerevisiae chromosomes. Under fermentation condition, each spore clone from a tetrad developed a mixed population characterized by different genotypic and phenotypic properties. The S. cerevisiae x S. uvarum spore clones revealed large modifications at the sequence level of the S. cerevisiae sub-genome, and some of the clones lost a few additional S. cerevisiae and S. uvarum chromosomes. The S. cerevisiae x S. kudriavzevii segregants were subjected to consecutive loss of the S. kudriavzevii markers and chromosomes. Both the hybrid types showed increased ethanol and glycerol production as well as better sugar consumption than their parental strains. The hybrid segregants responded differently to stress and a correlation was found between the observed genotypes and fermentation performances.
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http://dx.doi.org/10.1007/s00253-016-7481-0DOI Listing
July 2016

Pathogenic Yet Environmentally Friendly? Black Fungal Candidates for Bioremediation of Pollutants.

Geomicrobiol J 2016 Mar 25;33(3-4):308-317. Epub 2016 Feb 25.

University of Natural Resources and Life Sciences Vienna, VIBT-Extremophile Center , Vienna , Austria.

A collection of 163 strains of black yeast-like fungi from the CBS Fungal Biodiversity Center (Utrecht, The Netherlands), has been screened for the ability to grow on hexadecane, toluene and polychlorinated biphenyl 126 (PCB126) as the sole carbon and energy source. These compounds were chosen as representatives of relevant environmental pollutants. A microtiter plate-based culture assay was set up in order to screen the fungal strains for growth on the selected xenobiotics versus glucose, as a positive control. Growth was observed in 25 strains on at least two of the tested substrates. Confirmation of substrate assimilation was performed by cultivation on closed vials and analysis of the headspace composition with regard to the added volatile substrates and the generated carbon dioxide. (CBS 120910) and (CBS 110551), both of the order and isolated from a patient with chronic sinusitis and a polluted soil sample, respectively, showed the ability to grow on toluene as the sole carbon and energy source. Toluene assimilation has previously been described for but this is the first account for . Also, this is the first time that the capacity to grow on alkylbenzenes has been demonstrated for a clinical isolate. Assimilation of toluene could not be demonstrated for the human opportunistic pathogen (CBS 115.59, ), but the results from microtiter plate assays suggest that strains of this species are promising candidates for further studies. The outstanding abilities of black yeast-like fungi to thrive in extreme environments makes them ideal agents for the bioremediation of polluted soils, and for the treatment of contaminated gas streams in biofilters. However, interrelations between hydrocarbonoclastic and potentially pathogenic strains need to be elucidated in order to avoid the possibility of biohazards occurring.
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http://dx.doi.org/10.1080/01490451.2015.1052118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4786828PMC
March 2016

Protein functional analysis data in support of comparative proteomics of the pathogenic black yeast Exophiala dermatitidis under different temperature conditions.

Data Brief 2015 Dec 2;5:372-5. Epub 2015 Sep 2.

VIBT Extremophile Center, Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.

In the current study a comparative proteomic approach was used to investigate the response of the human pathogen black yeast Exophiala dermatitidis toward temperature treatment. Protein functional analysis - based on cellular process GO terms - was performed on the 32 temperature-responsive identified proteins. The bioinformatics analyses and data presented here provided novel insights into the cellular pathways at the base of the fungus temperature tolerance. A detailed analysis and interpretation of the data can be found in "Proteome of tolerance fine-tuning in the human pathogen black yeast Exophiala dermatitidis" by Tesei et al. (2015) [1].
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http://dx.doi.org/10.1016/j.dib.2015.08.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4773397PMC
December 2015

Biodeterioration Risk Threatens the 3100 Year Old Staircase of Hallstatt (Austria): Possible Involvement of Halophilic Microorganisms.

PLoS One 2016 17;11(2):e0148279. Epub 2016 Feb 17.

Department of Biotechnology, VIBT-Vienna Institute of BioTechnology, University of Natural Resources and Life Sciences, Vienna, Austria.

Background: The prosperity of Hallstatt (Salzkammergut region, Austria) is based on the richness of salt in the surrounding mountains and salt mining, which is documented as far back as 1500 years B.C. Substantial archaeological evidence of Bronze and Iron Age salt mining has been discovered, with a wooden staircase (1108 B.C.) being one of the most impressive and well preserved finds. However, after its discovery, fungal mycelia have been observed on the surface of the staircase, most probably due to airborne contamination after its find.

Objective: As a basis for the further preservation of this valuable object, the active micro-flora was examined to investigate the presence of potentially biodegradative microorganisms.

Results: Most of the strains isolated from the staircase showed to be halotolerant and halophilic microorganisms, due to the saline environment of the mine. Results derived from culture-dependent assays revealed a high fungal diversity, including both halotolerant and halophilic fungi, the most dominant strains being members of the genus Phialosimplex (synonym: Aspergillus). Additionally, some typical cellulose degraders, namely Stachybotrys sp. and Cladosporium sp. were detected. Numerous bacterial strains were isolated and identified as members of 12 different genera, most of them being moderately halophilic species. The most dominant isolates affiliated with species of the genera Halovibrio and Marinococcus. Halophilic archaea were also isolated and identified as species of the genera Halococcus and Halorubrum. Molecular analyses complemented the cultivation assays, enabling the identification of some uncultivable archaea of the genera Halolamina, Haloplanus and Halobacterium. Results derived from fungi and bacteria supported those obtained by cultivation methods, exhibiting the same dominant members in the communities.

Conclusion: The results clearly showed the presence of some cellulose degraders that may become active if the requirements for growth and the environmental conditions turn suitable; therefore, these microorganisms must be regarded as a threat to the wood.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0148279PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4757552PMC
July 2016

Draft Genome of Debaryomyces fabryi CBS 789T, Isolated from a Human Interdigital Mycotic Lesion.

Genome Announc 2016 Feb 4;4(1). Epub 2016 Feb 4.

University of Natural Resources and Life Sciences Vienna, VIBT-Extremophile Center, Vienna, Austria

The yeast genus Debaryomyces comprises species isolated from various natural habitats, man-made environments, and clinical materials. Here, the draft genome of D. fabryi CBS 789(T), isolated from a human interdigital mycotic lesion, is presented.
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http://dx.doi.org/10.1128/genomeA.01580-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4742678PMC
February 2016

Molecular Tools for Monitoring the Ecological Sustainability of a Stone Bio-Consolidation Treatment at the Royal Chapel, Granada.

PLoS One 2015 29;10(7):e0132465. Epub 2015 Jul 29.

Department of Biotechnology, University of Natural Resources and Life Sciences, VIBT-BOKU, Vienna, Austria.

Background: Biomineralization processes have recently been applied in situ to protect and consolidate decayed ornamental stone of the Royal Chapel in Granada (Spain). While this promising method has demonstrated its efficacy regarding strengthening of the stone, little is known about its ecological sustainability.

Methodology/principal Findings: Here, we report molecular monitoring of the stone-autochthonous microbiota before and at 5, 12 and 30 months after the bio-consolidation treatment (medium/long-term monitoring), employing the well-known molecular strategy of DGGE analyses. Before the bio-consolidation treatment, the bacterial diversity showed the exclusive dominance of Actinobacteria (100%), which decreased in the community (44.2%) after 5 months, and Gamma-proteobacteria (30.24%) and Chloroflexi (25.56%) appeared. After 12 months, Gamma-proteobacteria vanished from the community and Cyanobacteria (22.1%) appeared and remained dominant after thirty months, when the microbiota consisted of Actinobacteria (42.2%) and Cyanobacteria (57.8%) only. Fungal diversity showed that the Ascomycota phylum was dominant before treatment (100%), while, after five months, Basidiomycota (6.38%) appeared on the stone, and vanished again after twelve months. Thirty months after the treatment, the fungal population started to stabilize and Ascomycota dominated on the stone (83.33%) once again. Members of green algae (Chlorophyta, Viridiplantae) appeared on the stone at 5, 12 and 30 months after the treatment and accounted for 4.25%, 84.77% and 16.77%, respectively.

Conclusions: The results clearly show that, although a temporary shift in the bacterial and fungal diversity was observed during the first five months, most probably promoted by the application of the bio-consolidation treatment, the microbiota tends to regain its initial stability in a few months. Thus, the treatment does not seem to have any negative side effects on the stone-autochthonous microbiota over that time. The molecular strategy employed here is suggested as an efficient monitoring tool to assess the impact on the stone-autochthonous microbiota of the application of biomineralization processes as a restoration/conservation procedure.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0132465PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4519126PMC
April 2016

Proteome of tolerance fine-tuning in the human pathogen black yeast Exophiala dermatitidis.

J Proteomics 2015 Oct 17;128:39-57. Epub 2015 Jul 17.

VIBT Extremophile Center, Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.

Unlabelled: The black yeast Exophiala dermatitidis is a worldwide distributed agent of primary and secondary diseases in both immunocompromised and healthy humans, with a high prevalence in human-made environments. Since thermo-tolerance has a crucial role in the fungus persistence in man-dominated habitat and in its pathogenicity, three incubation temperatures (37, 45, 1 °C) and two time spans (1 h, 1 week) were selected to simulate different environmental conditions and to investigate the effect of temperature on the proteome of E. dermatitidis CBS 525.76. Using a novel protocol for protein extraction from black yeasts, 2-D DIGE could be applied for characterization of changes in total protein spot abundance among the experimental conditions. A total of 32 variable proteins were identified by mass spectrometry. Data about protein functions, localization and pathways were also obtained. A typical stress response under non-optimal temperature could not be observed at the proteome level, whereas a reduction of the metabolic activity, mostly concerning processes as the general carbon metabolism, was detected after exposure to cold. These results suggest that a fine protein modulation takes place following temperature treatment and a repertoire of stable protein might be at the base of E. dermatitidis adaptation to altered growth conditions.

Significance: E. dermatitidis is a pathogenic black yeast causing neurotropic infections, systemic and subcutaneous disease in a wide range of hosts, including humans. The discovery of the fungus high prevalence in man-made habitats, including sauna facilities, drinking water and dishwashers, generated concern and raised questions about the infection route. In the present work - which is the first contribution on E. dermatitidis proteome - the effect of different temperature conditions on the fungus protein pattern have been analyzed by using a gel-based approach and the temperature responsive proteins have been identified. The absence of a typical stress response following the exposure to non-optimal temperature was detected at the proteome level, along with a general reduction of the metabolic activity after exposure to cold. These results suggest that a very fine regulation of the protein expression as well as adaptations involving a basic set of stable proteins may be at the base of E. dermatitidis enormous ecological plasticity, which plays a role in the fungus distribution, also enabling the transition from natural to human habitat and to the human host.
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http://dx.doi.org/10.1016/j.jprot.2015.07.007DOI Listing
October 2015

Amid the possible causes of a very famous foxing: molecular and microscopic insight into Leonardo da Vinci's self-portrait.

Environ Microbiol Rep 2015 Dec 19;7(6):849-59. Epub 2015 Aug 19.

Istituto Centrale per il Restauro e la Conservazione del Patrimonio Archivistico e Librario (ICRCPAL), Ministero per i Beni e le Attivita Culturali, Via Milano 76, Rome, 00184, Italy.

Leonardo da Vinci's self-portrait is affected by foxing spots. The portrait has no fungal or bacterial infections in place, but is contaminated with airborne spores and fungal material that could play a role in its disfigurement. The knowledge of the nature of the stains is of great concern because future conservation treatments should be derived from scientific investigations. The lack of reliable scientific data, due to the non-culturability of the microorganisms inhabiting the portrait, prompted the investigation of the drawing using non-invasive and micro-invasive sampling, in combination with scanning electron microscope (SEM) imaging and molecular techniques. The fungus Eurotium halophilicum was found in foxing spots using SEM analyses. Oxalates of fungal origin were also documented. Both findings are consistent with the hypothesis that tonophilic fungi germinate on paper metabolizing organic acids, oligosaccharides and proteic compounds, which react chemically with the material at a low water activity, forming brown products and oxidative reactions resulting in foxing spots. Additionally, molecular techniques enabled a screening of the fungi inhabiting the portrait and showed differences when different sampling techniques were employed. Swabs samples showed a high abundance of lichenized Ascomycota, while the membrane filters showed a dominance of Acremonium sp. colonizing the drawing.
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http://dx.doi.org/10.1111/1758-2229.12313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4959533PMC
December 2015

From Glacier to Sauna: RNA-Seq of the Human Pathogen Black Fungus Exophiala dermatitidis under Varying Temperature Conditions Exhibits Common and Novel Fungal Response.

PLoS One 2015 10;10(6):e0127103. Epub 2015 Jun 10.

VIBT-Extremophile Center, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria.

Exophiala dermatitidis (Wangiella dermatitidis) belongs to the group of the so-called black yeasts. Thanks in part to its thick and strongly melanized cell walls, E. dermatitidis is extremely tolerant to various kinds of stress, including extreme pH, temperature and desiccation. E. dermatitidis is also the agent responsible for various severe illnesses in humans, such as pneumonia and keratitis, and might lead to fatal brain infections. Due to its association with the human environment, its poly-extremophilic lifestyle and its pathogenicity in humans, E. dermatitidis has become an important model organism. In this study we present the functional analysis of the transcriptional response of the fungus at 1°C and 45°C, in comparison with that at 37°C, for two different exposition times, i.e. 1 hour and 1 week. At 1°C, E. dermatitidis uses a large repertoire of tools to acclimatize, such as lipid membrane fluidization, trehalose production or cytoskeleton rearrangement, which allows the fungus to remain metabolically active. At 45°C, the fungus drifts into a replicative state and increases the activity of the Golgi apparatus. As a novel finding, our study provides evidence that, apart from the protein coding genes, non-coding RNAs, circular RNAs as well as fusion-transcripts are differentially regulated and that the function of the fusion-transcripts can be related to the corresponding temperature condition. This work establishes that E. dermatitidis adapts to its environment by modulating coding and non-coding gene transcription levels and through the regulation of chimeric and circular RNAs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0127103PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4463862PMC
March 2016