Course FOOD and AIRBORNE FUNGI CBS-KNAW Fungal Biodiversity Centre, Utrecht Course leaders and assistants Rob Samson Jos Houbraken Tineke van Doorn Martin Meijer Neriman Yilmaz Programme of the course Telephone, fax and Email Telephone 030 2122600 Telefax 030 2512097 CBS has a wireless network for laptops (ask password) 1
Programme of the course 8-12 October 2012 Programme Monday October 8 10.00 Opening 10.10 Introduction (R.A. Samson) 11.30 Zygomycetes 12.00-15.00 Microscopy Zygomycetes 15.00 Aspergillus 15.20-17.00 Microscopy Aspergillus Tuesday October 9 10.00 Methods for isolation of food-borne Fungi (J. Houbraken) 11.00 Ascomycetes 11.30-15.00. Microscopy Ascomycetes 15.00 Penicillium 15.30 17.00 Microscopy Penicillium Wednesday October 10 10.00 Molecular methods for identifying food-borne fungi. An overview (J. Houbraken) 11.00 Microscopy Penicillium 13.30 Fusarium 14.30 17.00 Microscopy Fusarium 15.00 Conidiogenesis of Deuteromycetes Thursday October 11 10.00 Detection of Airborne fungi (R.A. Samson) 11.30 17.00 Microscopy Deuteromycetes Friday October 12 10.00 Mycotoxins 10.45 Yeasts 11.00-14.00 continuation Microscopy Deuteromycetes 14.00 General Discussion Dinner together with the course participants 3 course menu is Euro 19.75 For some dishes there is an extra charge Programme of the course Course from 9.00-17.00 Lecture mostly starting 10.00 and 15.30 Coffee and tea breaks Lunch 13.00-13.30 2
Programme of the course Lunch 13.00 Programme of the course CBS rules Please wear name badges all the time No smoking or food in the laboratory Please wear laboratory coat when possible NO mobile phones in the course room Use only one strain each time and return the plates Do not leave the plates open to avoid release of spores in the air Be careful with the burners and put them off when you are not using it Try to work with your neighbour(s) Programme of the course Introduction to the fungal flora of food and the indoor environment Introduction to the important groups of Zygomycetes, Ascomycetes and Deuteromycetes Microscopical examination of cultures Lectures on applied aspects 3
Rotterdam Amsterdam Baarn Utrecht CBS was founded in 1904 and is since 1968 an institute of the Royal Netherlands Academy of Arts and Sciences. The institute moved in December 2000 from Baarn to Utrecht Frits Went (1863-1935) professor in botany at the University of Utrecht The initiator of CBS Prof. Johanna Westerdijk (1883-1961) Phytopathologist and the first female professor in the Netherlands Centraalbureau voor Schimmelcultures the oldest and largest culture collection of filamentous fungi and yeasts Since 2001 also including the NCCB bacteria and phages collections more than 65.000 strains are preserved for long-term maintenance 4
Research programmes Applied and Industrial Mycology (R.A. Samson) Bioinformatics group (V. Robert) Fungal Physiology (R. de Vries) Origin of Pathogenicity in Clinical Fungi (G.S. de Hoog) Collection, Preservation and Digitalisation (G. Verkleij) Evolutionary Phytopathology (P.W. Crous) Yeast and Basidiomycete Research (T. Boekhout) 5
Penicillium roqueforti on rye bread 6
Bathroom Pathogenic Fungi There are relatively very few fungi which are dangerous for man and animals However when the immune system is affected opportunistic species which can commonly occur become pathogenic Aspergillus fumigatus mostly not common in indoorenvironments important in clinical environments when present, it can be easily be detected in air samples incubation temperature important quantitative data should be treated carefully 7
Penicilllium chrysogenum An important and useful mould because of the production of the antibiotic penicillin Very common species on food and in indoor environments can produce several mycotoxins Aflatoxins Aspergillus flavus Aspergillus parasiticus Aspergillus nomius Tremorgenic activity Haemorrhage of the lung and brain Immune surpression Carcinogenic Nephrotoxic Oestrogenic Hepatic necrosis Haemaorrhage of liver MYCOTOXINS Gastro-intestinal disturbances Idiopathic pulmonary hemosiderosis Dermal toxicosis Parasympathetic nerve system 8
Stachybotrys chartarum common in temperate and humid climates produces mycotoxins (e.g. satratoxins) respiratory tract bleeding non-respiratory symptoms Indication to cause death in infants Mycotoxins in the indoor environments Fungi which occur in indoor environments are similar to those found in food These fungi are also able to produce mycotoxins These toxic metabolites can be inhaled directly or via the spores Where are the Fungi? In principle everywhere on earth 9
Growth of fungi Source Carbon Temperature 0-60 C, optimal 25-30 C ph 1-12, optimal at neutral ph water oxygen The biodiversity of fungi on food and in indoorenvironments Saccharomyces cerevisiae, a yeast with one celled structures 10
Penicillium expansum in culture Apples contaminated with Penicillium expansum 11
hypha hypha Sporangiophore Mycelium of a Mucor Coenocytic mycelium of a Zygomycete Mycelium of a hyphomycete (Wallemia sebi) 12
Coniophora puteana hyphe with clamp connections Numbers of known and estimated total species of organisms group known taxa total taxa vascular plants 220.000 270.000 bryophytes 17.000 25.000 algae 40.000 60.000 fungi 75.000 >1.500.000 bacteria 3.000 >1.000.000 viruses 5.000 130.000 13
Zygomycetes: Mucor, Rhizopus, Absidia Ascomycetes: Eurotium, Emericella, Neosartorya Basidiomycetes: Agaricus, Serpula Deuteromycetes belong either to the Ascomycetes or Basidiomycetes but they do not have a sexual form (teleomorph): Penicillium, Aspergillus, Cladosporium Yeasts are Ascomycetes or Basidiomycetes Zygomycota or Zygomycetes 14
rhizoids stolon Rhizopus stolonifer Rhizopus columella apophysis Sexual production in the Zygomycetes 15
Ascomycota; Ascomycetes Diverse group of fungi, multiplying in a yeast-like manner without fruit-bodies or produce mycelium with fruit-bodies (= ascomata) Worldwide distribution; soil, dung, plants, food and feed Saprobes or parasites to humans, animals, plants In food or indoor environments a limited number of genera occur and are mostly very simple. Eurotium: ascoma and asci Ascus with ascospores Ascoma (cleistothecium) Sexual reproduction in the Ascomycetes 16
Ascospores of Eurotium herbariorum Emericella nidulans Anamorph: Aspergillus nidulans Hülle cells Deuteromycetes or anamorphic fungi 17
Conidiogenesis Conidiogenesis = the mode of conidium formation Conidia = specialized non-motile asexual spore Conidiogenous cell = specialized cell which give rise to the conidia Conidiophore = entire system of fertile hyphae Thallic and blastic conidiogenesis Thallic conidia in Geotrichum candidum 18
Blastic conidiogensis in Botrytis Poroconidia in Alternaria Phialides with conidia in chains 19
The common mycoflora in food and in indoor environments Zygomycetes (about 10-15 species) Ascomycetes (about 10-15 species) Deuteromycetes (about 100-150 species) List of fungal species occurring in indoor environments Curvularia lunata Absidia corymbifera Emericella nidulans Penicillium olsonii Acremonium mumorum Epicoccum nigrum Penicillium rugulosum Acremonium strictum Eurotium amstelodami Penicillium simplicissimum Alternaria alternaria Eurotium chevalieri Penicillium spinulosum Aspergillus candidus Eurotium herbariorum Penicillium variabile Aspergillus clavatus Exophalia dermatitidis Phialophora fastigiata Aspergillus flavus Fusarium culmorum Phialophora verrucosa Aspergillus flavipes Fusarium solani Phoma glomerata Aspergillus fumigatus Fusarium verticillioides Phoma macrostoma Aspergillus niger Geomyces pannorum Pithomyces chartarum Aspergillus ochraceus Geotrichum candidum Pyronema domesticum Aspergillus penicillioides Memnoniella echinata Rhizopus stolonifer Aspergillus restrictus Mucor hiemalis Rhodotorula mucilaginosa Aspergillus sydowii Mucor plumbeus Schizophyllum commune Aspergillus terreus Mucor racemosus Scopulariopsis brevicaulis Aspergillus versicolor Oidiodendron griseum Scopulariopsis candida Aureobasidium pullulans Oidiodendron rhodogenum Scopulariopsis fusca Botrytis cinerea Paecilomyces lilacinus Serpula lacrymans Candida peltata Paecilomyces variotii Sistotrema brinkmanii Chaetomium aureum Penicillium Sporobolomyces roseus Chaetomium globosum aurantiogriseum Stachybotrys chartarum Chaetomium indicum Penicillium Syncephalastrum racemosum Chrysonillia sitophila brevicompactum Trichoderma harzianum Cladosporium cladosporioides Penicillium chrysogenum Trichoderma koningii Cladosporium herbarum Penicillium citrinum Trichoderma viride Cladosporium Penicillium commune Tritirachium oryzae sphaerospermum Penicillium corylophilum Ulocladium chartarum Clonostachys rosea Penicillium expansum Verticillium lecanii Coprinus cordisporus Penicillium glabrum Wallemia sebi Cryptococcus laurentii Penicillium janthinellium Toxinogenic pathogenic xerophilic bath rooms and wet environments Identification of moulds There are presently no fast methods for identification of genus or species Morphological examination is essential New molecular approaches are developed The new morphological data show that some common species are species complexes Examples are Fusarium, Cladosporium, Aspergillus In Penicillium and Aspergillus a polyphasic approach is used, combining morphology, biochemistry and molecular data (for example β- tubulin and calmodulin sequences) 20
Beta tubulin phylogeny of section Fumigati 100 N. warcupii NRRL 35723 A. unilateralis CBS 126.56 N. papuensis CBS 841.96 N. assulata KACC 41691 100 N. aurata CBS 466.65 N. stramenia CBS 498.65 N. galapagensis CBS 117522 N. multiplicata CBS 646.95 N. australensis CBS 112.55 86 N. fennelliae CBS 598.74 100 N. denticulata CBS 652.73a N. ferenczii NRRL 4179 N. glabra CBS 111.55 N. nishimurae IFM 54133 A. turcosus KACC 42091 N. tatenoi CBS 407.93 97 N. aureola CBS 105.55 N. udagawae CBS 114217 A. viridinutans CBS 127.56 95 A. laciniosa KACC 41657 N. spinosa CBS 483.65 76 98 A. fumigatiaffinis IBT 12703 A. novofumigatus IBT 16806 N. coreana KACC 41659 86 N. fischeri CBS 544.65 85 A. fumigatus CBS 133.61 99 A. lentulus FH5 A. fumisynnematus IFM 42277 N. hiratsukae CBS 294.93 N. spathulata CBS 408.89 N. pseudofischeri CBS 208.92 N. quadricincta CBS 135.52 A. brevipes CBS 118.53 A. duricaulis CBS 481.65 A. clavatus CBS 513.65 0.02 Programme of the course Introduction to the Zygomycetes Microscopical examination of cultures 21