The Genus Penicillium a Guide for Historical, Classification and Identification of Penicilli, Their Industrial Applications and Detrimental Effects

Home , Penicillium adametzii, Penicillium atramentosum, Penicillium atrovenetum, Penicillium bilaiae, Penicillium brasilianum, Penicillium brocae

Monograph On The genus Penicillium A guide for historical, classification and identification of penicilli, their industrial applications and detrimental effects

By Mohamed Refai1, Heidy Abo El-Yazid1 and Wael Tawakkol2

1. Department of Microbiology, Faculty of Veterinary Medicine, Cairo University 2. Department of Microbiology, Faculty of Pharmacy, Misr University for Science and Technology

2015

Google: Examples of sites Ancient Egyptians and penicillin 22 Likes - Instagram instagram.com/p/sHcNA4R6YN/ ... as penicillium mold is used today). Imhotep an Egyptian was a polymath from the 27th century BC. He is one of the first physicians or doctors known to history. Ancient Egyptians used penicillin | Matter Of Facts https://matteroffactsblog.wordpress.com/.../ancient-egyptians-used-pencil... Oct 9, 2013 - Imhotep: a polymath from the 27th century BC. ... because of the presence of antibacterial molds (just as penicillium mold is used today).

Preface

When I was in El-Badrashen primary school (1944-1950) I used to visit my classmates living in the nearby villages of Memphis and Sakkara. There, I heard from girls and women that they liked to eat mouldy bread because they thought it rendered their hairs smooth and shiny. When, I searched the internet using the key words Ancient Egyptians and Penicillium. Surprisingly, I found several sites indicating that the Egyptian polymath, Imhoteb, was one of the first physicians or doctors known to history, who prescribed mouldy bread for healing wounds, i.e. he predicted the presence of antibiotics in moulds 4700 years ago.

Prof. Dr. Mohamed Refai, March 2015

Contents 1. Introduction , 4 2. Penicillium history, 5 3. Penicillium classification17 2. 1. Classification after Raper and Thom, 1949, 17 3.2. Claasification according to Ramirez 1982, 21 3. 3. Classification according Houbraken & Samson (2011) , 27 3. 4. Updated Classification, 39 4. Morphology of Penicillium species, 49 4.1. Macromorphology of Penicillium species, 51 4.2. Micromorphology of Penicillium, 58 5. Extrolites produced by various Penicillium species, 66 5.1. Antibiotics, 66 5.2. Anti-cancer,66 5. 3. Penicillium pigments, 68 5.4. Enzymes ,68 5.5. Mycotoxins ,69 6. Molecular genetics of Penicillium ,72 6.1. Genome sequencing,72 6.2. Genetic transformation, 76 6.3. The biosynthetic pathway, 79 6.4. Virulence genes of Penicillium digitatum, 81 6.5. Sexual life cycle of P. roqueforti ,82 6.6. Genome shuffling ,83 6.7. Mechanism of conidiation in P. decumbens, 83 6.8. Expression and characterization of enzymes, 84 6.9. Molecular identification, 88 7. Industral applications of Penicillium species, 89 7.1. Penicillin, 89 7. 2. Griseofulvin, 95 7. 3. Roquefort cheese, 98 7. 4. Camembert cheese ,103 7. 5. Gorgonzola cheese ,108 7. 6. Cambozola cheese ,110 7. 7. Penicillium-cured salami, I110 7. 8. Penicillium pigments ,113 7. 9. Penicillium enzymes, 119 7. 10. Production of metal nanoparticles, 119 8. Health risks and economic losses caused by Penicillium species, 120 9. Penicillium species identification, 120 10.Gallery of Penicillia, 136 11. References. 149

1. Introduction

Penicillium is a well known cosmopolitan genus of moulds that comprises more than 350 species playing various roles in natural ecosystems, agriculture and biotechnology. They have double faces, a good and beneficial one and a bad and economically destructive one. Examples of the beneficial roles are: Penicillium chrysogenum produces the antibacterial antibiotic penicillin, Penicillium griseofulvum produces the antifungal antibiotic griseofulvin, several Penicillium species produce anti-cancer substances such as Penicillium albocoremium (Andrastin A), P. decumbens (Bredenin). Penicillium roqueforti is used for the production of Roquefort cheese and Penicillium camemberti is used for the production of Camembert cheese. Several Penicillium species produce enzymes that are used in industry, e.g. cellulases and xylanases produced by Penicillium species have broad applications in food and feed, the textile industry, and the pulp and paper industries. Penicillium species are also used for biodegradation of oil and can be used in restoring the ecosystem when contaminated by oil. Peroxidase enzyme of Penicillium species have potential biodegradable activities that degrade Amaranth dye, Orange G, heterocyclic dyes like, Azure B and Lip dye. Morepver, some species function as decomposers of dead materials and can be used in recycling of waste products. Recently, Penicillium species, such as P. aurantiogriseum, P. citrinum, and P. waksmanii, are used for the eco-friendly biosynthesis of gold nanoparticles from a solution of AuCl. Gold nanoparticles are formed fairly uniform with spherical shape with the Z-average diameter of 153.3 nm, 172 nm and 160.1 nm for the 3 species, respectively. On the other hand, some species are known to cause postharvest diseases, e.g. Penicillium expansum is one of the most prevalent post-harvest rots that infects apples. Although it is a major economic problem in apples, this plant pathogen can be isolated from a wide host range, including pears, strawberries, tomatoes, corn, and rice. This mould also produces the carcinogenic metabolite patulin, a neurotoxin that is harmful in apple juice and apple products. patulin in food products is a health concern because many are consumed by young children. In addition, a second secondary metabolite citrinin is produced as well. Mould growth on citrus fruits during storage is a continuing problem that results in economic loss. Although several fungal species have been reported to be involved in the spoilage of citrus products, Penicillium digitatum (green mold) and Penicillium italicum (blue mold) are the primary organisms involved.

Penicillium is one of the first fungi to grow on water-damaged materials and has been implicated in causing allergic reactions, hypersensitivity pneumonitis, and a variety of severe lung complications. It may cause sarcoidosis, fibrosis, or allergic alveolitis in susceptible individuals, or patients who have been exposed over long periods of time, depending on the strain. P. oxalicum has also been reported to cause genital infection of water buffalo.

2. Penicillium history

1809, Link created the genus Penicillium, he described 3 species, namely P. candidum, P. expansum and P. glaucus 1824, Link called all green Penicillia, P. glaucum

Heinrich Friedrich Link

1837-1839, Corda illustrated the morphology of several P. species 1874, Brefeld published his detailed report on production of ascospore by P. glaucum 1880, Saccardo described P. digitatum

Saccardo August Karl Corda Julius Oscar Brefeld

1889, Zukal described P. luteum

1892, Ludwig proposed the name Eupenicillium for the ascosporic species

1895, Wehmer published his studies of Penicillia occurring upon rotting fruits. He described P. italicum

1901, Dierckx published his essai, in which he proposed 25 new P. species. Accepted species are: P. citreonigrum , P. corylophilum , P. roseopurpureum , P. brevicompactum , P. aurantiogriseum ,.P. hirsutum ., P. griseofulvum ,P. verrucosum

1906, Thom published his study on Penicillia in connection with chease

1911, Westling described series of the genus Penicillium: P. frequentans, P. lividum ,P. piscarium ,P. glabrum , P. turbatum , P. lanosum , P. viridicatum , P. solitum , P. cyclopium , P. palitans , P. frequentans

1912, Sopp described 60 P. species in his monograph including P.canescens , P. albidum, P.islandicum, P. variabile Sopp

1907-1912, Bainier published description of several species of P. Penicillium paxilli , P. herquei 1912., P. olsonii 1912.

1915, Thom published on the group concept of classification of the genus Penicillium

1915 Grig.-described P. multicolor

1923, Biourge redescribed 125 species of Penicillium, his monograph represented the most comprehensive and elaborate study of the genus made up to that time, with drawings and colored plates and classified Penicillium into sections, subsections and series, the accepted species are: P. aurantioviolaceum, P. roseomaculatum , P. fuscum , P. fellutanum P. sanguifluum , P. cinerascens P. chermesinum ,P. coeruleum, P. janthinellum , P. ochrochloron , P.cyaneum, P. dierckxii, P. sublateritium, P. rubens

1923, Demelius described P. glaucoroseum P. clavigerum

1927, Zaleski described 35 new P. species and one variety. Accepted species are: P. trzebinskii , P. miczynskii , P. chrzaszczii , P. godlewskii ,

P. steckii , P. waksmanii , P. westlingii , P. namyslowskii , P. raciborskii , P. adametzii , Penicillium janczewskii, P. jensenii, P. bialowiezense , P. soppii , P. swiecickii , P. polonicum

1927, J.C. Gilman & E.V. Abbott described Penicillium restrictum , Penicillium vinaceum .

1928, Flemming and Florey discovered the penicillin

Sir Alexander Fleming Lord Florey of Adelaide

Beyma 1929-1940: described P. baarnense, and P. javanicum J. 1929 , P. phoeniceum , 1933 , P. egyptiacum, 1933, P. velutinum . 1935 P. sclerotiorum , 1937, . Penicillium euglaucum . 1940 and Penicillium novae-zeelandiae . 1940

1906-1930, Thom, described many species, the following are the currently accepted species: P. camemberti . 1906, P. decumbens . 1910. P. spinulosum 1910, P. chrysogenum . 1910, P. citrinum 1910. P. commune, 1910.,P. biforme . 1910, P. atramentosum , 1910., P. lanosocoeruleum, 1930, P. oxalicum . 1915., P. crustosum . 1930, P. roqueforti . 1930, P. rolfsii, 1930 P. simplicissimum ,1930, P. melinii . 1930,

1930, Thom published his comprehensive monograph. He reexamined all material up to that time from cultures grown in the laboratory under uniform conditions. He emphasized the group concept of classification and separated Paecilomyces, Gliocladium and Scopulariopsis from Penicillum and treated them as related genera.

1931, Hetherington, A. C., and H. Raistrick. 1931. Were the first to isolate Citrinin from Penicillium citrinum Thom

Charles Thom

1934, Shear described P. asperum

1939, Oxford, Raistrick and Simonart discovered griseofulvin from P. griseofulvum. It was developed initially as an agricultural product and later as a systemic fungistatic for human and veterinary application, griseofulvin proved a revolutionary agent in treating Dermatophytoses.

1943, Anslow et al. isolated patulin, an antibiotic, from Penicillium patulum, known at present as a mycotoxin and the fungus is currently named P. griseofulvum. 1949, Raper and Thom published their famous Manual of the Penicillia, which described 99 accepted species and varieties of Penicillium sensu stricto and listed almost 700 species

Kenneth Raper 1950, Chalabuda.,. described the following species” P. glauco-lanosum, P. bilaiae, P. kurssanovii, P. cinereo-atrum albo-cinerascens, P. arenicola, Penicillium cremeogriseum

1955, Benjamin created the genus Talaromyces for the ascosporic species of Penicillium, which comprise the P. luteum series of Raper and Thom

1959-1983, Udagawa et al. P. cinnamopurpureum 1959. , P. ochrosalmoneum . 1959 , P. hirayamae . 1959 , P.fractum . 1968 , P. reticulisporum . 1968 , P. ornatum ,1968, P. ludwigii . 1969 , P. philippinense . 1972, P. rubidurum 1973 , P. rubidurum . 1973 , P. gracilentum . 1973 , P.caperatum . 1973 , P. meloforme . 1973, P. lineolatum . 1977 , P. sinaicum,1982, P. nepalense , . 1983, P. angustiporcatum , . 1983

1957-1974, Stolk and Samson described the following species: P. isariiforme 1957., P. anatolicum ,P. katangense , P. abidjanum, P. shearii 1967, P. striatisporum 1969, P. hordei . 1969., P. donkii . 1973..P. osmophilum . 1974

1968, D.B. Scott described the following species P. alutaceum , , P. erubescens , P. catenatum , P. meridianum , P. terrenum , P. inusitatum , P. senticosum , P. stolkiae

1968, Baghdadi described the following species P. arabicum, P. baradicum, P. eben-bitarianum, P. damascenum Baghdadi P. gorlenkoanum P. harmonense, P. sizovae, P. yarmokense, P. kabunicum

1971, Stolk and Samson created the genus Hamigera for the species of Talaromyces characterized by asci developing singly from croziers, and redefined the genus Talaromyces by restricting it to species which produce asci in chains.

1972, Stolk & Samson Published their book on the Genus Talaromyces 1973, Stolk & Samson Published their book on the Genus Eupenicillium

Stolk & Samson 1972 Stolk & Samson 1983

1973, Pitt published his appraisal of identification methods for Penicillium species:Novel taxonomic criteria based on temperature and water relations

1974, Pitt published a synoptic key to the genus Eupenicillium and to sclerotigenic Penicillium species

1978-1984C. Ramírez et al. described the following species” P. grancanariae . 1978. ,P. hispanicum 1978., P. palmense ,.P. fagi , P. malacaense ,P. valentinum . P. vasconiae . P. galliacum .,1980, P. onobense 1981, P.flavidostipitatum . 1984, P. jugoslavicum . 1984.

1979, Pitt published his book The genus Penicillium and its teleomorphic states Eupenicillium and Talaromyces. Pitt divided Penicillium into four subgenera based on conidiophore morphology and branching pattern: Aspergilloides, Biverticillium, Furcatum, and Penicillium.

John I Pitt

1981, Frisvad suggested that extrolites could be used directly in penicilli Taxonomy

1982, Carlos Ramirez published his Manual and Atlas of the Penicllia. It contains description of 227 species and varieties of Penicillium , 252 coloured plates of Penicillium cultures, each plate presents a type species grown upon three standard media(Czapek, Czapek-yeast and Malt extract agars, viewed from the obverse and reverse, and 38 SEM plates

Cuau Ramirez Angel T Martinez 1976-2011, Seifert et al.,. described the following species:

P. concentricum 1985, P. vulpinum 1985, P.glandicola 1985, P. coprophilum 1985, P. kananaskense . P. terrigenum 2011

1994, Tzean, Shean-Shong published Penicillium and related teleomorphs from Taiwan.

Tzean, Shean-Shong Long Wang

2000.-2014, Wang etal. described the following species. Penicillium ellipsoideosporum L2000, Penicillium macrosclerotiorum 2007, Penicillium saturniforme ,. Penicillium zhuangii . 2014., Penicillium fusisporum 2014

2000, Robert A. Samson and John Pitt published their book

1990-2013, Frisvad et al. described the following species P. coprobium . 1990., P. mononematosum, . 1990, P. scabrosum . 1990, P. confertum . 1990 , , P. tricolor 1994, P. carneum 1996 , P. paneum . 1996, P. flavigenum . 1997, P. discolor . 1997., P. albocoremium . 2000 , P. albocoremium . 2000 , P. venetum 2000 , P. cavernicola. , P. marinum . 2000, P. dipodomyicola 2000, P. dipodomyis 2000 P. radicicola . 2003., P. tulipae . 2003., P. thymicola,2004, P. freii ,2004, P., melanoconidium, 2004, P. ribium, 2006, P. jamesonlandense 2006, P. svalbardense 2007. P. halotolerans 2012., P. tardochrysogenum 2012, P. allii-sativi . 2012., P. vanluykii . 2012, P. desertorum 2012.,P. buchwaldii . 2013., P. spathulatum

2000-2014, Wang et al. described the following species P. ellipsoideosporum , 2000. ,. P. saturniforme , P. persicinum ., 2004, P. brevistipitatum , 2005, P. macrosclerotiorum . 2007Penicillium kongii . 2013, P. zhuangii . 2014., P. fusisporum 2014

2008, Genome sequencing and analysis of the filamentous fungus Penicillium chrysogenum were published by Marco A van den Berg1, Richard Albang2, Kaj Albermann2, Jonathan H Badger3, Jean- Marc Daran4,5, Arnold J M Driessen4,6, Carlos Garcia-Estrada7, Natalie D Fedorova3, Diana M Harris4,5, Wilbert H M Heijne8, Vinita Joardar3, Jan A K W Kiel9, Andriy Kovalchuk6, Juan F Martín7,10, William C Nierman3,11, Jeroen G Nijland6, Jack T Pronk4,5, Johannes A Roubos8, Ida J van der Klei4,9, Noël N M E van Peij8, Marten Veenhuis9, Hans von Döhren12, Christian Wagner2, Jennifer Wortman3 & Roel A L Bovenberg 2010-2014, Houbraken et al, described the following species” P. psychrosexualis . P. hetheringtonii , P. tropicoides , P. tropicum ,P. argentinense , P. atrofulvum , P. aurantiacobrunneum , P. cairnsense , P. christenseniae , . P. copticola , P. cosmopolitanum , . P. nothofagi , P. pancosmium , P. pasqualense , P. quebecense , P. raphiae , P. ubiquetum , P. vancouverense , P. araracuaraense , P. elleniae , P. penarojense , P. vanderhammenii ,P. wotroi , P. hennebertii , P. longisporum , P. porphyreum , P. laeve , P. ovatum , P. bovifimosum , P.malachiteum, P. subrubescens, Penicillium armarii ), Penicillium athertonense , Penicillium austroafricanum , P. bussumense ,P. cartierense , P. contaminatum ,P. grevilleicola , P. hoeksii , P. kiamaense, P. pulvis , P. ranomafanaense , P. rudallense , P. sterculiniicola , P. sublectaticumm, P . subspinulosum , P. tsitsikammaense , P. vagum , P. verhagenii

2011, K.G. Rivera et al. described the following species P. cainii ,, P. guanacastense , P. mallochii , P. jacksonii ., Penicillium johnkrugii .

2011, Robert A. Samson and Jos Houbraken published their book : Phylogenetic and taxonomic studies on the genera Penicillium and Talaromyces (Studies in Mycology, 70 (September 2011)

2012, Complete mitochondrial genome of compactin-producing fungus Penicillium solitum and comparative analysis ofTrichocomaceae mitochondrial genomes Michael A. Eldarov , Andrey V. Mardanov , Alexey V. Beletsky , Vakhtang V. Dzhavakhiya , Nikolai V. Ravin , Konstantin G. Skryabin

Marco A van den Berg1 Michael A. Eldarov Marina

2012, Genome sequence of the necrotrophic fungus Penicillium digitatum, the main postharvest pathogen of citrus Marina Marcet-Houben, Ana-Rosa Ballester, Beatriz de la Fuente, Eleonora Harries, Jose F Marcos, Luis González- Candelas3* and Toni Gabaldón

2013, Genomic and Secretomic Analyses Reveal Unique Features of the Lignocellulolytic Enzyme System of Penicillium decumbens Guodong Liu., Lei Zhang., Xiaomin Wei., Gen Zou., Yuqi Qin, Liang Ma , Jie Li, Huajun Zheng , Shengyue Wang , Chengshu Wang , Luying Xun, Guo- Ping Zhao, Zhihua Zhou, Yinbo Qu

Marcet-Houben Guodong Liu

2013-2014, Visagie, et al.,. described the following species P. arianeae ,. P. alexiae , P. maximae , P. amaliae , P. vanoranjei ,P. infra-aurantiacum , P. clavistipitatum , P. flavisclerotiatum , P. brunneoconidiatum , P. longicatenatum , P. malmesburiense , P. turcosoconidiatum , P. infrapurpureum , P. sucrivorum . P. dunedinense , P. magnielliptisporum , P. mexicanum , P. lenticrescens, P. alfredii

Robert Samson Jos Houbraken

2014, Robert A Samson, Cobus M Visagie, Jos Houbraken published a book on Species Diversity in Aspergillus, Penicillium and Talaromyces. 49 Penicillium and 18 Talaromyces; 22 that were described as new) have been identified and barcoded using the ITS gene, as well as Beta-tubulin for Penicillium and Talaromyces.

Cobus M Visagie Jens Frisvad

2014, Draft Genome Sequence of Penicillium expansum Strain R19, Which Causes Postharvest Decay of Apple Fruit Jiujiang Yu, Wayne M. Jurick II, Huansheng Cao, Yanbin Yin,Verneta L. Gaskins, Liliana Losada, Nikhat Zafar, Maria Kim,Joan W. Bennettd, William C. Nierman

2014, Multiple recent horizontal transfers of a large genomic region in cheese making fungi by Kevin Cheeseman, Jeanne Ropars, Pierre Renault, Joëlle Dupont, Jérôme GouzyAntoine Branca, Anne-Laure Abraham, Maurizio Ceppi, Emmanuel Conseiller, Robert Debuchy, Fabienne Malagnac, Anne Goarin, Philippe Silar, Sandrine Lacoste, Erika Sallet, Aaro Bensimon,Tatiana Giraud & Yves Brygoo

2014, Genome sequencing and analysis of the paclitaxel-producing endophytic fungus Penicillium aurantiogriseumNRRL 62431 Yanfang Yang , Hainan Zhao , Roberto A Barrero, Baohong Zhang, Guiling Sun, Iain W Wilson, Fuliang Xie, Kevin D Walker, Joshua W Parks, Robert Bruce, Guangwu Guo, Li Chen, Yong Zhang, Xin Huang, Qi Tang, Hongwei Liu, Matthew Bellgard, Deyou Qiu*, Jinsheng Lai and Angela Hoffman

Huansheng Cao Jeanne Ropars Roberto A Barrero3

3. Penicillium classification 2. 1. Classification after Raper and Thom, 1949

1. The Monoverticillata

Penicillium javanicum Series 1. Penicillium javanicum van Beyma, 2. PenicilUum parvum Raper and Fennel 3. Peniculium brefeldiamon Dodge 4. Penicillium ehrlichii Klebahn 5. Peniciuium levitum Raper and Fennell Penicillium thomii Series 6. Penicillium thomii Maire 7. Penicillium scleroliorum van Beyma 8. Penicillium lapidosum Raper and Fennell 9. Penicillium turbatum Westling, Penicillium frequentans series 10. Penicillium frequentans Westling 11. Penicillium purpurrescens (Sopp) 12. Penicillium spinulosum Thom Penicillium lividum Series 13. PenicilUum lividum Westling 14. PenicilUum aurantio-violaceum Biourge 15. PenicilUum trzebinskii Zaleski Penicillium implicatum Series 16. Penicilum implicatum Biourge 17. Peniciluum multicolor 18. PeniciluUum sublateritium Biourge Penicillium decumbens Series 19. Penicillium decumhens Thom, 20. Penicillium fellutanum Biourge 21. Penicilum chermesinum Biourge 22. Penicilium citreo-viride Biourge 23. Penicilium roseo-purpureum Dierckx Penicillium restrictum Series 24. Penicillium restrictum Oilman and Abbott 25. Penicillium fuscum (Sopp) Penicillium adametzi Series 26. Penicillium adametzi Zaleski 27. Penicillium terlikowskii Zaleski 28. Penicillium vinaceum Oilman and Abbott 29. Penicillium phoeniceum van Beyma, The Ramigena Series 30. Penicillium capsulatnm Raper and Fennell 31. Penicillium cyaneum (B. and S.) Biourge 32. Penidllium waksmani Zaleski 33. Penicillium charlesii Smith 34. Penicillium velutinum van Beyma

2. The ASYMMETRICA

2.1.ASYMMETRICA Sub-section: DIVARICATA

Carpenteles Series 35. Penicillium asperum (Shear) 36. Penicillium haarnense van Beyma 37. Penicillium egyptiacum van Beyma Penicillium raistrickii series 38. Penicillium raistrickii Smith 39. Penicillium pulvillorum Turfitt 40. Penicillium soppi Zaleski 41. Penicillium rolfsii Thom Penicillium lilacinum Series 42. Penicillium lilacinum Thom, Penicillium janthinellum Series 43. Penicillium daleae Zaleski 44. Penicillium janthinellum Biourge 45. Penicillium simplicissimum (Oud.) Thorn 46. Penicillium ochro-chloron Biourge 47. Penicillium miczyjnskii Zaleski Penicillium canescens series 48. Penicillium canescens Sopp 49. Penicillium nalgiovensis 50. Penicillium jensenii Penicillium nigricans Series 51. P. nigricans (Bainier) Thorn 52. P. albidum Sopp 53. P. kapuscinskii Zaleski 54. P. melinii Thom 55. P. raciborskii Zaleski

2.2. ASYMMETRICA Sub-section:VELUTINA P. citrinum series 56. P. corylophilum Dierckx 57. P. citrinum Thom 58. P. steckii Zaleski P. chrysogenum series 59. P. chrysogenum Thom 60. P. meleagrinum Biourge 61. P. notatum Westling 62. P. cyaneofulvum Biourge P. oxalicum series 63. P. oxalicum Currie and Thom 64. P. atramentosum Thom P. digitatum series 65. P. digitatum Sacc. P. roqueforti series 66. P. roqueforti Thom 67. P. casei Staub P. brevi-compactum series 68. P. hrevi-compactum Dierckx 69. P. stoloniferum Thom 70. P. paxilli Bainier

2.3. ASYMMETRICA Sub-section: LANATA

P. camemberti series 71. Penicillium caseicolum Bainier 72. Penicillium camemherti Thom P. commune series 73. Penicillium commune Thom 74. Penicillium lanosum Westling 75. Penicillium. lanosoviride 76. Penicillium. lanosocoeruleum 77. Penicillium lanosogriseum 78. Penicillium biforme 79. Penicillium. aurantiocandidum

2.4. ASYMMETRICA Sub-section: FUNICULOSA

Penicillium terrestre Series 80. Penicillium psittacinum Thom, 81. Penicillium terrestre Jensen 82. Penicillium solitum Westling 83. Penicillium resticulosum Birkinshaw Penicillium pallidum Series 84. Penicillium pallidum Smith 85. Penicillium putterillii Thom 86. Penicillium lavendulum Raper and Fennell 87. Penicillium namysloivskii Zaleski

2.5. ASYMMETRICA Sub-section: FASCICULATA

P. gladioli series 86. P. gladioli Machacek P. raistrickii series 87. Penicillium raistrickii P. ochraceum series 88. Penicillium ochraceum (Bainier) Thorn 89. Penicillium carneo-lutescens Smith P. viridicatum series 90. Penicillium viridicatum Westling 91. Penicillium olivino-viride Biourge 92. Penicillium palitans Westling, P. cyclopium series 93. Penicillium cyclopium Westling 94. P. cyclopiumvar . echinulatum 95. Penicillium. martensii Biourge 96. Penicillium aurantio-virens Biourge 97. Penicillium puberulum Bainier P. expansum series 98. Penicillium expansum Link 99. P. crustosum Thorn P. ilalicum series 100. P. italicum Wehmer P. urticae series

101. P. urticae Bainier P. granulatum series 102. PenicilUum corymhiferum 103. Penicillium granulatmn Bainier Penicillium claviforme Series 104. Penicillium claviforme Bainier

3. BIVERTICILLATA-SYMMETRICA

Penicillium luteum Series 105. Penicillium luteum Zukal 106. Penicillium striatum Raper and Fennell Penicillium duclauxi Series 107. Pemcillium dudauxi Delacroix Penicillium funiculosum Series 108. Penicillium funiculosum. Thom 109. Penicillium verruculosum Peyronel 110. Penicillium islandicum Sopp 111. Penicillium varians Smith 112. Penicillium piceum Raper and Fennell Penicillium purpurogenum Series 113. Penicillium purpurogenum Stoll 114. Penicillium rubrum Stoll 115. Penicillium variahile Sopp Penicillium rugulosum Series 116. Penicillium rugulosum Thom 117. Pemcillium diversum Raper and Fennell Penicillium herquei Series 118. Penicillium novae-zeelandiae van Beyma

4. POLYVERTICILLATA GLIOCLADIUM, PAECILOMYCES, AND SCOPULARIOPSIS

3.2. Claasification according to Ramirez 1982

1. The Monoverticillata P. thomii series 1. P. thomii Matre 2. P. scleratiorum van Beyma 3. P. syriacum Baghdadi 4. P. turbatum Westling 5. P. pusillum Smith 6. P. indicum Sandhu et Sandhu 7. P. donkii Stolk 8. P. grancanariae Ramirez, Martinez et Ferrer P. frequentans series 9. P. frequentans Westling 10. P. purpurescens (Sopp) Raper et Thom 11. P. spinulosum thom 12. P. cremeo-griseum Chalabuda 13. P. griseo-azureum Moreau et MOREAU ex Ramirez 14. P. luteo-aurantium Smith 15. P. abeanum Smith 16. P. odoratum Christensen et Backus 17. P. tarrconense Ramirez et Martinez P. lividum series 18. P. lividum Westling 19. P. aurantio-violaceum Biourge 20. P. trzebinskii Zaleski 21. P. trezbinskianaum Abe ex Ramirez 22. P. valentinum Ramirez et Martinez 23. P. multicolor Grigorieva-Manoilova et Poradielova 24. P. implicatum Biourge 25. P. sublateritium Biourge 26. P. aeneum Smith 27. P. ramusculum Batista et MAAIA 28. P. palmensis Martinez et Ferrer 29. P. hispanicum Martinez, Ferrer et Martinez 30. P. ardesiacum Novobranova 31. P. gallaicum Ramirez, Martinez et Berenguer P. decumbens series 32. P. decumbens Thom 33. P. chermesinum Biourge 34. P. citreo-viride Biourge 35. P. fellutanum Biourge 36. P. roseo-purpureum Dierckx 37. P. glauco-lanosum Chalabuda 38. P. brevissimum Rai et Wadhwani 39. P. bilaiae Chalabuda 40. P. kurssanovii Chalabuda 41. P. cinereo-atrum Chalabuda 42. P. taxicarium Miyake et R%amirez 43. P. gerundense Ramirez et Martinez 44. P. alicantinum Ramirez et Martinez 45. P. malacaense Ramirez et Martinez P. restrictum series 46. P. restrictum Gilman et Abbott

47. P. montanense Christensen et Backus 48. P. atra-virens Smith 49. P. fuscum (Sopp)Raper et Thom 50. P. arabicum Baghdadi 51. P. raperi Smith 52. P. dimorphosporum Swart 53. P. striatisporum Stolk 54. P. vascaniae Ramirez et Martinez P. adametzii series 55. P. adametzii Zaleski 56. P. terlikowskii Zaleski 57. P. spinulo-ramigenum Sasaki et Ramirez 58. P. griseolum Smith 59. P. adametzioides Abe et Smith 60. P. resedanum McLennan, Ducker et Thrower 61. P. lilacino-echinulatum Abe 62. P. kazachstanicum Novobranova 63. P. albo-cinerascens Chalabuda 64. P. sacculum Dale 65. P. vinaceum Gilman et Abbott 66. P. phoeniceum van Beyma The Ramigena series 67. P. capsulatum Raper et Fennell 68. P. cyaneum (Bainier et Sartory) Biourge 69. P. waksmanii Zaleski 70. P. charlesii Smith 71. P. velutinum van Beyma 72. P. sartoryi Thom

2. The Asymmetrica 2.1. The Asymmetrica-divaricata sub-section

P. raistrickii series 73. P. raistrickii Smith 74. P. pulvillorum Turfitt 75. P. soppii Zaleski 76. P. rolfsii Thom 77. P.rolfii Thom var. sclerotiale Novobranova 78. P. pedemontanum Luppi-Mosca et Fontana 79. P. mirabile Beeljakova Milko P. lilacinum series 80. P. lilacinum Thom 81. P. humuli van Beyma 82. P. argillaceum Stolk 83. P. cylindrosporum Smith P. janthinellum series 84. P. janthinellum Biourge 85. P. daleae Zaleski 86. P. simplicissima (oudemans) Thom 87. P. ochro-chloron Biourge 88. P. piscarium Westling 89. P. miczynskii Zaleski 90. P. asturianum Ramirez et Martinez 91. P. aragonense Ramirez et Martinez

P. godlewskii series 92. P. P. godlewskii Zaleski 93. P.botryosum Batista et Maia 94. P. baradicum Baghdadi 95. P. eben-bitarianum Baghdadi 96. P. damascenum Baghdadi 97. P. gorlenkoanum Baghdadi 98. P. harmonense Baghdadi 99. P. sizovae Baghdadi 100. P. novae-caledoniae Smith 101. P. novae-caledoniae Smith var album Ramirez et Martinez 102. P. citreo-virens Abe ex Ramirez P. canescens series 103. P. canescens Sopp 104. P.nalgiovense Laxa 105. P. jensenii Zaleski 106. P.murcianum Ramirez et Martinez 107. P. turolense Ramirez et Martinez P. nigricans series 108. P. nigricans (Bainier) Thom 109. P. albidum Sopp 110. P. kapuscinskii Zaleski 111. P. melinii Thom 112. P. raciborskii Zalenski 113. P. radulatum Smith 114. P. granatense Ramirez, Martinez et Berenguer 115. P. inflatum Stolk et Malla 116. P. megasporum Orpurt et Fennell 117. P. avetense Ramirez et Martinez 118. P. yarmokense Baghdadi P. atro-sanguineum series 119. P. atro-sanguineum Dong 120. P. griseo-purpureum Smith P. brasilianum series 121. P. brasilianum Batista 122. P. kabunicum Baghdadi 123. P.moldavicum Milko et Beljakova 124. P. onobense Ramirez et Martinez 125. P. castellonense Ramirez et Martinez

2.2. Asymmetrica-velutina sub-section

P. citrinum series 126. P. citrinum Thom 127. P. corylophilum Diercks 128. P. stecki Zaleski 129. P. matriti Smith 130. P. chrysogenum Thom 131. P. chrysogenum Thom mut. Fulvescens Takashima P. oxalicum series 132. P. oxalicum Currie et Thom 133. P. atramentosum Thom 134. P. fennelliae Stolk 135. P. digitatum Saccardo 136. P. japonicum Smith

P. roqueforti series 137. P. roqueforti Thom 138. P. fagi Martinez et Ramirez 139. P. cardubense Ramirez et Martinez 140. P. mali Novobranova 141. P. farinsum Novobranova P. brevi-compactum 142. P. brevi-compactum Diercks 143. P. stoloniferum Thom 144. P. paxilli Bainier 145. P. volgaense Baljakova 146. P. skrjabinii Shmotina et Golovleva 147. P. arenicola Chalabuda 148. P. brunneo-stoloniferum Abe et Ramirez

2.3. Asymmetrica-lanata sub-section

P. camembertii series 149. P. camembertii Thom 150. P. griseum Bonorden P. commune series 151. P. commune Thom 152. P. lanosum Westling 153. P. echinosporum Nehira et Ramirez 154. P. giganteum Roy et Singh

2.4.Asymmetrica-funiculosa sub-section

P. pallidum series 155. P. pallidum Smith 156. P. putterilli Thom 157. P. namyslowskii Zaleski 158. P. lavendulum Raper et Fennell 159. P. gladioli McCulloch et Thom

2.5.Asymmetrica-fasciculata sub-section

P. verrucosum Diercks complex 160. P. verrucosum Diercks var cyclopium (Westling) Samson, Stolk et Hadlok 161. P. verrucosum Diercks var verrucosum Samson, Stolk et Hadlok 162. P. verrucosum Diercks var album (Smith) Samson, Stolk et Hadlok 163. P. verrucosum Diercks var melanochlorum Samson, Stolk et Hadlok 164. P. verrucosum Diercks var ochraceum (Thom) Samson, Stolk et Hadlok 165. P. verrucosum Diercks var corymbiferum (Westling) Samson, Stolk et Hadlok 166. P. echinulatum Fassatiova 167. P. verrucosum Diercks var cyclopium Samson et al.strain ananas-olens Ramirez 168. P. expansum Link 169. P. italicum Wehmer var. italicum Samson, Stolk et Hadlok 170. P. italicum Wehmer var. avellaneum Samson, et Gutter 171. P. hispalense Ramirez et Martinez 172. P.griseo-fulvum Diercks

173. P.granulatum Bainier 174. P. claviforme Bainier 175. P. claviforme Bainier mut. Candicans 176. P. claviforme Bainier mut. olivicolor 177. P. clavigerum 178. P.hordei Stolk 179. P. isariforme Stolk et Meyer 180. P. concentricum Samson, Stolk et Hadlok 181. P. kojigenum Smith 182. P.marneffei Segretain, Capponi et Sureau ex Ramirez

3. The Biverticillata-symmetrica or Lanceolata

P. herquei series 183. P. herquei Bainier et Sartory 184. P. novae-zeelandiae van Beyma 185. P.para-herquei Abe ex Smith 186. P.olsonii Bainier et Sartory 187. P. caralligerum Nicot et Pionnat 188. P.atro-venetum Smith 189. P. estinogenum Komatsu et Abe ex Smith P. funiculosum series 190. P. funiculosum Thom 191. P. verruculosum Peyronei 192. P.islandicum Sopp 193. P. variana Smith 194. P. piceum Raper et Fennell 195. P.allahabadense Mehrotra et Kumar 196. P. lignorum Stolk 197. P. korosum Rai, Wadhwani et Tewari 198. P. brunneum Udagawa 199. P.ilerdanum Ramirez,Martinez et Berenguer 200. P.aurantiacum Miller, Giddens et Foster 201. P.asperosporum Smith 202. P. rubicundum Miller, Goddena etFoster 203. P.gaditanum Ramirez et Martinez P. duclauxii series 204. P. P. duclauxii Delacroix 205. P. pseudostromaticum Hodges, Werner et Rogerson P. purpurogenum series 206. P. purpurogenum Stoll 207. P. rubrum Stoll 208. P.aculeatum Raper et Fennell 209. P. variabile Sopp 210. P. aurantio-flammiferum Ramirez, Berenguer et Martinez 211. P.brasiliense Thom P. rugulosum series 212. P. rugulosum Thom 213. P.tardum Thom 214. P.diversum Raper et Fennell 215. P.diversum var. aureum Raper et Fennell

Closely related species 216. P. zacinthae Ramirez et Martinez 217. P. phialosporum Udagawa 218. P. albo-aurantium Smith 219. P. ingelheimense van Beyma 4. The Polyverticillata 220. P. albicans Bainier 221. P. canadense Smith 222. P. sclerotigenum Yamamoto 223. P. brevi-compactum Diercks var. magnum var nov 224. P. avellaneum Thom et Turesson Addendum 225. P. dendriticum Pitt 226. P. loliense Pitt 227. P. erythromellis Hocking

 Based on the inferred phylogenetic relationships among the Penicillia, they proposed a sectional classification and subdivided Penicillium into two subgenera and 25 sections, with section Aspergilloides being one of them.  Following the concepts of nomenclatural priority and single name nomenclature,  All accepted species of Penicillium subgenus Biverticillium were transferred to Talaromyces. including teleomorph and anamorph characters  Eupenicillium and several other genera were considered as synonyms to Penicillium species(Houbraken & Samson 2011),  Aspergillus paradoxus, A. crystallinus and A. malodoratus were shown to belong to Penicillium and were transferred in Visagie et al. (2014).  Several species described as Penicillium belongs to other genera and not to Penicillium.

3. 3. Classification according Houbraken & Samson (2011)

The genus Aspergillus is classified into two subgenera and 25 sections.

A. Penicillium subgenus Aspergilloides.

1: Section Aspergilloides:

1. Penicillium ardesiacum Novobranova, Novosti Sist. Nizs. Rast. 11: 228. 1974. 2. Penicillium asperosporum Smith, Trans. Br. Mycol. Soc. 48: 275. 1965. 3. Penicillium crocicola Yamamoto, Scient. Rep. Hyogo Univ. Agric., Agric. Biol. Ser. 2, 2: 28. 1956. 4. Penicillium fuscum (Sopp) Biourge, Cellule 33: 103. 1923 (Stolk & Samson 1983). 5. Penicillium georgiense Peterson & Horn, Mycologia 101: 79. 2009. 6. Penicillium glabrum (Wehmer) Westling, Ark. Bot. 11: 131. 1911 (syn. P. terlikowskii; Barreto et al. 2011). 7. Penicillium kananaskense Seifert, Frisvad & McLean, Can. J. Bot. 72: 20. 1994 (unpubl. data, K.A. Seifert). 8. Penicillium lapatayae Ramírez, Mycopathol. 91: 96. 1985 (Frisvad et al. 1990c). 9. Penicillium lividum Westling, Ark. Bot. 11: 134. 1911. 10. Penicillium montanense Christensen & Backus, Mycologia 54: 574. 1963. 11. Penicillium odoratum Christensen & Backus, Mycologia 53: 459. 1962 (this study, Fig. 8). 12. Penicillium palmense Ramírez & Martínez, Mycopathol. 66: 80. 1978. 13. Penicillium patens Pitt & Hocking, Mycotaxon 22: 197. 1985. 14. Penicillium quercetorum Baghdadi, Nov. Sist. Niz. Rast. 5: 110. 1968. 15. Penicillium saturniforme (Wang & Zhuang) Houbraken & Samson, Stud. Mycol. 70: 48. 2011 (this study). 16. Penicillium spinulosum Thom, Bull. Bur. Anim. Ind. U.S. Dep. Agric. 118: 76. 1910. 17. Penicillium subericola Barreto, Frisvad & Samson, Fungal Diversity 49: 32. 2011. 18. Penicillium thiersii Peterson, Bayer & Wicklow, Mycologia 96: 1283. 2004. 19. Penicillium thomii Maire, Bull. Soc. Hist. Nat. Afrique N. 8: 189. 1917.

2: section Sclerotiora Houbraken & Samson, sect. nov.MycoBank

20. Penicillium adametzii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 507. 1927. 21. Penicillium adametzioides Abe ex Smith, Trans. Br. Mycol. Soc. 46: 335. 1963. 22. Penicillium angulare Peterson, Bayer & Wicklow, Mycologia 96: 1289. 2004. 23. Penicillium bilaiae Chalabuda, Bot. Mater. Otd. Sporov. Rast. 6: 165. 1950. 24. Penicillium brocae Peterson, Pérez, Vega & Infante, Mycologia 95: 143. 2003.

25. Penicillium cainii Rivera & Seifert, Stud. Mycol. 70: 147. 2011. 26. Penicillium guanacastense Rivera, Urb & Seifert, Mycotaxon, in press. 2011. 27. Penicillium herquei Bainier & Sartory, Bull. Soc. Mycol. France 28: 121. 1912. 28. Penicillium hirayamae Udagawa, J. Agric. Sci. Tokyo Nogyo Daigaku 5: 6. 1959. 29. Penicillium jacksonii Rivera & Seifert, Stud. Mycol. 70: 151. 2011. 30. Penicillium johnkrugii Rivera & Seifert, Stud. Mycol. 70: 151. 2011. 31. Penicillium jugoslavicum Ramírez & Muntañola-Cvetkovic, Mycopathol. 88: 65. 1984. 32. Penicillium malachiteum (Yaguchi & Udagawa) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 (this study). 33. Penicillium mallochii Rivera, Urb & Seifert Mycotaxon, in press. 2011. 34. Penicillium nodositatum Valla, Plant and Soil 114: 146. 1989. 35. Penicillium sclerotiorum van Beyma, Zentralbl. Bakteriol., 2. Abt., 96: 418. 1937. 36. Penicillium viticola Nonaka & Masuma, Mycoscience 52: 339. 2011.

3: section Charlesia Houbraken & Samson, sect. nov. MycoBank

37. Penicillium charlesii Smith, Trans. Br. Mycol. Soc. 18: 90. 1933. 38. Penicillium coffeae Peterson, Vega, Posada & Nagai, Mycologia 97: 662. 2005. 39. Penicillium fellutanum Biourge, Cellule 33: 262. 1923. 40. Penicillium georgiense Peterson & Horn, Mycologia 101: 79. 2009. 41. Penicillium indicum Sandhu & Sandhu, Can. J. Bot. 41: 1273. 1963 (syn. P. gerundense, Peterson & Horn 2009). 42. Penicillium phoeniceum van Beyma, Zentralbl. Bakteriol., 2. Abt., 88: 136. 1933.

4: section Thysanophora Houbraken & Samson, sect. nov. MycoBank

43. Penicillium asymmetricum (Subramanian & Sudha) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 . 44. Penicillium coniferophilum Houbraken & Samson, Stud. Mycol. 70: 47. 2011 . 45. Penicillium glaucoalbidum (Desmazières) Houbraken & Samson, Stud. Mycol.70: 47. 2011 . 46. Penicillium hennebertii Houbraken & Samson, Stud. Mycol. 70: 47. 2011 . 47. Penicillium longisporum (Kendrick) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 48. Penicillium melanostipe Houbraken & Samson, Stud. Mycol. 70: 47. 2011. 49. Penicillium taiwanense (Matsushima) Houbraken & Samson, Stud. Mycol. 70: 48. 50. Penicillium taxi Schneider, Zentralblatt für Bakteriologie und Parasitenkunde, Abteilung 2, 110: 43. 1956.

5: section Ochrosalmonea Houbraken & Samson, sect. nov. MycoBank

51. Penicillium isariiforme Stolk & Meyer, Trans. Br. Mycol. Soc. 40: 187. 1957. 52. Penicillium ochrosalmoneum Udagawa, J. Agric. Sci. Tokyo Nogyo Daigaku 5: 10. 1959.

6: Section Cinnamopurpurea Houbraken & Samson, sect. nov. MycoBank

53. Penicillium chermesinum Biourge, Cellule 33: 284. 1923. 54. Penicillium cinnamopurpureum Udagawa, J. Agric. Food Sci., Tokyo 5: 1. 1959. 55. Penicillium ellipsoideosporum Wang & Kong, Mycosystema 19: 463. 2000. 56. Penicillium idahoense Paden, Mycopath. Mycol. Appl. 43: 261. 1971 (Peterson & Horn 2009, this study). 57. Penicillium incoloratum Huang & Qi, Acta Mycol. Sin. 13: 264. 1994. 58. Penicillium malacaense Ramírez & Martínez, Mycopathologia 72: 186. 1980 (syn. P. ovetense, this study) (Peterson & Horn 2009). 59. Penicillium nodulum Kong & Qi, Mycosystema 1: 108. 1988. 60. Penicillium parvulum Peterson & Horn, Mycologia 101: 75. 2009. 61. Penicillium shennangjianum Kong & Qi, Mycosystema 1: 110. 1988.

7: section Ramigena Thom, The Penicillia: 225. 1930.

62. Penicillium capsulatum Raper & Fennell, Mycologia 40: 528. 1948. 63. Penicillium cyaneum (Bainier & Sartory) Biourge, Cellule 33: 102. 1923. 64. Penicillium dierckxii Biourge, Cellule 33: 313. 1923. 65. Penicillium hispanicum Ramírez, Martínez & Ferrer, Mycopathol. 66: 77. 1978 66. Penicillium ornatum Udagawa, Trans. Mycol. Soc. Japan 9: 49.1968. 67. Penicillium ramusculum Batista & Maia, Anais Soc. Biol. Pernamb. 13: 27. 1955 ( 68. Penicillium sublateritium Biourge, Cellule 33: 315. 1923.

8: section Torulomyces (Delitsch) Stolk & Samson, Adv. Pen. Asp. Syst.: 169. 1985.

69. Penicillium cryptum Gochenaur, Mycotaxon 26: 349. 1986. 70. Penicillium lagena (Delitsch) Stolk & Samson, Stud. Mycol. 23:100. 1983. 71. Penicillium laeve (K. Ando & Manoch) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 72. Penicillium lassenii Paden, Mycopathol. Mycol. Appl. 43: 266. 1971. 73. Penicillium ovatum (K. Ando & Nawawi) Houbraken & Samson, Stud. Mycol. 70: 48. 2011

74. Penicillium parviverrucosum (K. Ando & Pitt) Houbraken & Samson, Stud. Mycol. 70: 48. 2011 75. Penicillium porphyreum Houbraken & Samson, Stud. Mycol. 70: 48. 2011 (this study).

9: section Fracta Houbraken & Samson, sect. nov. MycoBank

76. Penicillium fractum Udagawa, Trans. Mycol. Soc. Japan 9: 51. 1968. 77. Penicillium inusitatum Scott, Mycopathol. Mycol. Appl. 36: 20. 1968.

10: section Exilicaulis Pitt, The Genus Penicillium: 205. 1980.

78. Penicillium alutaceum Scott, Mycopathol. Mycol. Appl. 36: 17. 1968. 79. Penicillium atrosanguineum Dong, Ceská Mycol. 27: 174. 1973. 80. Penicillium burgense Quintanilla, Avances Nutr. Mejora Anim. Aliment. 30: 176. 1990. 81. Penicillium catenatum Scott, Mycopathol. Mycol. Appl. 36: 24. 1968. 82. Penicillium chalybeum Pitt & Hocking, Mycotaxon 22: 204. 1985. 83. Penicillium cinerascens Biourge, Cellule 33: 308. 1923. 84. Penicillium cinereoatrum Chalabuda, Bot. Mater. Otd. Sporov. Rast. 6: 167, 1950 (Frisvad et al. 1990c). 85. Penicillium citreonigrum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901. 86. Penicillium corylophilum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901. 87. Penicillium decumbens Thom, Bull. Bur. Anim. Ind. U.S. Dep. Agric. 118: 71. 1910. 88. Penicillium dimorphosporum Swart, Trans. Br. Mycol. Soc. 55: 310. 1970. 89. Penicillium dravuni Janso, Mycologia 97: 445. 2005. 90. Penicillium erubescens Scott, Mycopathol. Mycol. Appl. 36: 14. 1968. 91. Penicillium fagi Ramírez & Martínez, Mycopathol. 63: 57. 1978. 92. Penicillium flavidostipitatum Ramírez & González, Mycopathol. 88: 3. 1984 (preliminary sequencing results show that this species is closely related to P. namyslowskii). 93. Penicillium guttulosum Gilman & Abbott, Iowa State Coll. J. Sci. 1: 298. 1927 (Peterson et al. 2011). 94. Penicillium heteromorphum Kong & Qi, Mycosystema 1: 107. 1988. 95. Penicillium katangense Stolk, Ant. van Leeuwenhoek 34: 42. 1968. 96. Penicillium lapidosum Raper & Fennell, Mycologia 40: 524. 1948. 97. Penicillium maclennaniae Yip, Trans. Br. Mycol. Soc. 77: 202. 1981. 98. Penicillium melinii Thom, Penicillia: 273. 1930. 99. Penicillium menonorum Peterson, IMA Fungus 2: 122. 2011. 100. Penicillium meridianum Scott, Mycopathol. Mycol. Appl. 36: 12. 1968. 101. Penicillium namyslowskii Zaleski, Bull. Int. Aead. Polonc. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 479. 1927. 102. Penicillium nepalense Takada & Udagawa, Trans. Mycol. Soc. Japan 24: 146. 1983.

103. Penicillium parvum Raper & Fennell, Mycologia 40: 508. 1948 (this study). 104. Penicillium philippinense Udagawa & Y. Horie, J. Jap. Bot. 47: 341. 1972. 105. Penicillium pimiteouiense Peterson, Mycologia 91: 271. 1999. 106. Penicillium raciborskii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 454. 1927. 107. Penicillium restrictum Gilman & Abbott, Iowa State Coll. J. Sci. 1: 297. 1927. 108. Penicillium rubefaciens Quintanilla, Mycopathol. 80: 73. 1982. 109. Penicillium rubidurum Udagawa & Horie, Trans. Mycol. Soc. Japan 14: 381. 1973. 110. Penicillium smithii Quintanilla, Avances Nutr. Mejora Anim. Aliment. 23: 340. 1982 111. Penicillium striatisporum Stolk, Ant. van Leeuwenhoek 35: 268. 1969. 112. Penicillium terrenum Scott, Mycopathol. Mycol. Appl. 36: 1. 1968. 113. Penicillium toxicarium Miyake, Rep. Res. Inst. Rice Improvement 1: 1. 1940 (nom. inval., Art. 36) (Serra et al. 2008). 114. Penicillium velutinum van Beyma, Zentralbl. Bakteriol., 2. Abt., 91: 353. 1935. 115. Penicillium vinaceum Gilman & Abbott, Iowa State Coll. J. Sci. 1: 299. 1927.

11: Section Lanata-divaricata Thom, The Penicillia: 328. 1930.

116. Penicillium abidjanum Stolk, Ant. van Leeuwenhoek 34: 49. 1968. 117. Penicillium araracuarense Houbraken, C. López-Q, Frisvad & Samson, Int. J. Syst. Evol. Microbiol. 61: 1469. 2011. 118. Penicillium brasilianum Batista, Anais Soc. Biol. Pernambuco 15: 162. 1957. 119. Penicillium brefeldianum Dodge, Mycologia 25: 92. 1933 (syn. P. dodgei). 120. Penicillium caperatum Udagawa & Horie, Trans. Mycol. Soc. Japan 14: 371. 1973 (syn. E. brefeldianum sensu Pitt). 121. Penicillium cluniae Quintanilla, Avances Nutr. Mejora Anim. Aliment. 30: 174. 1990. 122. Penicillium coeruleum Sopp apud Biourge, Cellule 33: 102. 1923. 123. Penicillium cremeogriseum Chalabuda, Bot. Mater. Otd. Sporov. Rast. 6: 168. 1950. 124. Penicillium daleae Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 495. 1927. 125. Penicillium ehrlichii Klebahn, Ber. Deutsch. Bot. Ges. 48: 374. 1930. 126. Penicillium elleniae Houbraken, C. López-Q, Frisvad & Samson, Int. J. Syst. Evol. Microbiol. 61: 1470. 2011. 127. Penicillium glaucoroseum Demelius, Verh. Zool.-Bot. Ges. Wien 72: 72. 1923. (unpubl. data) 128. Penicillium griseopurpureum Smith, Trans. Br. Mycol. Soc. 48: 275. 1965 (unpubl. data). 129. Penicillium janthinellum Biourge, Cellule 33: 258. 1923. 130. Penicillium javanicum van Beyma, Verh. Kon. Ned. Akad. Wetensch., Afd. Natuurk., Tweede Sect., 26: 17. 1929 (syn. P. oligosporum, P. indonesiae).

131. Penicillium levitum Raper & Fennell, Mycologia 40: 511. 1948 (syn. P. rasile). 132. Penicillium limosum Ueda, Mycoscience 36: 451. 1995. 133. Penicillium lineolatum Udagawa & Horie, Mycotaxon 5: 493. 1977. 134. Penicillium ludwigii Udagawa, Trans. Mycol. Soc. Japan 10: 2. 1969. 135. Penicillium mariaecrucis Quintanilla, Avances Nutr. Mejora Anim. Aliment. 23: 334. 1982. 136. Penicillium meloforme Udagawa & Horie, Trans. Mycol. Soc. Japan 14: 376. 1973. 137. Penicillium ochrochloron Biourge, Cellule 33: 269. 1923. 138. Penicillium onobense Ramírez & Martínez, Mycopathol. 74: 44. 1981. 139. Penicillium oxalicum Currie & Thom, J. Biol. Chem. 22: 289. 1915. 140. Penicillium paraherquei Abe ex Smith, Trans. Br. Mycol. Soc. 46: 335. 1963. 141. Penicillium penarojense Houbraken, C. López-Q, Frisvad & Samson, Int. J. Syst. Evol. Microbiol. 61: 1471. 2011. 142. Penicillium piscarium Westling, Ark. Bot. 11: 86. 1911. 143. Penicillium pulvillorum Turfitt, Trans. Br. Mycol. Soc. 23: 186. 1939 (Syn. P. ciegleri). 144. Penicillium raperi Smith, Trans. Br. Mycol. Soc. 40: 486. 1957. 145. Penicillium reticulisporum Udagawa, Trans. Mycol. Soc. Japan 9: 52. 1968. 146. Penicillium rolfsii Thom, Penicillia: 489. 1930. 147. Penicillium simplicissimum (Oudemans) Thom, Penicillia: 335. 1930. 148. Penicillium skrjabinii Schmotina & Golovleva, Mikol. Fitopatol. 8: 530. 1974. 149. Penicillium svalbardense Frisvad, Sonjak & Gunde-Cimerman, Ant. van Leeuwenhoek 92: 48. 2007. 150. Penicillium vanderhammenii Houbraken, C. López-Q, Frisvad & Samson, Int. J. Syst. Evol. Microbiol. 61: 1473. 2011. 151. Penicillium vasconiae Ramírez & Martínez, Mycopathol. 72: 189. 1980. 152. Penicillium wotroi Houbraken, C. López-Q, Frisvad & Samson, Int. J. Syst. Evol. Microbiol. 61: 1474. 2011. 153. Penicillium zonatum Hodges & Perry, Mycologia 65: 697. 1973.

12: section Stolkia Houbraken & Samson, sect. nov. MycoBank

154. Penicillium boreae Peterson & Sigler, Mycol. Res. 106: 1112. 2002. 155. Penicillium canariense Peterson & Sigler, Mycol. Res. 106: 1113. 2002. 156. Penicillium donkii Stolk, Persoonia 7: 333. 1973. 157. Penicillium pullum Peterson & Sigler, Mycol. Res. 106: 1115. 2002. 158. Penicillium stolkiae Scott, Mycopathol. Mycol. Appl. 36: 8. 1968. 159. Penicillium subarcticum Peterson & Sigler, Mycol. Res. 106: 1116. 2002.

13: section Gracilenta Houbraken & Samson, sect. nov. MycoBank

160. Penicillium angustiporcatum Takada & Udagawa, Trans. Mycol. Soc. Japan 24: 143. 1983. 161. Penicillium estinogenum Komatsu & Abe ex Smith, Trans. Br. Mycol. Soc. 46: 335. 1963. 162. Penicillium macrosclerotiorum Wang, Zhang & Zhuang, Mycol. Res. 111: 1244. 2007. 163. Penicillium gracilentum Udagawa & Horie, Trans. Mycol. Soc. Japan 14: 373. 1973.

14: section Citrina Houbraken & Samson, sect. nov. MycoBank

164. Penicillium anatolicum Stolk, Ant. van Leeuwenhoek 34: 46. 1968. 165. Penicillium argentinense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 78. 2011. 166. Penicillium atrofulvum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 80. 2011. 167. Penicillium aurantiacobrunneum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 80. 2011. 168. Penicillium cairnsense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 83. 2011. 169. Penicillium christenseniae Houbraken, Frisvad & Samson, Stud. Mycol. 70: 85. 2011. 170. Penicillium chrzaszczii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 464. 1927. 171. Penicillium citrinum Thom, Bull. Bur. Anim. Ind. U.S. Dep. Agric. 118: 61. 1910. 172. Penicillium copticola Houbraken, Frisvad & Samson, Stud. Mycol. 70: 88. 2011. 173. Penicillium cosmopolitanum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 91. 2011. 174. Penicillium decaturense Peterson, Bayer & Wicklow, Mycologia 96: 1290. 2004. 175. Penicillium euglaucum van Beyma, Ant. van Leeuwenhoek 6: 269. 1940. 176. Penicillium galliacum Ramírez, Martínez & Berenguer, Mycopathol. 72: 30. 1980. 177. Penicillium godlewskii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 466. 1927. 178. Penicillium gorlenkoanum Baghdadi, Nov. Sist. Niz. Rast. 5: 97. 1968. 179. Penicillium hetheringtonii Houbraken, Frisvad & Samson, Fung. Div. 44: 125. 2010. 180. Penicillium manginii Duché & Heim, Trav. Cryptog. Louis L. Mangin: 450. 1931 (syn. P. pedemontanum, Houbraken et al. 2011b). 181. Penicillium miczynskii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 482. 1927. 182. Penicillium neomiczynskii Cole, Houbraken, Frisvad & Samson, Stud. Mycol. 70: 105. 2011. 183. Penicillium nothofagi Houbraken, Frisvad & Samson, Stud. Mycol. 70: 105. 2011. 184. Penicillium pancosmium Houbraken, Frisvad & Samson, Stud. Mycol. 70: 108. 2011. 185. Penicillium pasqualense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 108. 2011. 186. Penicillium paxilli Bainier, Bull. Soc. Mycol. France 23: 95. 1907. 187. Penicillium quebecense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 111. 2011. 188. Penicillium raphiae Houbraken, Frisvad & Samson, Stud. Mycol. 70: 114. 2011.

189. Penicillium roseopurpureum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901. 190. Penicillium sanguifluum (Sopp) Biourge, La Cellule 33: 105. 1923. 191. Penicillium shearii Stolk & Scott, Persoonia 4: 396. 1967. 192. Penicillium sizovae Baghdadi, Novosti Sist. Nizs. Rast. 1968: 103. 1968. 193. Penicillium steckii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 469. 1927. 194. Penicillium sumatrense Szilvinyi, Archiv. Hydrobiol. 14, Suppl. 6: 535. 1936. 195. Penicillium terrigenum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 125. 2011. 196. Penicillium tropicoides Houbraken, Frisvad & Samson, Fung. Div. 44: 127. 2010. 197. Penicillium tropicum Houbraken, Frisvad & Samson, Fung. Div. 44: 129. 2010. 198. Penicillium ubiquetum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 127. 2011. 199. Penicillium vancouverense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 131. 2011. 200. Penicillium waksmanii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 468. 1927. 201. Penicillium wellingtonense Cole, Houbraken, Frisvad & Samson, Stud. Mycol. 70: 133. 2011. 202. Penicillium westlingii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 473. 1927.

B. Penicillium subgenus Penicillium

15. Section Fasciculata Thom, The Penicillia: 374. 1930.

203. Penicillium albocoremium (Frisvad) Frisvad, Int. Mod. Tax. Meth. Pen. Asp. Clas.: 275. 2000. 204. Penicillium allii Vincent & Pitt, Mycologia 81: 300. 1989. 205. Penicillium aurantiogriseum Dierckx, Ann. Soc. Scient. Brux. 25: 88. 1901. 206. Penicillium camemberti Thom, Bull. Bur. Anim. Ind. USDA 82: 33. 1906. 207. Penicillium caseifulvum Lund, Filt. & Frisvad, J. Food Mycol. 1: 97. 1998. 208. Penicillium cavernicola Frisvad & Samson, Stud. Mycol. 49: 31. 2004. 209. Penicillium commune Thom, Bull. Bur. Anim. Ind. USDA 118: 56. 1910. 210. Penicillium crustosum Thom, Penicillia: 399. 1930. 211. Penicillium cyclopium Westling, Ark. Bot. 11: 90. 1911. 212. Penicillium discolor Frisvad & Samson, Ant. Van Leeuwenhoek, 72: 120. 1997. 213. Penicillium echinulatum Fassatiová, Acta Univ. Carol. Biol. 12: 326. 1977. 214. Penicillium freii Frisvad & Samson, Stud. Mycol. 49: 28. 2004. 215. Penicillium hirsutum Dierckx, Ann. Soc. Scient. Brux. 25: 89. 1901. 216. Penicillium hordei Stolk, Ant. van Leeuwenhoek 35: 270. 1969. 217. Penicillium melanoconidium (Frisvad) Frisvad & Samson, Stud. Mycol. 49: 28. 2004.

218. Penicillium neoechinulatum (Frisvad, Filt. & Wicklow) Frisvad & Samson, Stud. Mycol. 49: 28. 2004. 219. Penicillium nordicum Dragoni & Cantoni ex Ramírez, Adv. Pen. Asp. Syst.: 139. 1985. 220. Penicillium osmophilum Stolk & Veenbaas-Rijks, Ant. van Leeuwenhoek 40: 1. 1974. 221. Penicillium palitans Westling, Ark. Bot. 11: 83. 1911. 222. Penicillium polonicum Zaleski, Bull. Int. Acad. Pol. Sci. Lett., Sér. B 1927: 445. 1927. 223. Penicillium radicicola Overy & Frisvad, Syst. Appl. Microbiol.: 633. 2003. 224. Penicillium solitum Westling, Ark. Bot. 11: 65. 1911. 225. Penicillium thymicola Frisvad & Samson, Stud. Mycol. 49: 29. 2004. 226. Penicillium tricolor Frisvad, Seifert, Samson & Mills, Can. J. Bot. 72: 937. 1994. 227. Penicillium tulipae Overy & Frisvad, Syst. Appl. Microbiol. 634. 2003. 228. Penicillium venetum (Frisvad) Frisvad, Int. Mod. Tax. Meth. Pen. Asp. Clas.: 275. 2000. 229. Penicillium verrucosum Dierckx, Ann. Soc. Scient. Brux. 25: 88. 1901. 230. Penicillium viridicatum Westling, Ark. Bot. 11: 88. 1911.

16. Section Digitata Frisvad & Samson, Stud. Mycol. 49: 26. 2004.

231. Penicillium digitatum (Pers.:Fr.) Sacc., Fung. Ital.: 894. 1881.

17. Section Penicillium

232. Penicillium brevistipitatum Wang & Zhuang, Mycotaxon 93: 234. 2005. 233. Penicillium clavigerum Demelius, Verh. Zool.-Bot. Ges. Wien 72: 74. 1922. 234. Penicillium concentricum Samson, Stolk & Hadlok, Stud. Mycol. 11: 17. 1976. 235. Penicillium coprobium Frisvad, Mycologia 81: 853. 1989. 236. Penicillium coprophilum (Berk. & Curt.) Seifert & Samson, Adv.Pen. Asp. Syst.: 145. 1985. 237. Penicillium dipodomyicola (Frisvad, Filt. & Wicklow) Frisvad, Int. Mod. Meth. Pen. Asp. Clas.: 275. 2000. 238. Penicillium expansum Link, Ges. Naturf. Freunde Berlin Mag. Neuesten Entdeck. Gesammten Naturk. 3: 16. 1809. 239. Penicillium formosanum Hsieh, Su & Tzean, Trans. Mycol. Soc. R.O.C. 2: 159. 1987. 240. Penicillium gladioli McCulloch & Thom, Science, N.Y. 67: 217. 1928. 241. Penicillium glandicola (Oud.) Seifert & Samson, Adv. Pen. Asp. Syst.: 147. 1985. 242. Penicillium griseofulvum Dierckx, Ann. Soc. Scient. Brux. 25: 88. 1901. 243. Penicillium italicum Wehmer, Hedwigia 33: 211. 1894. 244. Penicillium marinum Frisvad & Samson, Stud. Mycol. 49: 20. 2004. 245. Penicillium sclerotigenum Yamamoto, Scient. Rep. Hyogo Univ. Agric., Agric. Biol. Ser. 2, 1: 69. 1955. 246. Penicillium ulaiense Hsieh, Su & Tzean, Trans. Mycol. Soc. R.O.C. 2: 161. 1987.

247. Penicillium vulpinum (Cooke & Massee) Seifert & Samson, Adv. Pen. Asp. Syst.: 144. 1985.

18. Section Roquefortorum (as “Roqueforti”) Frisvad & Samson, Stud. Mycol. 49: 16. 2004.

248. Penicillium carneum (Frisvad) Frisvad, Microbiology, UK, 142: 546. 1996. 249. Penicillium paneum Frisvad, Microbiology (UK) 142: 546. 1996. 250. Penicillium psychrosexualis Houbraken & Samson, IMA Fungus 1:174. 2010. 251. Penicillium roqueforti Thom, Bull. Bur. Anim. Ind. US Dept. Agric. 82: 35, 1906.

19. Section Chrysogena Frisvad & Samson, Stud. Mycol. 49: 17. 2004.

252. Penicillium aethiopicum Frisvad, Mycologia 81: 848. 1990. 253. Penicillium chrysogenum Thom, Bull. Bur. Anim. Ind. U.S. Dep. Agric. 118: 58. 1910. 254. Penicillium confertum (Frisvad et al.) Frisvad, Mycologia 81: 852. 1990. 255. Penicillium dipodomyis (Frisvad, Filtenborg & Wicklow) Banke, Frisvad & Rosendahl, Int. Mod. Meth. Pen. Asp. Clas., 270. 2000. 256. Penicillium egyptiacum van Beyma, Zentralbl. Bakteriol., 2. Abt., 88: 137. 1933. (syn. P. nilense). 257. Penicillium flavigenum Frisvad & Samson, Mycol. Res. 101: 620. 1997. 258. Penicillium kewense Smith, Trans. Br. Mycol. Soc. 44: 42. 1961 (syn. E. crustaceum). 259. Penicillium molle Pitt, The Genus Penicillium: 148, 1980 [“1979”]. 260. Penicillium mononematosum (Frisvad et al.) Frisvad, Mycologia 81:857. 1990. 261. Penicillium nalgiovense Laxa, Zentralbl. Bakteriol., 2. Abt., 86: 160. 1932. 262. Penicillium persicinum Wang, Zhou, Frisvad & Samson, Ant. van Leeuwenhoek 86: 177. 2004. 263. Penicillium rubens Biourge, Cellule 33: 265. 1923. 264. Penicillium sinaicum Udagawa & Ueda, Mycotaxon 14: 266. 1982.

20: section Turbata Houbraken & Samson, sect. nov. MycoBank .

265. Penicillium bovifimosum (Tuthill & Frisvad) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 (this study). 266. Penicillium matriti Smith, Trans. Br. Mycol. Soc. 44: 44. 1961. 267. Penicillium turbatum Westling, Ark. Bot. 11: 128. 1911 (syn. E. baarnense, P. baarnense, this study).

21: section Paradoxa Houbraken & Samson, sect. nov. MycoBank

268. Penicillium atramentosum Thom, Bull. Bur. Anim. Ind. US Dept. Agric. 118: 65. 1910. 269. Aspergillus crystallinus Kwon-Chung & Fennell, The Genus Aspergillus: 471. 1965. 270. Aspergillus malodoratus (Kwon-Chung & Fennell), The Genus Aspergillus: 468. 1965. 271. Aspergillus paradoxus Fennell & Raper, Mycologia 47: 69.

22: section Brevicompacta Thom, The Penicillia: 289. 1930.

272. Penicillium astrolobatum Serra & Peterson, Mycologia 99: 80. 2007. 273. Penicillium bialowiezense Zaleski, Bull. Int. Acad. Pol. Sci. Lett., Sér. B, 1927: 462. 1927 (syn. P. biourgeianum). 274. Penicillium brevicompactum Dierckx, Ann. Soc. Scient. Brux. 25: 88. 1901. 275. Penicillium fennelliae Stolk, Ant. van Leeuwenhoek 35: 261. 1969. 276. Penicillium neocrassum Serra & Peterson, Mycologia 99: 81. 2007. 277. Penicillium olsonii Bainier & Sartory, Ann. Mycol. 10: 398. 1912. 278. Penicillium tularense Paden, Mycopathol. Mycol. Appl. 43: 264. 1971.

23: section Ramosa (as “Ramosum”) Stolk & Samson, Adv. Pen. Asp. Syst.: 179. 1985.

279. Penicillium jamesonlandense Frisvad & Overy, Int. J. Syst. Evol. Microbiol. 56: 1435. 2006. 280. Penicillium kojigenum Smith, Trans. Br. Mycol. Soc. 44: 43. 1961. 281. Penicillium lanosum Westling, Ark. Bot. 11: 97. 1911. 282. Penicillium raistrickii Smith, Trans. Br. Mycol. Soc.18: 90. 1933. 283. Penicillium ribeum Frisvad & Overy, Int. J. Syst. Evol. Microbiol. 56: 1436. 2006. 284. Penicillium sajarovii Quintanilla, Avances Nutr. Mejora Anim. Aliment. 22: 539. 1981. 285. Penicillium scabrosum Frisvad, Samson & Stolk, Persoonia 14: 177. 1990. 286. Penicillium simile Davolos, Pietrangeli, Persiani & Maggi, J. Syst. Evol. Microbiol., in press. 287. Penicillium soppii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 476. 1927. 288. Penicillium swiecickii Zaleski, Bull. Int. Acad. Pol. Sci. Lett., Sér. B 1927: 474. 1927. 289. Penicillium virgatum Nirenberg & Kwasna, Mycol. Res. 109: 977. 2005.

24: section Canescentia Houbraken & Samson, sect. nov. MycoBank MB563135.

290. Penicillium canescens Sopp, Skr. Vidensk.-Selsk. Christiana, Math.-Naturvidensk. Kl. 11: 181. 1912. 291. Penicillium jensenii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 494. 1927. 292. Penicillium yarmokense Baghdadi, Nov. Sist. Niz. Rast. 5: 99. 1968.

293. Penicillium janczewskii Zaleski, Bull. Int. Acad. Polon. Sci., Cl. Sci. Math., Sér. B, Sci. Nat., 1927: 488. 1927. 294. Penicillium antarcticum Hocking & McRae, Polar Biology 21: 103. 1999. 295. Penicillium atrovenetum Smith, Trans. Br. Mycol. Soc. 39: 112. 1956. 296. Penicillium novae-zeelandiae van Beyma, Ant. van Leeuwenhoek 6: 275. 1940. 297. Penicillium coralligerum Nicot & Pionnat, Bull. Soc. Mycol. France 78: 245. 1963 [“1962”].

25: section Eladia (Smith) Stolk & Samson, Adv. Pen. Asp. Syst.: 169. 1985.

298. Penicillium sacculum Dale apud Biourge, Cellule 33: 323. 1923. 299. Penicillium senticosum Scott, Mycopathol. Mycol. Appl. 36: 5. 1968.

3. 4. Updated Classification

This is the sectional classification published by Visagie et al. (2014), arranged in the two subgenera published by Houbraken & Samson (2011)

1. Penicillium subgenus : Aspergilloides

1. Section Aspergilloides

1. Penicillium ardesiacum Novobr., Novosti Sist. Nizsh. Rast. 11: 228. 1974. 2. Penicillium armarii Houbraken, et al. Stud. Mycol. 78: 410. 2014 3. Penicillium athertonense Houbraken, Stud. Mycol. 78: 412. 2014 4. Penicillium aurantioviolaceum Biourge, Cellule 33: 282. 1923. 5. Penicillium austroafricanum Houbraken & Visagie, Stud. Mycol. 78: 412. 2014. 6. Penicillium brunneoconidiatum Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 415. 2014. 7. Penicillium bussumense Houbraken, Stud. Mycol. 78: 415. 2014 8. Penicillium cartierense Houbraken, Stud. Mycol. 78: 415. 2014. 9. Penicillium clavistipitatum Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 419. 2014. 10. Penicillium contaminatum Houbraken, Stud. Mycol. 78: 419. 2014 11. Penicillium crocicola W. Yamam., Sci. Rep. Hyogo Univ. Agric. 2: 28. 1956. 12. Penicillium flavisclerotiatum Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 419. 2014. 13. Penicillium frequentans Westling, Ark. Bot. 11: 133. 1911. 14. Penicillium fuscum (Sopp) Biourge, Cellule 33: 103. 1923 15. Penicillium fusisporum L. Wang, PloS ONE 9: e101454-P2. 2014. 16. Penicillium glabrum (Wehmer) Westling, Ark. Bot. 11: 131. 1911 17. Penicillium grancanariae C. Ramírez, A.T. Martínez & Ferrer, Mycopathologia 66: 79. 1978. 18. Penicillium grevilleicola Houbraken & Quaedvlieg, Stud. Mycol. 78: 423. 2014. 19. Penicillium hoeksii Houbraken, Stud. Mycol. 78: 423. 2014 20. Penicillium infra-aurantiacum Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 426. 2014. 21. Penicillium kananaskense Seifert, Frisvad & McLean, Can. J. Bot. 72: 20. 1994. 22. Penicillium kiamaense Houbraken & Pitt, Stud. Mycol. 78: 426. 2014 23. Penicillium lividum Westling, Ark. Bot. 11: 134. 1911 24. Penicillium longicatenatum Visagie, et al., Stud. Mycol. 78: 429. 2014 25. Penicillium malmesburiense Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 429. 2014. 26. Penicillium montanense M. Chr. & Backus, Mycologia 54: 574. 1962 27. Penicillium odoratum M. Chr. & Backus, Mycologia 53: 459. 1961 28. Penicillium palmense C. Ramírez & A.T. Martínez, Mycopathologia 66: 80. 1978. 29. Penicillium patens Pitt & A.D. Hocking, Mycotaxon 22: 205. 1985 30. Penicillium pulvis Houbraken, Visagie, Samson & Seifert, Stud. Mycol. 78: 429. 2014. 31. Penicillium purpurescens (Sopp) Raper & Thom, A manual of the Penicillia: 177. 1949 2. 32. Penicillium quercetorum Baghd., Novosti Sist. Nizsh. Rast. 5: 110. 1968 33. Penicillium ranomafanaense Houbraken & Hagen, Stud. Mycol. 78: 433. 2014 34. Penicillium roseomaculatum Biourge, Cellule 33: 301. 1923 35. Penicillium roseoviride Stapp & Bortels, Zentralbl. Bakteriol. Parasitenk., Abt. 2 93: 51. 1935. 36. Penicillium rudallense Houbraken, Visagie & Pitt, Stud. Mycol. 78: 433. 2014

37. Penicillium saturniforme (L. Wang & W.Y. Zhuang) Houbraken & Samson, Stud. Mycol. 70: 48. 2011 . 38. Penicillium spinulosum Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 76. 1910. 39. Penicillium sterculiniicola Houbraken, Stud. Mycol. 78: 436. 2014 40. Penicillium sublectaticum Houbraken, et al., Stud. Mycol. 78: 436. 2014 41. Penicillium subspinulosum Houbraken, Stud. Mycol. 78: 436. 2014. 42. Penicillium thiersii S.W. Peterson, E.M. Bayer & Wicklow, Mycologia 96: 1283. 2004. 43. Penicillium thomii Maire, Bull. Soc. Hist. Nat. Afrique N. 8: 189. 44. Penicillium trzebinskii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 498. 1927. 45. Penicillium tsitsikammaense Houbraken, Stud. Mycol. 78: 440. 2014 46. Penicillium turcosoconidiatum Visagie, Houbraken & K. Jacobs, Stud. Mycol. 78: 440. 2014. 47. Penicillium vagum Houbraken, et al., Stud. Mycol. 78: 443. 2014 48. Penicillium valentinum C. Ramírez and A.T. Martínez, Mycopathologia 72: 183. 1980. 49. Penicillium verhagenii Houbraken, Stud. Mycol. 78: 443. 2014. 50. Penicillium yezoense Hanzawa ex Houbraken, Stud. Mycol. 78: 443. 2014 51. Penicillium zhuangii L. Wang, PloS ONE 9: e101454-P4. 2014

2. SectionCharlesia

52. Penicillium charlesii G. Sm., Trans. Brit. Mycol. Soc. 18: 90. 1933 53. Penicillium coffeae S.W. Peterson et al., Mycologia 97: 662. 2005. 54. Penicillium fellutanum Biourge, Cellule 33: 262. 1923. 55. Penicillium georgiense S.W. Peterson & B.W. Horn, Mycologia 101: 79. 2009. 56. Penicillium indicum D.K. Sandhu & R.S. Sandhu, Can. J. Bot. 41: 1273. 1963 57. Penicillium multicolor Grig.-Man. & Porad., Arch. des Sciences Biol. Leningrad 19: 120. 1915. 58. Penicillium phoeniceum J.F.H. Beyma, Zentralbl. Bakteriol. Parasitenk., Abt. 2 88: 136. 1933

3..Section Cinnamopurpurea

59. Penicillium chermesinum Biourge, Cellule 33: 284. 1923. 60. Penicillium cinnamopurpureum Udagawa, J. Agric. Food Sci., Tokyo 5: 1. 1959. 61. Penicillium ellipsoideosporum L. Wang & W.Y. Kong, Mycosystema 19: 463. 2000. 62. Penicillium idahoense Paden, Mycopathol. Mycol. Appl. 43: 259. 1971 63. Penicillium incoloratum L.Q. Huang & Z.T. Qi, Acta Mycol. Sin. 13: 264. 1994. 64. Penicillium infrapurpureum Visagie, Seifert & Samson, Stud. Mycol. 78: 116. 2014. 65. Penicillium jiangxiense H.Z. Kong & Z.Q. Liang, Mycosystema 22: 4. 2003 66. Penicillium malacaense C. Ramírez & A.T. Martínez, Mycopathologia 72: 186. 1980. 67. Penicillium nodulum H.Z. Kong & Z.T. Qi, Mycosystema 1: 108. 1988. 68. Penicillium parvulum S.W. Peterson & B.W. Horn, Mycologia 101: 75. 2009 69. Penicillium shennangjianum H.Z. Kong & Z.T. Qi, Mycosystema 1: 110. 1988.

4. Section Citrina

70. Penicillium anatolicum Stolk, Antonie van Leenwenhoek 34: 46. 1968 71. Penicillium argentinense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 78. 2011. 72. Penicillium atrofulvum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 80. 2011. 73. Penicillium aurantiacobrunneum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 80. 2011. 74. Penicillium cairnsense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 83. 2011. 75. Penicillium christenseniae Houbraken, Frisvad & Samson, Stud. Mycol. 70: 85. 2011.

76. Penicillium chrzaszczii K.M. Zalessky, Bull. Int. Acad. Polon. Sci 77. Penicillium citrinum Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 61. 1910. 78. Penicillium copticola Houbraken, Frisvad & Samson, Stud. Mycol. 70: 88. 2011. 79. Penicillium cosmopolitanum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 91. 2011. 80. Penicillium decaturense S.W. Peterson, E.M. Bayer & Wicklow, Mycologia 96: 1290. 2004. 81. Penicillium euglaucum J.F.H. Beyma, Antonie van Leeuewenhoek 6: 269. 1940 82. Penicillium galliacum C. Ramírez, A.T. Martínez & Berer., Mycopathologia 72: 30. 1980. 83. Penicillium godlewskii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 466. 1927. 84. Penicillium gorlenkoanum Baghd., Novosti Sist. Nizsh. Rast. 5: 97. 1968. 85. Penicillium hetheringtonii Houbraken, Frisvad & Samson, Fungal Divers. 44: 125. 2010. 86. Penicillium manginii Duché & R. Heim, Trav. Cryptog.: 450. 1931 87. Penicillium miczynskii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 482. 1927. 88. Penicillium neomiczynskii A.L.J. Cole et al., Stud. Mycol. 70: 105. 2011 89. Penicillium nothofagi Houbraken, Frisvad & Samson, Stud. Mycol. 70: 105. 2011. 90. Penicillium pancosmium Houbraken, Frisvad & Samson, Stud. Mycol. 70: 108. 2011. 91. Penicillium pasqualense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 108. 2011. 92. Penicillium paxilli Bainier, Bull. Soc. Mycol. France 23: 95. 1907 93. Penicillium quebecense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 111. 2011. 94. Penicillium raphiae Houbraken, Frisvad & Samson, Stud. Mycol. 70: 114. 2011. 95. Penicillium roseopurpureum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901. 96. Penicillium sanguifluum (Sopp) Biourge, Cellule 33: 105. 97. Penicillium shearii Stolk & D.B. Scott, Persoonia 4: 396. 1967 98. Penicillium sizovae Baghd., Novosti Sist. Nizsh. Rast. 1968: 103. 1968 99. Penicillium steckii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 469. 1927. 100. Penicillium sucrivorum Visagie & K. Jacobs, Mycologia 106: 546. 2014 101. Penicillium sumatraense Szilvinyi [as ‘sumatrense’], Archiv. Hydrobiol. 14, Suppl. 6: 535. 1936. 102. Penicillium terrigenum Seifert et al., Stud. Mycol. 70: 125. 2011 103. Penicillium tropicoides Houbraken, Frisvad & Samson, Fungal Divers. 44: 127. 2010. 104. Penicillium tropicum Houbraken, Frisvad & Samson, Fungal Divers. 44: 129. 2010 105. Penicillium ubiquetum Houbraken, Frisvad & Samson, Stud. Mycol. 70: 127. 2011. 106. Penicillium vancouverense Houbraken, Frisvad & Samson, Stud. Mycol. 70: 131. 2011. 107. Penicillium waksmanii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat.: 468. 1927. 108. Penicillium wellingtonense A.L.J. Cole et al., Stud. Mycol. 70: 133. 2011. 109. Penicillium westlingii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 473. 1927.

5. Section Exilicaulis

110. Penicillium alutaceum D.B. Scott, Mycopathol. Mycol. Appl. 36: 17. 111. Penicillium atrosanguineum B.X. Dong, Ceská Mycol. 27: 174. 1973 112. Penicillium burgense Quintan., Av. Aliment. Majora Anim. 30: 176. 1990 113. Penicillium catenatum D.B. Scott, Mycopathol. Mycol. Appl. 36: 24. 1968 114. Penicillium chalybeum Pitt & A.D. Hocking, Mycotaxon 22: 204. 1985. 115. Penicillium cinerascens Biourge, Cellule 33: 308. 1923. 116. Penicillium cinereoatrum Chalab., Bot. Mater. Otd. Sporov. Rast. 6: 167. 1950. 117. Penicillium citreonigrum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901 118. Penicillium corylophilum Dierckx, Ann. Soc. Sci. Bruxelles 25: 86. 1901. 119. Penicillium decumbens Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 71. 1910.

120. Penicillium dimorphosporum H.J. Swart, Trans. Brit. Mycol. Soc. 55: 310. 1970 121. Penicillium dravuni Janso, Mycologia 97: 445. 2005. 122. Penicillium erubescens D.B. Scott, Mycopathol. Mycol. Appl. 36: 14. 1968 123. Penicillium fagi C. Ramírez & A.T. Martínez, Mycopathologia 63: 57. 1978 124. Penicillium flavidostipitatum C. Ramírez & C.C. González, Mycopathologia 88: 3. 1984. 125. Penicillium guttulosum J.C. Gilman & E.V. Abbott, Iowa St. Coll. J. Sci. 1: 298. 1927. 126. Penicillium heteromorphum H.Z. Kong & Z.T. Qi, Mycosystema 1: 107. 1988. 127. Penicillium katangense Stolk, Antonie van Leeuwenhoek 34: 42. 1968 128. Penicillium laeve (K. Ando & Manoch) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 129. Penicillium lapidosum Raper & Fennell, Mycologia 40: 524. 1948 130. Penicillium maclennaniae H.Y. Yip, Trans. Brit. Mycol. Soc. 77: 202. 1981. 131. Penicillium melinii Thom, Penicillia: 273. 1930 132. Penicillium menonorum S.W. Peterson, IMA Fungus 2: 122. 2011 133. Penicillium meridianum D.B. Scott, Mycopathol. Mycol. Appl. 36: 12. 1968 134. Penicillium namyslowskii K.M. Zalessky, Bull. Int. Aead. Polonc. Sci., 1927 135. Penicillium nepalense Takada & Udagawa, Trans. Mycol. Soc. Japan 24: 146. 1983 136. Penicillium ovatum (K. Ando & Nawawi) Houbraken & Samson, Stud. Mycol. 70: 48. 2011 137. Penicillium parvum Raper & Fennell, Mycologia 40: 508. 1948 138. Penicillium philippinense Udagawa & Y. Horie, J. Jap. Bot. 47: 341. 1972 139. Penicillium pimiteouiense S.W. Peterson, Mycologia 91: 271. 1999 140. Penicillium raciborskii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., 1927: 454. 1927. 141. Penicillium restrictum J.C. Gilman & E.V. Abbott, Iowa St. Coll. J. Sci. 1: 297. 1927. 142. Penicillium rubefaciens Quintan., Mycopathologia 80: 73. 1982. 143. Penicillium rubidurum Udagawa & Y. Horie, Trans. Mycol. Soc. Japan 14: 381. 1973 144. Penicillium smithii Quintan., Av. Aliment. Majora Anim. 23: 340. 1982 145. Penicillium striatisporum Stolk, Antonie van Leeuwenhoek 35: 268. 1969 146. Penicillium terrenum D.B. Scott, Mycopathol. Mycol. Appl. 36: 1. 1968 147. Penicillium velutinum J.F.H. Beyma, Zentralbl. Bakteriol. Parasitenk., Abt. 2 91: 353. 1935. 148. Penicillium vinaceum J.C. Gilman & E.V. Abbott, Iowa St. Coll. J. Sci. 1: 299. 1927.

6. Section Fracta

149. Penicillium fractum Udagawa, Trans. Mycol. Soc. Japan 9: 51. 1968 ≡ Eupenicillium fractumUdagawa, Trans. Mycol. Soc. Japan 9: 51. 1968. 150. Penicillium inusitatum D.B. Scott, Mycopathol. Mycol. Appl. 36: 20. 1968 ≡ Eupenicillium inusitatumD.B. Scott, Mycopathol. Mycol. Appl. 36: 20. 1968.

7. Section Gracilenta

151. Penicillium angustiporcatum Takada & Udagawa, 1983 152. Penicillium estinogenum A. Komatsu & S. Abe ex G. Sm., Trans. Brit. Mycol. Soc. 46: 335. 1963 153. Penicillium gracilentum Udagawa & Y. Horie, Trans. Mycol. Soc. Japan 14: 373. 1973 . 154. Penicillium macrosclerotiorum L. Wang, X.M. Zhang & W.Y. Zhuang, Mycol. Res. 111: 1244. 2007.\

8. Section Lanata-Divaricata

155. Penicillium abidjanum Stolk, Antonie van Leeuwenhoek 34: 49. 1968 156. Penicillium araracuaraense Houbraken, et al., Int. J. Syst. Evol. Microbiol. 61: 1469. 2011. 157. Penicillium brasilianum Bat., Anais Soc. Biol. Pernambuco 15: 162. 1957 158. Penicillium brefeldianum B.O. Dodge, Mycologia 25: 92. 1933 159. Penicillium caperatum Udagawa & Y. Horie, Trans. Mycol. Soc. Japan 14: 371. 1973 160. Penicillium cluniae Quintan., Av. Aliment. Mejora Anim. 30: 174. 1990 161. Penicillium coeruleum Sopp apud Biourge, Cellule 33: 102. 1923. 162. Penicillium cremeogriseum Chalab., Bot. Mater. Otd. Sporov. Rast. 6: 168. 1950. 163. Penicillium daleae K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 495. 1927. 164. Penicillium ehrlichii Kleb., Ber. Deutsch. Bot. Ges. 48: 374. 1930 165. Penicillium elleniae Houbraken et al., Int. J. Syst. Evol. Microbiol. 61: 1470. 2011. 166. Penicillium glaucoroseum Demelius, Verh. Zool.-Bot. Ges. Wien 72: 72. 1923 (1922). 167. Penicillium griseopurpureum G. Sm., Trans. Brit. Mycol. Soc. 48: 275. 1965. 168. Penicillium janthinellum Biourge, Cellule 33: 258. 1923. 169. Penicillium javanicum J.F.H. Beyma, Verh. Kon. Ned. Akad. Wetensch., Afd. Natuurk. 26: 17. 1929 170. Penicillium levitum Raper & Fennell, Mycologia 40: 511. 171. Penicillium limosum S. Ueda, Mycoscience 36: 451. 1995 172. Penicillium lineolatum Udagawa & Y. Horie, Mycotaxon 5: 493. 1977 173. Penicillium ludwigii Udagawa, Trans. Mycol. Soc. Japan 10: 2. 1969 174. Penicillium mariae-crucis Quintan., Av. Aliment. Majora Anim. 23: 334. 1982. 175. Penicillium meloforme Udagawa & Y. Horie, Trans. Mycol. Soc. Japan 14: 376. 1973 176. Penicillium ochrochloron Biourge, Cellule 33: 269. 1923. 177. Penicillium onobense C. Ramírez & A.T. Martínez, Mycopathologia 74: 44. 1981. 178. Penicillium oxalicum Currie & Thom, J. Biol. Chem. 22: 289. 1915 179. Penicillium paraherquei S. Abe ex G. Sm., Trans. Brit. Mycol. Soc. 46: 335. 1963 180. Penicillium penarojense Houbraken et al., Int. J. Syst. Evol. Microbiol. 61: 1471. 2011. 181. Penicillium piscarium Westling, Ark. Bot. 11: 86. 1911 182. Penicillium pulvillorum Turfitt, Trans. Brit. Mycol. Soc. 23: 186. 1939 183. Penicillium raperi G. Sm., Trans. Brit. Mycol. Soc. 40: 486. 1957 184. Penicillium reticulisporum Udagawa, Trans. Mycol. Soc. Japan 9: 52. 1968 185. Penicillium rolfsii Thom, Penicillia: 489. 1930 186. Penicillium simplicissimum (Oudem.) Thom, Penicillia: 335. 1930 187. Penicillium singorense Visagie, Seifert & Samson, Stud. Mycol. 78: 119. 2014. 188. Penicillium skrjabinii Schmotina & Golovleva, Mikol. Fitopatol. 8: 530. 1974. 189. Penicillium subrubescens Houbraken et al., Antonie van Leeuwenhoek 103: 1354. 2013. 190. Penicillium svalbardense Frisvad, Sonjak & Gundae-Cim, Antonie van Leeuwenhoek 92: 48. 2007. 191. Penicillium vanderhammenii Houbraken et al., Int. J. Syst. Evol. Microbiol. 61: 1473. 2011. 192. Penicillium vasconiae C. Ramírez & A.T. Martínez, Mycopathologia 72: 189. 1980 193. Penicillium wotroi Houbraken et al., Int. J. Syst. Evol. Microbiol. 61: 1474. 2011. 194. Penicillium zonatum Hodges & J.J. Perry, Mycologia 65: 697. 1973 ≡ Eupenicillium zonatum Hodges & J.J. Perry, Mycologia 65: 697. 1973.

9. Section Ochrosalmonea

195. Penicillium isariiforme Stolk & J.A. Mey., Trans. Brit. Mycol. Soc. 40: 187. 1957. 196. Penicillium ochrosalmoneum Udagawa, J. Agric. Sci. Tokyo Nogyo Daig. 5: 10. 1959

10. Section Ramigera

197. Penicillium capsulatum Raper & Fennell, Mycologia 40: 528. 1948 198. Penicillium cyaneum (Bainier & Sartory) Biourge, Cellule 33: 102. 1923 199. Penicillium dierckxii Biourge, Cellule 33: 313. 1923. 200. Penicillium hispanicum C. Ramírez, A.T. Martínez & Ferrer, Mycopathologia 66: 77. 1978. 201. Penicillium ornatum Udagawa, Trans. Mycol. Soc. Japan 9: 49. 1968 202. Penicillium ramusculum Bat. & H. Maia, Anais Soc. Biol. Pernambuco 13: 27. 1955. 203. Penicillium sublateritium Biourge, Cellule 33: 315. 1923.

11. Section Sclerotiora

204. Penicillium adametzii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat., 1927: 205. Penicillium adametzioides S. Abe ex G. Sm., Trans. Brit. Mycol. Soc. 46: 335. 206. Penicillium alexiae Visagie Houbraken & Samson, Persoonia 31: 59. 2013. 207. Penicillium amaliae Visagie, Houbraken & K. Jacobs, 208. Penicillium angulare S.W. Peterson, E.M. Bayer & Wicklow, 209. Penicillium arianeae Visagie, Houbraken & Samson, Persoonia 31: 59. 2013. 210. Penicillium bilaiae Chalab., Bot. Mater. Otd. Sporov. Rast. 6: 165. 1950. 211. Penicillium brocae S.W. Peterson et al., Mycologia 95: 143. 2003. 212. Penicillium cainii K.G. Rivera, Malloch & Seifert, Stud. Mycol. 70: 147. 2011 213. Penicillium daejeonium S.H. Yu & H.K. Sang, J. Microbiol. 51: 537. 2013. 214. Penicillium guanacastense K.G. Rivera, Urb & Seifert, Mycotaxon 119: 324. 2011 215. Penicillium herquei Bainier & Sartory, Bull. Soc. Mycol. France 28: 121. 1912. 216. Penicillium hirayamae Udagawa, J. Agric. Soc. Tokyo 5: 6. 1959 217. Penicillium jacksonii K.G. Rivera, Houbraken & Seifert, Stud. Mycol. 70: 151. 2011. 218. Penicillium johnkrugii K.G. Rivera, Houbraken & Seifert, Stud. Mycol. 70: 151. 2011. 219. Penicillium jugoslavicum C. Ramírez & Munt.-Cvetk., Mycopathologia 88: 65. 1984. 220. Penicillium lilacinoechinulatum S. Abe ex G. Sm., Trans. Brit. Mycol. Soc. 46: 335. 1963 221. Penicillium malachiteum (Yaguchi & Udagawa) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 222. Penicillium mallochii K.G. Rivera, Urb & Seifert, Mycotaxon 119: 322. 2012 223. Penicillium maximae Visagie, Houbraken & Samson, Persoonia 31: 52. 2013 224. Penicillium restingae J.P. Andrade et al., Persoonia 32: 293. 2014 225. Penicillium sclerotiorum J.F.H. Beyma, Zentralbl. Bakteriol. Parasitenk., Abt. 2 96: 418. 1937. 226. Penicillium vanoranjei Visagie, Houbraken & Samson, Persoonia 31: 46. 2013. 227. Penicillium viticola Nonaka & Masuma, Mycoscience 52: 339. 2011

12. Section Stolkia

228. Penicillium boreae S.W. Peterson & Sigler, Mycol. Res. 106: 1112. 2002 229. Penicillium canariense S.W. Peterson & Sigler, Mycol. Res. 106: 1113. 2002. 230. Penicillium donkii Stolk, Persoonia 7: 333. 1973. 231. Penicillium pullum S.W. Peterson & Sigler, Mycol. Res. 106: 1115. 2002. 232. Penicillium stolkiae D.B. Scott, Mycopathol. Mycol. Appl. 36: 8. 1968 233. Penicillium subarcticum S.W. Peterson & Sigler, Mycol. Res. 106: 1116. 2002.

13. Section Thysanophora

234. Penicillium hennebertii Houbraken & Samson, Stud. Mycol. 70: 47. 2011 235. Penicillium longisporum (W.B. Kend.) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 236. Penicillium taxi R. Schneid., Zentralbl. Bakteriol. Parasitenk., Abt. 2 110: 43. 1956

14.Section Torulomyces

237. Penicillium cryptum Goch., Mycotaxon 26: 349. 1986 ≡ Eupenicillium cryptum Goch., Mycotaxon 26: 349. 1986 238. Penicillium lagena (Delitsch) Stolk & Samson, Stud. Mycol. 23: 100. 1983 239. Penicillium lassenii Paden, Mycopathol. Mycol. Appl. 43: 266. 240. Penicillium porphyreum Houbraken & Samson, Stud. Mycol. 70: 48. 2011

2. Penicillium subgenus Penicillium

15. Section Brevicompacta

241. Penicillium astrolabium R. Serra & S.W. Peterson, Mycologia 99: 80. 2007 242. Penicillium bialowiezense K.M. Zalessky, Bull. Int. Acad. Polon. Sci., 243. Penicillium brevicompactum Dierckx, Ann. Soc. Sci. Bruxelles 25: 88. 1901 244. Penicillium buchwaldii Frisvad & Samson, FEMS Microbiol. Lett. 339: 86. 2013. 245. Penicillium fennelliae Stolk, Antonie van Leeuwenhoek 35: 261. 1969. 246. Penicillium kongii L. Wang, Mycologia 105: 1549. 2013 247. Penicillium neocrassum R. Serra & S.W. Peterson, Mycologia 99: 81. 2007. 248. Penicillium olsonii Bainier & Sartory, Ann. Mycol. 10: 398. 1912. 249. Penicillium spathulatum Frisvad & Samson, FEMS Microbiol. Lett. 339: 88. 2013 250. Penicillium tularense Paden, Mycopathol. Mycol. Appl. 43: 264. 1971

16. Section Canescentia

251. Penicillium antarcticum A.D. Hocking & C.F. McRae, Polar Biol. 21: 103. 1999 252. Penicillium atrovenetum G. Sm., Trans. Brit. Mycol. Soc. 39: 112. 1956. 253. Penicillium canescens Sopp, Skr. Vidensk.-Selsk. Christiana Math.- 11: 181. 1912. 254. Penicillium canis S.W. Peterson, J. Clin. Microbiol. (in press) 255. Penicillium coralligerum Nicot & Pionnat, Bull. Soc. Mycol. 78: 245. 1963 [1962]. 256. Penicillium dunedinense Visagie, Seifert & Samson, Stud. Mycol. 78: 121. 2014. 257. Penicillium janczewskii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., 258. Penicillium jensenii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., 1927. 259. Penicillium novae-zeelandiae J.F.H. Beyma, Antonie van Leeuwenhoek 6: 275. 1940. 260. Penicillium yarmokense Baghd., Novosti Sist. Nizsh. Rast. 5: 99. 1968

17. Section Chrysogena

261. Penicillium allii-sativi Frisvad, Houbraken & Samson, Persoonia 29: 89. 2012. 262. Penicillium chrysogenum Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 58. 1910. 263. Penicillium confertum (Frisvad, Filt. & Wicklow) Frisvad, Mycologia 81: 852. 1990

264. Penicillium desertorum Frisvad, Houbraken & Samson, Persoonia 29: 90. 2012. 265. Penicillium dipodomyis (Frisvad, Filt. & Wicklow) Banke, Firsvad & S. Rosend, Int. Mod. Meth. Pen. Asp. Clas. 270. 2000 266. Penicillium egyptiacum J.F.H. Beyma, Zentralbl. Bakteriol. Parasitenk., Abt. 2 88: 137. 1933 267. Penicillium flavigenum Frisvad & Samson, Mycol. Res. 101: 620. 1997 268. Penicillium glycyrrhizacola A.J. Chen, B.D. Sun & W.W. Gao, PloS ONE 8: e78285-P3. 2013. 269. Penicillium goetzii J. Rogers et al., Persoonia 29: 92. 2012. 270. Penicillium halotolerans Frisvad, Houbraken & Samson, Persoonia 29: 92. 2012. 271. Penicillium kewense G. Sm., Trans. Brit. Mycol. Soc. 44: 42. 1961. 272. Penicillium lanosocoeruleum Thom, Penicillia: 322. 1930 273. Penicillium mononematosum (Frisvad, Filt. & Wicklow) Frisvad, Mycologia 81: 857. 1990 274. Penicillium nalgiovense Laxa, Zentralbl. Bakteriol. Parasitenk., Abt. 2 86: 160. 1932 275. Penicillium persicinum L. Wang et al., Antonie van Leeuwenhoek 86: 177. 2004 276. Penicillium rubens Biourge, Cellule 33: 265. 1923. 277. Penicillium sinaicum Udagawa & S. Ueda, Mycotaxon 14: 266. 1982 278. Penicillium tardochrysogenum Frisvad, Houbraken & Samson, Persoonia 29: 93. 2012. 279. Penicillium vanluykii Frisvad, Houbraken & Samson, Persoonia 29: 97. 2012

18. Section Digitata

280. Penicillium digitatum (Pers.: Fr.) Sacc., Fung. Ital.: tab. 894. 1881

19. Section Eladia

281. Penicillium sacculum E. Dale, Ann. Mycol. 24: 137. 1926 282. Penicillium senticosum D.B. Scott, Mycopathol. Mycol. Appl. 36: 5. 1968

20. Section Fasciculata.

283. Penicillium albocoremium (Frisvad) Frisvad, Int. Mod. Tax. Meth. Pen. Asp. Clas.: 275. 2000 284. Penicillium allii Vincent & Pitt, Mycologia 81: 300. 1989 285. Penicillium aurantiogriseum Dierckx, Ann. Soc. Sci. Bruxelles 25: 88. 1901. 286. Penicillium biforme Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 54. 1910 287. Penicillium camemberti Thom, U.S.D.A. Bur. Animal Industr. Bull. 82: 33. 1906. 288. Penicillium caseifulvum Lund, Filt. & Frisvad, J. Food Mycol. 1: 97. 1998. 289. Penicillium cavernicola Frisvad & Samson, Stud. Mycol. 49: 31. 2004. 290. Penicillium commune Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 56. 1910. 291. Penicillium crustosum Thom, The Penicillia: 399. 1930 292. Penicillium cyclopium Westling, Ark. Bot. 11: 90. 1911 293. Penicillium discolor Frisvad & Samson, Antonie van Leeuwenhoek, 72: 120. 1997. 294. Penicillium echinulatum Raper & Thom ex Fassat., Acta Univ. Carol., Biol. 1974: 326. 1977. 295. Penicillium freii Frisvad & Samson, Stud. Mycol. 49: 28. 2004 296. Penicillium hirsutum Dierckx, Ann. Soc. Sci. Bruxelles 25: 89. 1901. 297. Penicillium hordei Stolk, Antonie van Leeuwenhoek 35: 270. 1969. 298. Penicillium melanoconidium (Frisvad) Frisvad & Samson, Stud. Mycol. 49: 28. 2004

299. Penicillium neoechinulatum (Frisvad, Filt. & Wicklow) Frisvad & Samson, Stud. Mycol. 49: 28. 2004 300. Penicillium nordicum Dragoni & Cantoni ex C. Ramírez, Adv. Penicillium Aspergillus Syst.: 139. Antonie van Leeuwenhoek 40: 1. 1974. 301. Penicillium osmophilum Stolk & Veenb.-Rijks, Antonie van Leenwenhoek 40: 1. 1974 302. Penicillium palitans Westling, Ark Bot. 11: 83. 1911 303. Penicillium polonicum K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 445. 1927 304. Penicillium radicicola Overy & Frisvad, Syst. Appl. Microbiol. 26: 633. 2003. 305. Penicillium solitum Westling, Ark. Bot. 11: 65. 1911. 306. Penicillium thymicola Frisvad & Samson, Stud. Mycol. 49: 29. 2004 307. Penicillium tricolor Frisvad et al., Can. J. Bot. 72: 937. 1994 308. Penicillium tulipae Overy & Frisvad, Syst. Appl. Microbiol. 26: 634. 2003 309. Penicillium venetum (Frisvad) Frisvad, Adv. Penicillium Aspergillus Syst.: 275. 2000 310. Penicillium verrucosum Dierckx, Ann. Soc. Sci. Bruxelles 25: 88. 1901 311. Penicillium viridicatum Westling, Ark. Bot. 11: 88. 1911

21. Section Paradoxa

312. Penicillium atramentosum Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 65. 1910. 313. Penicillium crystallinum (Kwon-Chung & Fennell) Samson et al., (published here) 314. Penicillium magnielliptisporum Visagie, Seifert & Samson, Stud. Mycol. 78: 127. 2014. 315. Penicillium malodoratum (Kwon-Chung & Fennell) Samson et al., (published here) 316. Penicillium mexicanum Visagie, Seifert & Samson, Stud. Mycol. 78: 125. 2014 317. Penicillium paradoxum (Fennell & Raper) Samson et al., (published here)

22. Section Penicillium

318. Penicillium brevistipitatum L. Wang & W.Y. Zhuang, Mycotaxon 93: 234. 2005 319. Penicillium clavigerum Demelius, Verh. Zool.-Bot. Ges. Wien 72: 74. 1923. 320. Penicillium concentricum Samson, Stolk & Hadlock, Stud. Mycol. 11: 17. 1976 321. Penicillium coprobium Frisvad, Mycologia 81: 853. 1990. 322. Penicillium coprophilum (Berk. & M.A. Curtis) Seifert & Samson, Adv. Penicillium Aspergillus Syst.: 145. 1985 323. Penicillium dipodomyicola (Frisvad, Filt. & Wicklow) Frisvad, Int. Mod. Meth. Pen. Asp. Clas.: 275. 2000. 324. Penicillium expansum Link, Mag. Ges. Naturf. Freunde Berlin 3: 16. 1809. 325. Penicillium formosanum H.M. Hsieh, H.J. Su & Tzean, Trans. Mycol. Soc. Republ. China 2: 159. 1987. 326. Penicillium gladioli L. McCulloch & Thom, Science 67: 217. 1928 327. Penicillium glandicola (Oudem.) Seifert & Samson, Adv. Penicillium Aspergillus Syst.: 147. 1985 . 328. Penicillium griseofulvum Dierckx, Ann. Soc. Sci. Bruxelles 25: 88. 1901 329. Penicillium italicum Wehmer, Hedwigia 33: 211. 1894 330. Penicillium marinum Frisvad & Samson, Stud. Mycol. 49: 20. 200 331. Penicillium sclerotigenum W. Yamam., Sci. Rep. Hyogo Univ Agric. 1: 69. 1955 332. Penicillium ulaiense H.M. Hsieh, H.J. Su & Tzean, Trans. Mycol. Soc. Rep. China 2: 161. 1987.

333. Penicillium vulpinum (Cooke & Massee) Seifert & Samson, Adv. Penicillium Aspergillus Syst.: 144. 1985

23. Section Ramosa

334. Penicillium jamesonlandense Frisvad & Overy, Int. J. Syst. Evol. Microbiol. 56: 1435. 2006. 335. Penicillium kojigenum G. Sm., Trans. Brit. Mycol. Soc. 44: 43. 1961. 336. Penicillium lanosum Westling, Ark. Bot. 11: 97. 1911. 337. Penicillium lenticrescens Visagie, Seifert & Samson, Stud. Mycol. 78: 123. 2014. 338. Penicillium raistrickii G. Sm., Trans. Brit. Mycol. Soc. 18: 90. 1933 339. Penicillium ribium Frisvad & Overy, Int. J. Syst. Evol. Microbiol. 56: 1436. 2006 340. Penicillium sajarovii Quintan., Av. Aliment. Majora Anim. 22: 539. 1981. 341. Penicillium scabrosum Frisvad, Samson & Stolk, Persoonia 14: 177. 1990. 342. Penicillium simile Davolos et al., Int. J. Syst. Evol. Microbiol. 62: 457. 2012. 343. Penicillium soppii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 476. 1927. 344. Penicillium swiecickii K.M. Zalessky, Bull. Int. Acad. Polon. Sci., Sér. B., Sci. Nat. 1927: 474. 1927. 345. Penicillium virgatum Nirenberg & Kwasna, Mycol. Res. 109: 977. 2005

24. Section Roquefortorum

346. Penicillium carneum (Frisvad) Frisvad, Microbiology, UK, 142: 546. 1996 347. Penicillium paneum Frisvad, Microbiology 142: 546. 1996 348. Penicillium psychrosexualis Houbraken & Samson, IMA Fungus 1: 174. 2010 349. Penicillium roqueforti Thom, U.S.D.A. Bur. Animal Industr. Bull. 82: 35. 1906 25. Section Turbata

350. Penicillium bovifimosum (Tuthill & Frisvad) Houbraken & Samson, Stud. Mycol. 70: 47. 2011 351. Penicillium madriti G. Sm., Trans. Brit. Mycol. Soc. 44: 44. 1961. 352. Penicillium turbatum Westling, Ark. Bot. 11: 128. 1911

Section unclassified 353. Penicillium alfredii Visagie, Seifert & Samson, Stud. Mycol. 78: 116. 2014. 354. Penicillium griseolum G. Sm., Trans. Br. Mycol. Soc. 40: 485. 1957

4. Morphology of Penicillium species

Media and stock solutions used for morphological characterization (C.M. Visagie et al. (2014)

Czapek's agar (CZ,Raper & Thom 1949) Czapek concentrate 10 ml Sucrose 30 g Trace elements stock solution 1 ml Agar 20 g

dH2O 1000 ml *Mix well and autoclave at 121 °C for 15 min.

Czapek Yeast Autolysate agar (CYA, Pitt 1979) Czapek concentrate 10 ml Sucrose 30 g Yeast extract (Difco) 5 g

K2HPO4 1 g Trace elements stock solution 1 ml Agar 20 g

dH2O 1000 ml *Mix well and autoclave at 121 °C for 15 min. pH 6.2 ± 0.2.

Czapek Yeast Autolysate agar with 5 % NaCl (CYAS) Czapek concentrate 10 ml Sucrose 30 g Yeast extract (Difco) 5 g

K2HPO4K2HPO4 1 g Trace elements stock solution 1 ml NaCl 50 g Agar 20 g

dH2OdH2O 1000 ml *Mix well and autoclave at 121 °C for 15 min. pH 6.2 ± 0.2.

Malt Extract agar (MEA,Samson et al. 2010) Malt extract (Oxoid CM0059) 50 g Trace elements stock solution 1 ml dH2O 1000 ml *Mix well and autoclave at 115 °C for 10 min. pH 5.4 ± 0.2.

Oatmeal agar (OA,Samson et al. 2010) Oatmeal / flakes 30 g Trace elements stock solution 1 ml Agar 20 g dH2O 1000 ml

*First autoclave flakes (121 °C for 15 min) in 1000 ml dH2O. Squeeze mixture through cheese cloth and use flow through, topping up to 1000 ml with dH2O with 20 g agar. Autoclave at 121 °C for 15 min. pH 6.5 ± 0.2.

Yeast extract sucrose agar (YES,Frisvad 1981) Yeast extract (Difco) 20 g Sucrose 150 g

MgSO4·7H2O 0.5 g Trace elements stock solution 1 ml Agar 20 g dH2O 885 ml *Mix well and autoclave at 121 °C for 15 min. pH 6.5 ± 0.2. YES is the recommended medium for extrolite profiling of species.

Creatine sucrose agar (CREA,Frisvad 1981) Sucrose 30 g

Creatine·1H2O 3 g

K3PO4·7H2O 1.6 g

MgSO4·7H2O 0.5 g KCl 0.5 g

FeSO4·7H2O 0.01 g Trace elements stock solution 1 ml Bromocresol purple 0.05 g Agar 20 g dH2O 1000 ml *Mix well and autoclave at 121 °C for 15 min. pH

8.0 ± 0.2. Acid production is observed by the colour reaction in CREA (from purple to yellow).

For consistent conidial colours, the addition of zinc-sulphate and copper-sulphate as trace elements (1 g ZnSO4.7H2O and 0.5 g CuSO4.5H2O in 100 ml distilled water) is of utmost importance

4.1. Macromorphology of Penicillium species

1. Growth rate Some species, e.g. P. wellingtonense, grow very restricted on CYA (5–15 mm), while others grow rapidly (P. sumatrense, P. decaturense, P. quebecense, 30–45 mm).

P. wellingtonense P. sumatrense

The growth rate and size of the colonies may vary also in the same species, when grown on different media at the same temperature and for the same period, e,g, in case of P. brasilianum FKI-3368. colonies grown at 25 °C for 7 days.on CYA(a) on malt extract agar (b) on 25% glycerol nitrate agar (c).

www.nature.com Junji Inokoshi et al., Journal of Antibiotics 66, 37-41 (January 2013) |

Incubation temperature affects growth rate and size of the colonies of the the same species on the same medium, e.g. P. wellingtonense has maximum growth temperatures at or below 30 °C and an optimum between 21 and 24 °C. on CYA

21°C 24°C 27°C 30°C 33°C 36°C 37°C P. wellingtonense Houbraken et al.,Stud Mycol. 2011 Nov 15; 70(1): 53–138.

Other species, e.g. P. decaturense. grows well at 30 °C (5–15 mm) optimum and maximum growth temperatures between 21 and 24 °C and even at 30°C

21°C 24°C 27°C 30°C 33°C 36°C 37°C P. decaturense Houbraken et al.,Stud Mycol. 2011 Nov 15; 70(1): 53–138.

Other species, e.g. P. citrinum, on the other hand, can grow at 37°C

21°C 24°C 27°C 30°C 33°C 36°C 37°C P. citrinum Houbraken et al.,Stud Mycol. 2011 Nov 15; 70(1): 53–138.

The size of the colony depends on the space available for the growth, when the fungus is cultured as one inoculum in the centre the colony spreads and covers the whole surface, when several inocula are inoculated the colonies are smaller and are equal in sizes and none of the colonies overgrows the neighbouring colonies as in the following figures:

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2. Appearance and texture of colonies

Velvety colonies are regarded as velvety or velutinous if all, or nearly all, of the vegetative hyphae are submerged in the nutrient substratum, and if the conidiophores rise above the surface in a fairly dense and even stand. Such colonies give the appearance of a surface of velvet, or of a field of grain in miniature. Velvety colonies are, as a rule, comparatively thin but may, upon occa- sion, range up to 0.5 to 1.0 mm. or more in depth, Colonies of this type are characteristically produced in such species as Penicillium frequentans, P. oxalicum, P. roqueforti and P. chrysogenum

P. frequentans, P. oxalicum, P. roqueforti P. chrysogenum

Floccose or lanose colonies produce a cottony mass of branching and interlacing hyphae evenly or unevenly over the surface of the nutrient medium. At a characteristic period in their development., Example: Penicillium camemherti. It may be floccose to funiculose as in case of Penicillium citreonigrum, fasciculate as in P. gladioli

P. camemherti P. citreonigrum P. gladioli

Fasciculate or Coremiform colonies: the marginal areas of rapidly growing colonies to be rough or granular. Microscopic examination in such cases shows that the rough or mealy appearance is due to the aggregation or fasciculation of conidiophores. In some species, fascicles may be at first clearly noticeable in marginal colony areas, only to become largely obscured later by the crowded development of simple conidiophores in the intervening spaces. In a few cases, , the coremiform structures characteristically appear late in colony development. In other species, the vast majority of the conidiophores are from the first aggregated into well- defined fascicles. Occasionally, conidiophores develop almost exclusively in compact columns, or "stalks", which produce single integrated conidium-bearing heads, e.g. Penicillium expansum and Penicillium vulpinum (P. claviforme

Penicillium expansum Penicillium vulpinum

Colonies may be flat, crateriform, subcenter raised with conspicuous zonation or wrinkled, or may show radial sulcation as in P. simile

P. corylophilum MBL Labs Penicillium glabrum Tzean, SS et al.

P. Chrysogenum fineartamerica.com Penicillium simile, Davolos et al. 2011

3. Colour of colonies

Colours within the genus Penicillium is quite wide:  Colonies may be white, e.g. P. simplicissimum , grey as in P. glabrum, grey or bluisc grey as in P. menonorum,.

P. simplicissimum P. glabrum P. menonorum, Peterson

The colour may be orange as in as in P. vanoranjei ,orange to pink , e.g. Penicillium maximae or yellowish brown, e.g. P. thiersi to various grades of brown

P. vanoranjei. Cobus et al. . P. maximae (CBS) P. thiersii Peterson et al., 2004

P. tropicum Houbraken et al P. tropicoides. Houbraken et al

The great majority develop colors in green, e.g. Penicillium expansum , yellow-greens, e.g. P. chrysogenum, and greyish-greens, e.g Penicillium anatolicum , dark-green or dull green , e.g. Penicillium atrofulvum , bluish green, e.g. Penicillium spinolosum or bluish grey

P. crustosum Tzean, SS et al. 1994 P. glaucum P. anatolicum

Many of these Penicillia lose all of their green color in age and assume various shades of yellowish brown, reddish brown, olive gray as in Penicillium cainii ,to almost fuscous.

Penicillium atrofulvum Penicillium spinolosum Penicillium cainii

The colour of the colonies of the same species differ on the different media

P. glabrum on CYA Tzean, SS et al. 199 P/ glabrum on MEA

P. pinophilum, Satoshi Ohte, 2011

The submerged mycelium of various species shows a series of yellow, orange, red, brown, lilac colours with occasional green and even black areas appearing. Such colours are best seen from below and are routinely designated as color of reverse, or simply "reverse."

4.2. Micromorphology of Penicillium

1. The Conidiophores

Essential data regarding conidiophores include their length, septation, the diameter of their cells, and especially their origin in relation to the substratum and to each other. The walls of the conidiophores may be smooth and thin or may be variously roughened, with aerial portions appearing delicately echinulate, granular, or asperulate. In some cases, such as Penicillium roqueforti, these may be marked by conspicuous concretions or warts.

The conidiophore may be simple and unbranched, or variously branched. It may arise from vegetative hyphae within the substratum or from aerial vegetative hyphae variously arranged. It may stand alone, or be more or less closely aggregated into clusters, fascicles, or definite coremia.

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P. spinolosum, MycobankConidiophore stipes 100-300 µm long, smooth- to rough-walled; penicilli monoverticillate P. roqueforti Mycobank the stipes typically ornamented with conspicuous warts, P. chrysogenum Mycobank Conidiophores arising from the substrate, mononematous, usually ter- to quaterverticillate, in some strains more-stage branched; P. griseofulvum, Mycobank: smooth-walled, hyaline, irregular ter- to quaterverticillate, with branches strongly divergent. Stipes undulate,

2. The penicillus (German: "pinsel") is understood to cover the whole branching system and is measured from the lowest branch upon the main axis to the tips of the sterigmata, or conidium bearing cells.

Morphological structures and types of conidiophore branching in Penicillium. a. simple; b. one-stage branched; c. two-stage branched; d. three-stage branched (Samson et al., 1984).

Typical monoverticillate penicillus consists of a terminal verticil of sterigmata only. Typical asymmetric penicillus as seen in P. expansum, shows metulae and branches below the sterigmata.

3. The Sterigmata (Phialides)

The differentiated conidium-producing cell, characteristic of Penicillium and related genera, is variously named the sterigma (plural, sterigmata). The sterigma as it applies to the Penicillia, may be defined as a transformed and highly differentiated cell with a tubular body of fairly typical length and diameter that is characteristically narrowed to a conidium producing tube, or tip, from which unicellular conidia are cut off successively to form a chain of varying length, depending upon the species and the conditions of culture. The resulting chain is characterized by fully ripe cells at its distal end and developing cells at its base, and may contain up to several hundred conidia.

pinterest.com aem.asm.orgwww. pixgood.com Scanning electron microscope (SEM) image of a spore-forming conidiophore of the fungus Penicillium

...

The number of sterigmata in the verticil may be few and readily deter- minable, or suflficiently large to render determination entirely impractic-

The first sterigma in a verticil is a terminal cell which becomes transformed into a spore producing organ, the second pushes out from the cell below at the base of the first, and successive sterigmata bud out to form a whorl, verticil or cluster, on the apex of the main axis or some secondary branch thereof. The apex may be unchanged in size or variously enlarged toward the appearance and proportions of the vesicle of a small Aspergillus as in the following species.

Penicillium paradoxum. Penicillium crystallinum Penicillium malodoratum.

Visagie et al.,Identification and nomenclature of the genus Penicillium. Studies in Mycology, 78, 2014,

4. The metulae

The cells bearing the sterigmata in biverticillate forms are known as metulae . These are characteristically borne in a terminal verticil on the main axis of the conidiophore, or on the main axis and one or more branches from it. If we accept the sterigma as the primary conidium-producing organ, the term metula may properly be restricted to apply to the characteristically differentiated members of the second series of branches, each supporting a group or verticil of sterigmata. They have been variously termed branches, basidia, secondary sterigmata and finally metulae by Westling (1911).

The metulae usually follow closely the diameter of the conidiophore and whatever branches it may produce; less commonly they may become- spicuously smaller in diameter. They are often somewhat enlarged at the tip. In length they may vary quite appreciably or be more or less constant depending upon the species and strain. They generally average a little longer than the sterigmata and their arrangement is fairly characteristic of the species. Variations in shape are usually such as may be attributed to the effect of crowding several elongated cells into a compact verticil upon the apex of the fertile branch. Whenever the walls of the conidiophore are smooth, those of the metulae are also consistently smooth. When the walls of the conidiphore are pitted or rough, the walls of the metulae may or may not be similarly roughened.

Metulae, zean, SS et al. 1994

5. The branches (rami)

In the larger penicilli produced by some of the Asymmetrica, where more than one verticil of metulae is developed, the cells, other than the main axis which bear such metulae, are referred to as branches. In certain species the penicillus is usually typified by a single branch; in others one or more branches may be produced. Biourge referred to branches as rami.

Branches in P. brevicompactum, P. griseofulvum.Mycobank and Penicillium hirsutum, Tzean, SS et al.

6. The conidia According to surface ornamentation, conidia have been classified by Martinez et al. (1982) into six groups: A, smooth-walled (7% of the species); B, delicately roughened (13%); C, warty (28%); D, echinate 910%); E, striate with low irregular ridges (36%); and F, striate with scarce high ridges or bars (6%). Whereas the first two groups are closely related in both shape and average size, a gradual reduction was observed in size and in the length/width (l/w) ratio in the remaining groups. Echinate conidia were globose, having the largest average size. Only four species produced conidia not surpassing 2 micrometers in diameter. Maximum length observed was 8 micrometers, and most elongated conidia had a l/w ratio of 3.5. Forty per cent of the species studied had globose conidia. Conidia of the monoverticillate species were generally smaller, more globose and frequently with ridges. In the Asymmetrica, the conidia were generally larger, and showed ridges in comparatively few species. Conidia of the Symmetrica, which were frequently striate with ridges, presented the most elongated forms. The largest average size was found in

63 the conidia of the Polyverticillata which were generally warty. Finally, we have considered the variations in surface ornamentation of conidia during the evolution of the genus Penicillium and drawn attention to their possible relationship with certain habitats and ways of conidial dispersion.

P. citrinum Penicillium anatolicum Conidia globose to subglobose, smooth Conidia globose, finely roughened

Penicillium cosmopolitanum. Penicillium steckii Conidia globose, rough, Conidia broadly ellipsoidal, slightly fusiform, smooth

Penicillium calidicanium J.L. Chen et al . 2002. P spinulosum conidia ellipsoidal to fusiform, rarely subglobose, conidia spherical, sub-spheroidal, spinose, spinulose

64 roughened or spiral-striated,

P. echinulatum Conidia echinate Conidia of P. glaucum P. chrysogenum conidia Mycobank www.gatan.com pixshark.com

pixgood.com www.phylomedb.org www.sciencephoto.com www.wallpaperdownloader.com long chain conidia of Penicillium species

5. Extrolites produced by various Penicillium species

5.1. Antibiotics:

1.1. Penicillin: P. chrysogenum, P. flavigenum, P. nalgiovense, P. dipodomyis , P. griseofulvum 1.2. Griseofulvin : P. aethiopicum, P. coprophilum , P. dipodomyicola , P. griseofulvum , P. persicinum , P. sclerotigenum

5.2. Anti-cancer:

2.1. Andrastin A : Penicillium species including Penicillium albocoremium[, . P. crustosum , P. allii , P. peneum, P.radicola, P. roqueforti,., P. tulpae, 2.2. Asperphenamate: Penicillium buchwaldii and Penicillium spathulatum 2.3. Asteric acid: P glabrum, P. vulpinum 2.4. Barceloneic acid B : Penicillium albocoremium 2.5. Bredinin: P. brefeldianum. , P. decumbens, P.cyaneum, P. simplicissimum, P. ehrlichii, P. piscarium 2.6. Chrysophanic acid, also known as chrysophanol: Penicillium islandicum, Penicillium citrinum, P. wortmanii 2.7. Chaetoglobosin A: Penicillium aurantiogriseum, Penicillium expansum:, P. marinum, P. discolor 2.8. Chrysophanol: P. islandicum, P. wortmannii 2.9. Citromycin : Penicillium glabrum, P. biliae, Penicillium bilaii . Penicillium striatisporum 2.10. Communesins : Penicillium expansum , Penicillium marinum, Penicillium buchwaldii,Penicillium waksmanii , Penicillium simplicissimum .Penicillium rivulum.. 2.11. Compactin: Penicillium brevicompactum., Penicillium aurantiogriseum , P. cyclopium, P. hirsutum, P. solitum, P. lanosum, P. janczewskii, Penicillium citrinum 2.12. Cyclopiazonic acid: P. camemberti, P. commune, P. dipodomyicola, P. griseofulvum and P. palitans

2.13. Duclauxin: Penicillium duclauxii, Penicillium herquei 2.14. Dehydroaltenusin: Penicillium verruculosum , Penicillium simplicissimum 2.15. Emodin: Penicillium islandicum, P. brunneum, P. janthinellum, Penicillium herquei 2.16. Fumagillin: Penicillium janczewski, P. soppii, Penicillium ribium, Penicillium jensenii. 2.17. Gladiolic acid: Penicillium gladioli 2.18. Gliotoxin : P. glabrum, P. corylophilum 2.19. Mycophenolic acid : Penicillium stoloniferum., Penicillium roqueforti, Penicillium brevicompactum, Penicillium pinophilum , P. bialowiezense, P. carneum, P. rugulosum 2.20. Neooxalin : Penicillium oxalicum 2.21. Oxaline: Penicillium paraherquei 2.22. Paclitaxel: Penicillium raistrickii , Penicillium aurantiogriseum 2.23. Paxilline: Penicillium paxilli 2.24. Pentostatin : Penicillium sclerotigenum, P. marinum 2.25. Secalonic acid D: Penicillium oxalicum,Penicillium isariiforme, P. chrysogenum 2.26. Sesquiterpene Quinone, named penicilliumin A, Penicillium sp. F00120 2.27. Verruculogen: Penicillium verruculosum. Penicillium estinogenum Penicillium simplicissimum, Penicillium raistrickii, Penicillium paxilli 2.28. Wortmannin: Penicillium funiculosum, Penicillium wortmannii

5. 3. Penicillium pigments

3.1. Blue pigments: P. herquei, P. chrysogenum, Penicillium expansum, Penicillium oxalicum 3.2. Blue-green: Penicillium roqueforti, P. aurantiogriseum 3.3. Yellow pigments: P. citrinum, P. chrysogenum, P.oxalicum, P. herquei, P. echinulatum. P. brevicompactum, P. atrovenetum, P. atrosanguineum, Penicillum sclerotiorum 3.4. Orange pigments: P. viridicatum, P. freii, P. cyclopium 3.5. Reddish-brown pigments: P. viridicatum, P. freii, P. cyclopium 3.6. Brown pigments: Penicillium chrysogenum 3.7. Dark brown: P. atramentosum 3.8. Red pigments: P. paneum, P.oxalicum, P. herquei, P. atrovenetum, P. atrosanguineum, Penicillium phoeniceum 3.9. Cherry red pigments: P. persicinum 3.10.Peach-red pigment: Penicillium persicinum

5.4. Enzymes

4.1. Cellulases: P. brasilianum , P. chrysogenum , P. decumbens, Penicillium janthinellum, P. occitanis , 4.2. β-glucanase enzyme: Penicillium oxalicum, Penicillium oxalicum and Penicillium citrinum, Penicillium miczynskii, Penicillium simplicissimum, Penicillium crustosum, 4.3. Pectinase enzyme: Penicillium chrysogenum, Penicillium oxalicum 4.4. Protease enzyme: Penicillium janthinellum, Penicillium roqueforti 4.5. Lipase enzyme: Penicillium expansum , Penicillium camembertii 4.6. a-Amylase enzyme: P. chrysogenum, Penicillium expansum, Penicillium fellutanum 4.7. Xylanase enzyme:

Penicillium sclerotiorum 4.8. Endoglucanase enzyme: Penicillium atrovenetum 4.9. Xylanase enzyme: Penicillium janczewskii , P. janthinellum 4.10. β-Xylosidase enzyme: Penicillium janczewskii 4.11. Penicillopepsin-JT2: Penicillium janthinellum 4.12. Nuclease P enzyme : Penicillium citrinum

5.5. Mycotoxins

5.1. Aflatoxin : P. polonicum 5.2. Anacine: P. aurantiogriseum, P. nordicum 5.3. Asteltoxin: P. cavernicola, P. concentricum, P. confertum, P. formosanum and P. tricolor. 5.4. Auranthine: P. aurantiogriseum 5.5. Aurantiamine: P. aurantiogriseum, P. freii 5.6. Botryodiploidin: P. brevicompactum and P. paneum 5.7. Brevianamide A: P. brevicompactum, P. viridicatum 5.8. Chaetoglobosin A, B: P. discolor, P. expansum 5.9. Chrysogine: P. chrysogenum, P. nalgiovense 5.10. Citrinin: P. citrinum , P. expansum, P. radicicola and P. verrucosum. 5.11. Citromycetin: P. glabrum 5.12. Communesins: P. expansum 5.13. Compactins: P. solitum 5.14. Cyclochlorotine:

P. islandicum 5.15. Cyclopaldic acid: P. commune 5.16. Cyclopenin: P. aurantiocandidum, P. crustosum, P. discolor, P. echinulatum, P. freii, P. polonicum, P. solitum 5.17. Cyclopenol: P. aurantiocandidum, P. crustosum, P. discolor, P. echinulatum, P. freii, P. polonicum, P. solitum 5.18. Cyclopiazonic acid: P. camemberti, P. commune, P. dipodomyicola, P. griseofulvum and P. palitans 5.19. Emodin: P. islandicum 5.20. Erythroskyrin: P. islandicum 5.21. Fumitremorgin A* & B: P. brasilianum, P. palitans 5.22. Islanditoxin: P. islandicum 5.23. Isofumigaclavine A & B: P. roqueforti 5.24. Italicic acid: P. italicum 5.25. Luteoskyrin: P. islandicum 5.26. Marcfortines: P. paneum 5.27. Meleagrin: P. chrysogenum 5.28. Mycophenolic acid: Penicillium stoloniferum, Penicillium roqueforti, Penicillium brevicompactum, Penicillium pinophilum , P. bialowiezense, P. carneum, P. rugulosum 5.29. Ochratoxin A: P. nordicum, P. verrucosum 5.30. Oxaline: P. atramentoseum, P. oxalicum 5.31. Patulin: P. carneum, clavigerum, P. concentricum, P. coprobium , P. dipodomyicola , P. expansum, P. glandicola, P. gladioli, P. griseofulvum, P. marinum, P. paneum,, P. sclerotigenum and P. vulpinum. 5.32. Penitrem A: P. carneum, P. clavigerum, P. crustosum, P. flavigenum, P. glandicola, P. melanoconidium and P. tulipae. 5.33. Penicillic acid:

P. aurantiogriseum, P. aurantiocandidum, P. brasilianum, P. carneum, P. cyclopium, P. freii, P. melanoconidium, P. neoechinulatum, P. polonicum, P. radicicola, P. tulipae and P.viridicatum 5.34. Penitrem A: P. crustosum, P. melanoconidium 5.35. PR-toxin : P. chrysogenum and P. roqueforti. 5.36. Puberulic acid: P. aurantiocandidum 5.37. Raistrick phenols: P. brevicompactum 5.38. Roquefortine C: P. carneum, P. chrysogenum, P. crustosum, P. expansum, P. griseofulvum, P. hirsutum, P. hordei, P. melanoconidium, P. paneum, P. roqueforti 5.39. Rubratoxin: P. crateriforme 5.40. Rugulosin: P. islandicum, P. rugulosum, P. variabile 5.41. Rugulovasine A & B: P. crateriforme, P. atramentoseum, P. commune 5.42. Secalonic acid A: P. chrysogenum and P. confertum 5.43. Secalonic acid D & F: P. oxalicum 5.44. Spiculisporic acid: P. crateriforme 5.46. Tanzawaic acid A: P. citrinum 5.47. Terrestric acid: P. crustosum, P. hirsutum, P. hordei 5.48. Territrems: P. cavernicola and P. echinulatum. 5.49. Tryptoquivalins: P. digitatum 5.50. Verrucins: P. verrucosum 5.51. Verrucologen: P. brasilianum 5.52. Verrucolone: P. italicum, P. nordicum, P. olsonii, P. verrucosum 5.53. Verrucosidin: P. aurantiogriseum, P. melanoconidium, P. polonicum 5.54. Viomellein:

P. clavigerum, P. cyclopium, P. freii, P. melanoconidium, P. tricolor and P. viridicatum 5.55. Vioxanthin: P. clavigerum, P. cyclopium, P. freii, P. melanoconidium, P. tricolor and P. viridicatum Viridic acid: P. nordicum and P. viridicatum 5.56. Viridicatumtoxin: P. brasilianum, P. aethiopicum. 5.57. Xanthomegnin: P. clavigerum, P. cyclopium, P. freii, P. melanoconidium, P. tricolor and P. viridicatum.

6. Molecular genetics of Penicillium

6.1. Genome sequencing

6.1.1. P. chrysogenum van den Berg et al. (2008) sequenced the 32.19 Mb genome of P. chrysogenum Wisconsin54-1255 and identified numerous genes responsible for key steps in penicillin production. DNA microarrays were used to compare the transcriptomes of the sequenced strain and a penicillinG high-producing strain, grown in the presence and absence of the side-chain precursor phenylacetic acid. Transcription of genes involved in biosynthesis of valine, cysteine and alpha-aminoadipic acid-precursors for penicillin biosynthesis-as well as of genes encoding microbody proteins, was increased in the high-producing strain. Some gene products were shown to be directly controlling beta- lactam output. Many key cellular transport processes involving penicillins and intermediates remain to be characterized at the molecular level. Genes predicted to encode transporters were strongly overrepresented among the genes transcriptionally upregulated under conditions that stimulate penicillinG production, illustrating potential for future genomics-driven metabolic engineering. Jónás et al. (2014) investigated the catabolism of lactose by Penicillium chrysogenum. In silico analysis of the genome sequences revealed that P. chrysogenum features at least five putative β-galactosidase (bGal)-encoding genes at the annotated loci Pc22g14540, Pc12g11750, Pc16g12750, Pc14g01510 and Pc06g00600. The first two proteins appear to be orthologs of two Aspergillus nidulans family 2 intracellular glycosyl hydrolases expressed on lactose. The latter three P. chrysogenum proteins appear to be distinct paralogs of the extracellular bGal from A. niger, LacA, a family 35 glycosyl hydrolase. The P. chrysogenum genome also specifies two putative lactose transporter genes at the annotated loci Pc16g06850 and Pc13g08630. These are orthologs of paralogs of the gene encoding the high-affinity lactose permease (lacpA) in

A. nidulans for which P. chrysogenum appears to lack the ortholog. Transcript analysis of Pc22g14540 showed that it was expressed exclusively on lactose, whereas Pc12g11750 was weakly expressed on all carbon sources tested, including D-glucose. Pc16g12750 was co-expressed with the two putative intracellular bGal genes on lactose and also responded on L-arabinose. The Pc13g08630 transcript was formed exclusively on lactose. The data strongly suggest that P. chrysogenum exhibits a dual assimilation strategy for lactose, simultaneously employing extracellular and intracellular hydrolysis, without any correlation to the penicillin-producing potential of the studied strains.

6.1.2. Penicillium cyclopium The complete gene (PG37 lipI) encoding an alkaline lipase (PG37 LipI) was cloned from the genomic DNA of Penicillium cyclopium PG37 (Zhang et al., 2011). The cloned PG37 lipI is 2020 bp in length, consisting of 632 bp of the 5' flanking promoter region and 1388 bp of the downstream fragment that contains 6 exons and 5 short introns. The promoter region harbours putative TATA box, CAAT box and several transcription factor binding sites. The open reading frame (ORF) encodes a PG37 LipI of 285 amino acid residues, which was predicted to contain a 20-aa signal peptide, a 7- aa propeptide and a 258-aa mature peptide with a conserved motif Gly-X-Ser-X-Gly. The PG37 LipI showed only 32%, 30%, 28% and 26% identity with lipases of Aspergillus parasiticus, Penicillium camembertii, Thermomyces lanuginosus and Rhizomucor miehei, respectively. It was predicted that the main secondary structures of PG37 LipI are alpha-helix and random coil. Three amino acid residues, Ser132-Asp188- His241, compose the enzymatic active center in the tertiary structure. 6.1.3. Penicillium digitatum The complete mitochondrial genome of Penicillium digitatum (Pers.:Fr) Sacc is reported by Sun et al. (2011). Comparative analysis revealed its close relationship to mitochondrial genomes of other Penicillium and Aspergillus species, both in gene content and in arrangement. The intron content of protein coding genes revealed several differences. The different exon-intron organization of Cytochrome Oxidase Subunit 1 genes indicated their common origin before the divergence of Penicillium and Aspergillus, and that, largely, their introns were transmitted vertically. Marcet-Houben (2012) sequenced two strains of P. chrysogenum , which were isolated in Spain, and another isolated in China. Comparison with the closely-related but non-phytopathogenic P. chrysogenum revealed a much smaller gene content in P. digitatum, consistent with a more specialized lifestyle. They showed that large regions of the P. chrysogenum genome, including entire supercontigs, are absent from P. digitatum, and that this is the result of large gene family expansions rather than acquisition through horizontal gene transfer. The analysis of the P. digitatum genome is

73 indicative of heterothallic sexual reproduction and revealed the molecular basis for the inability of this species to assimilate nitrate or produce the metabolites patulin and penicillin. Sun et al. (2013) sequenced the whole genome of the Penicillium digitatum R1 genotype strain Pd01-ZJU and investigated the genes and DNA elements highly associated with drug resistance. Variation in DNA elements related to drug resistance between P. digitatum strains was revealed in both copy number and chromosomal location, indicating that their recent and frequent translocation might have contributed to environmental adaptation. In addition, ABC transporter proteins in Pd01-ZJU were characterized, and the roles of typical subfamilies (ABCG, ABCC, and ABCB) in imazalil resistance were explored using real-time PCR. Seven ABC proteins, including the previously characterized PMR1 and PMR5, were induced by imazalil, which suggests a role in drug resistance. 6.1.4. Penicillium solitum Eldarov et al. (2012) determined the complete mitochondrial genome sequence of the compactin-producing fungus Penicillium solitum strain 20-01. The 28 601-base pair circular-mapping DNA molecule encodes a characteristic set of mitochondrial proteins and RNA genes and is intron-free. All 46 protein- and RNA-encoding genes are located on one strand and apparently transcribed in one direction. Comparative analysis of this mtDNA and previously sequenced but unannotated mitochondrial genomes of several medically and industrially important species of the Aspergillus/Penicillium group revealed their extensive similarity in terms of size, gene content and sequence, which is also reflected in the almost perfect conservation of mitochondrial gene order in Penicillium and Aspergillus. Phylogenetic analysis based on concatenated mitochondrial protein sequences confirmed the monophyletic origin of Eurotiomycetes.

6.1.5. Penicillium decumbens The genomic and secretomic analyses of Penicillium decumbens, that has been used in industrial production of lignocellulolytic enzymes in China for more than fifteen years, was ) presented by Liu et al. (2013). Comparative genomics analysis with the phylogenetically most similar species Penicillium chrysogenum revealed that P. decumbens has evolved with more genes involved in plant cell wall degradation, but fewer genes in cellular metabolism and regulation. Compared with the widely used cellulase producer Trichoderma reesei, P. decumbens has a lignocellulolytic enzyme system with more diverse components, particularly for cellulose binding domain- containing proteins and hemicellulases. Further, proteomic analysis of secretomes revealed that P. decumbens produced significantly more lignocellulolytic enzymes in the medium with cellulose-wheat bran as the carbon source than with glucose. Gene

74 curation from four gene predictors yielded 10,021 protein-coding gene models of which 35.2% were well supported by 454 transcriptome sequencing data (100% identity in full-length Totally, 6,186 proteins were assigned to Gene Ontology (GO) categories. 6.1.6. Penicillium aurantiogriseum The genome of Penicillium aurantiogriseum NRRL 62431 was sequenced and gene candidates that may be involved in paclitaxel biosynthesis were identified by comparison with the 13 known paclitaxel biosynthetic genes in Taxus (Yang et al., 2014).

6.1.7. P. roqueforti

Hidalgo et al. (2014) have cloned and sequenced a four gene cluster that includes the ari1 gene from P. roqueforti. Gene silencing of each of the four genes (named prx1 to prx4) resulted in a reduction of 65-75% in the production of PR-toxin indicating that the four genes encode enzymes involved in PR-toxin biosynthesis. Interestingly the four silenced mutants overproduce large amounts of mycophenolic acid, an antitumor compound formed by an unrelated pathway suggesting a cross-talk of PR-toxin and mycophenolic acid production. An eleven gene cluster that includes the above mentioned four prx genes and a 14-TMS drug/H(+) antiporter was found in the genome of Penicillium chrysogenum. This eleven gene cluster has been reported to be very poorly expressed in a transcriptomic study of P. chrysogenum genes under conditions of penicillin production (strongly aerated cultures). They found that this apparently silent gene cluster is able to produce PR-toxin in P. chrysogenum under static culture conditions on hydrated rice medium. Noteworthily, the production of PR- toxin was 2.6-fold higher in P. chrysogenum npe10, a strain deleted in the 56.8kb amplifiable region containing the pen gene cluster, than in the parental strain Wisconsin 54-1255 providing another example of cross-talk between secondary metabolite pathways in this fungus. Sequencing of the two leading filamentous fungi used in cheese making, P. roqueforti and P. camemberti, and comparison with the penicillin producer P. rubens reveals a 575 kb long genomic island in P. roqueforti--called Wallaby--present as identical fragments at non-homologous loci in P. camemberti and P. rubens. Wallaby is detected in Penicillium collections exclusively in strains from food environments. Wallaby encompasses about 250 predicted genes, some of which are probably involved in competition with microorganisms. The occurrence of multiple recent eukaryotic transfers in the food environment provides strong evidence for the importance of this understudied and probably underestimated phenomenon in eukaryotes (Cheeseman et al., 2014).

6.1.8. P. expansum Yu et al.(2014) reported the draft genome sequence of P. expansum R19 isolated in 2011 from a decomposing red delicious apple in Carlisle, PA, were inoculated in potato dextrose broth (PDB) at 25°C for 7 days. Genomic DNA was prepared with a DNeasy Plant Maxi Kit (Qiagen) according to the manufacturer’s instructions. Preliminary annotation results demonstrated that the P. expansum R19 genome harbors 10,554 predicted genes, with an average gene length of 1,599 bp. The total length of the coding sequence (genes) is 16,873,185 bp, which makes up 53.70% of the genome. There are 120 tRNA genes and 48 5S rRNA genes, respectively, as predicted by tRNAscan-SE 1.21 (5) and RNAmmer (6). The gene ontology (GO) analysis indicated that a total of 6,831 proteins fall in 985 major GO terms. It is estimated that there are 59 gene clusters putatively involved in the biosynthesis of secondary metabolites, as predicted by SMURF (7). This is similar to the result predicted by the AntiSMASH program (8), which resulted in 57 clusters. It is expected to identify groups of genes that are putatively involved in spore germination and fungal mycelial growth, as well as genes involved in mycotoxin biosynthesis Li et al. (2015) reported the whole genome sequence of P. expansum (33.52 Mb) and P. italicum (28.99 Mb), and identified differences in genome structure, important pathogenic characters, and secondary metabolite (SM) gene clusters in Penicillium species. They identified a total of 55 gene clusters potentially related to secondary metabolism, including a cluster of 15 genes (named PePatA ~ O), that may be involved in patulin biosynthesis in P. expansum. Functional studies confirmed that PePatL and PePatK play crucial roles in the biosynthesis of patulin, and that patulin production is not related to virulence of P. expansum. Collectively, P. expansum contains more pathogenic genes and SM gene clusters, in particular an intact patulin cluster, than P. italicum or P. digitatum. These findings provide important information relevant to understanding the molecular network of patulin biosynthesis and mechanisms of host-specificity in Penicillium species. The genomes of three Penicillium expansum strains, the main postharvest pathogen of pome fruit, and one Pencillium italicum strain, a postharvest pathogen of citrus fruit, were sequenced and compared with 24 other fungal species (Ballester et al., 2015). A genomic analysis of gene clusters responsible for the production of secondary metabolites was performed. Putative virulence factors in P. expansum were identified by means of a transcriptomic analysis of apple fruits during the course of infection. Despite a major genome contraction, P. expansum is the Penicillium species with the largest potential for the production of secondary metabolites. Results using knockout

76 mutants clearly demonstrated that neither patulin nor citrinin are required by P. expansum to successfully infect apples.

6.2. Genetic transformation

6.2.1. Penicillium nordicum A gfp reporter gene strain of Penicillium nordicum was constructed by Schmidt-Heydt et al. (2009) in order to be able to study the influence of environmental parameters on ochratoxin A biosynthesis. To introduce the gfp gene an Agrobacterium tumefaciens mediated transformation system (ATMT) for P. nordicum was established resulting in a transformation efficiency of about 60 transformants per microg of DNA. The selection principle was based on the hygromycin B resistance gene located on the TI-DNA fragment of the binary vector system. PCR and Southern blot hybridization revealed that the TI-DNA was integrated into the chromosome of P. nordicum. To show that the GFP protein can be used as a reporter gene in P. nordicum, this species was subsequently transformed with a vector, carrying a gfp gene under the control of the strong constitutive gpd promoter of A. nidulans. Moreover in this vector construction the gfp gene contained a stuA nuclear localization domain. Successful transformed strains showed a strong GFP activity located in the nuclei after light stimulation in contrast to the wild type which showed only very weak unspecific auto fluorescence under these conditions. Based on this proof of principle a vector was constructed in which the promoter of the otapksPN gene, the gene of the ochratoxin A polyketide synthase of P. nordicum was placed in front of the gfp gene. This construct was transformed into P. nordicum by ATMT and the resulting transformants were analysed by fluorescence microscopy. In these transformants the whole mycelial cells showed GFP activity after light stimulation, whereas the wild type strain did not. When the transformed strains were grown on medium which suppressed ochratoxin A biosynthesis, a very low level of fluorescence could be detected, whereas a high level of fluorescence was seen after growth on medium supportive for ochratoxin A biosynthesis. 6.2.2. Penicillium decumbens

In a study carried out by Ma et al. (2011), a beta-glucosidase I coding sequence from Penicillium decumbens was ligated with the cellobiohydrolase I (cbh1) promoter of T. reesei, a well-known cellulase producer and widely applied in enzyme industry, to increase its ability to efficiently decompose cellulose and introduced into the genome of T. reesei strain Rut-C30 by Agrobacterium-mediated transformation. In comparison to that from the parent strain, the beta-glucosidase activity of the enzyme complexes from two selected transformants increased 6- to 8-fold and their filter paper

77 activity (FPAs) was enhanced by 30% on average. The transformant's saccharifying ability towards pretreated cornstalk was also significantly enhanced. To further confirm the effect of heterologous beta-glucosidase on the cellulase activity of T. reesei, the heterologously expressed pBGL1 was purified and added to the enzyme complex produced by T. reesei Rut-C30. Supplementation of the Rut-C30 enzyme complex with pBGL1 brought about 80% increase of glucose yield during the saccharification of pretreated cornstalk. 6.2.3. Penicillium brevicompactum Ren et al. (2013) developed protoplast transformation methods mediated by polyethylene glycol in Penicillium brevicompactum, using phleomycin resistance gene (Sh ble) as a dominant selection marker. The frequency of transformation was up to 2 - 3 transformants per microg DNA; analysis of the transformants by PCR showed that the foreign DNA had been integrated into the host genome. The transformants retained stable after generation. They concluded that the establishment of the genetic transformation system of Penicillium brevicompactum could serve as the basis for the research of molecular biology and the breeding of gene engineering of the fungus. 6.2.4. P. griseoroseum Teixeira et al. (2014) indicated that analysis of plg1 and pgg2 pectinolytic genes expression and copy number in recombinant multi-copy strains of P. griseoroseum demonstrated that an increase in the number of gene copies could increase enzyme production, but the transcription could be affected by the gene integration position. Placing a copy of the plg1 gene under the control of the gpd promoter of Aspergillus nidulans yielded a 200-fold increase in transcription levels compared to the endogenous gene, and two copies of the pgg2 gene produced an 1100-fold increase compared with the endogenous gene. These results demonstrated that transcription, translation, and protein secretion in the fungus P. griseoroseum respond to an increased number of gene copies in the genome. The work of Goarin et al. (2014) provides the first example of gene replacement by homologous recombination in P. roqueforti, demonstrating that knockout experiments can be performed in this fungus. To do so, the authors improved the existing transformation method to integrate transgenes into P. roqueforti genome. In the meantime, they cloned the PrNiaD gene, which encodes a NADPH-dependent nitrate reductase that reduces nitrate to nitrite. Then, they performed a deletion of the PrNiaD gene from P. roqueforti strain AGO. The ΔPrNiaD mutant strain is more resistant to chlorate-containing medium than the wild-type strain, but did not grow on nitrate- containing medium. Because genomic data are now available, they believe d that generating selective deletions of candidate genes will be a key step to open the way for a comprehensive exploration of gene function in P. roqueforti.

6.2.5. Penicillium chrysogenum The phy gene, which encodes a phytase in Penicillium chrysogenum CCT 1273, was cloned into the vector pAN-52-1-phy and the resulting plasmid was used for the cotransformation of Penicillium griseoroseum PG63 protoplasts (Ribeiro Corrêa et al., 2015). Among the 91 transformants obtained, 23 were cotransformants. From there, the phytase activity of these 23 transformants was evaluated and P. griseoroseum T73 showed the highest. The recombinant strain P. griseoroseum T73 contained the phy gene integrated in at least three sites of the genome and showed a 5.1-fold increase in phytase activity in comparison to the host strain (from 0.56 ± 0.2 to 2.86 ± 0.4 U μg protein(-1)). The deduced PHY protein has 483 amino acids; an isoelectric point (pI) higher than that reported for phytases from filamentous fungi (7.6); higher activity at pH 2.0 (73%), pH 5.0 (100%) and 50 °C; and is stable at pH values 3.0-8.0 and temperatures 70-80 °C. PHY produced by the recombinant strain P. griseoroseum T73 was stable after four weeks of storage at -20, 8 and 25 °C and was effective in releasing Pi, especially from soybeans. The data presented here show that P. griseoroseum is a successful host for expression of heterologous protein and suggest the potential use of PHY in the animal nutrition industry.

6.3. The biosynthetic pathway

Major abundant metabolites of the roquefortine/meleagrin pathway were identified as histidyltryptophanyldiketopiperazine (HTD), dehydrohistidyltryptophanyldi- ketopiperazine (DHTD), roquefortine D, roquefortine C, glandicoline A, glandicoline B and meleagrin (Ali et al. (2013). Specific genes could be assigned to each enzymatic reaction step. The non-ribosomal peptide synthetase RoqA accepts L-histidine and L-tryptophan as substrates leading to the production of the diketopiperazine HTD. DHTD, previously suggested to be a degradation product of roquefortine C, was found to be derived from HTD involving the cytochrome P450 oxidoreductase RoqR. The dimethyl-allyltryptophan synthetase RoqD prenylates both HTD and DHTD yielding directly the products roquefortine D and roquefortine C without the synthesis of a previously suggested intermediate and the involvement of RoqM. This leads to a branch in the otherwise linear pathway. Roquefortine C is subsequently converted into glandicoline B with glandicoline A as intermediates, involving two monooxygenases (RoqM and RoqO) which were mixed up in an earlier attempt to elucidate the biosynthetic pathway.

Eventually, meleagrin is produced from glandicoline B involving a methyltransferase (RoqN). . The biosynthetic pathway of patulin is chemically well-characterized, but its genetic bases remain largely unknown with only few characterized genes in less economic relevant species. The identification and positional organization of the patulin gene cluster in P. expansum strain NRRL 35695 was done by Tannous et al. (2014). Several amplification reactions were performed with degenerative primers that were designed based on sequences from the orthologous genes available in other species. An improved genome Walking approach was used in order to sequence the remaining adjacent genes of the cluster. RACE-PCR was also carried out from mRNAs to determine the start and stop codons of the coding sequences. The patulin gene cluster in P. expansum was proved to consist of 15 genes in the following order: patH, patG, patF, patE, patD, patC, patB, patA, patM, patN, patO, patL, patI, patJ, and patK. The kinetics of patulin cluster genes expression was studied under patulin-permissive conditions (natural apple-based medium) and patulin-restrictive conditions (Eagle's minimal essential medium), and demonstrated a significant association between gene expression and patulin production. The independently synthesize paclitaxel Penicillium aurantiogriseum was established by Yang et al. (2014). The genome of Penicillium aurantiogriseum NRRL 62431 was sequenced and gene candidates that may be involved in paclitaxel biosynthesis were identified by comparison with the 13 known paclitaxel biosynthetic genes in Taxus. It was found that paclitaxel biosynthetic gene candidates in P. aurantiogriseum NRRL 62431 have evolved independently and that horizontal gene transfer between this endophytic fungus and its plant host is unlikely.

Reprogramming the antibiotics-producing fungus Penicillium chrysogenum, with discovery and engineering of a cytochrome P450 enzyme involved in the hydroxylation of the precursor compactin, enabled high level fermentation of the correct form of pravastatin to facilitate efficient industrial-scale statin drug production. Key steps leading to the successful outcome included the identification and deletion of a fungal gene responsible for degradation of compactin, in addition to evolution of the P450 to enable it to catalyse the desired stereoselective hydroxylation step required for high level pravastatin production. Statins are successful, widely used drugs that decrease the risk of coronary heart disease and strokes by lowering cholesterol levels. The development of this group of drugs has been one of the major breakthroughs in human healthcare over the last two decades (Kirsty et al., 2015).

6.4. Virulence genes of Penicillium digitatum

The phenotypes and genotypes of 403 isolates of P. digitatum, collected from packing houses and supermarkets in Zhejiang, China, during 2000 to 2010, were characterized in terms of their imazalil sensitivity (Sun et al., 2011). The frequency of detected imazalil- resistant (IMZ-R) isolates increased from 2.1% in 2000 to 60-84% during 2005-2010. Only 6.5% and 4.5% of the collected IMZ-R isolates belong to the previously described IMZ-R1 and IMZ-R2 genotypes, respectively. To determine the resistance mechanism of the predominant and novel IMZ-R isolates of P. digitatum (termed IMZ-R3), genes PdCYP51B and PdCYP51C, homologous to the sterol 14α-demethylase encoded gene PdCYP51, were cloned from six IMZ-R3 and eight imazalil-sensitive (IMZ-S) isolates of P. digitatum. A unique 199-bp insertion was observed in the promoter region of PdCYP51B in all IMZ-R3 isolates examined but in none of the tested IMZ-S isolates. Further analysis by PCR confirmed that this insertion was present in all IMZ-R3 isolates but absent in IMZ-S, IMZ-R1, and IMZ-R2 isolates. Transcription levels of PdCYP51B in three IMZ-R3 isolates were found to be 7.5- to 13.6-fold higher than that in two IMZ- S isolates of P. digitatum. Introduction of another copy of PdCYP51B ( s ) (from IMZ- S) into an IMZ-S isolate decreased the sensitivity of P. digitatum to 14α-demethylation inhibitors (DMIs) only to a small extent, but introduction of a copy of PdCYP51B ( R ) (from IMZ-R3) dramatically increased the resistance level of P. digitatum to DMIs. Regarding PdCYP51C, no consistent changes in either nucleotide sequence or expression level were correlated with imazalil resistance among IMZ-R and IMZ-S isolates. Based on these results, it is concluded that (1) the CYP51 family of P. digitatum contains the PdCYP51B and PdCYP51C genes, in addition to the known gene PdCYP51A (previously PdCYP51); (2) PdCYP51B is involved in DMI fungicide resistance; and (3) overexpression of PdCYP51B resulting from a 199-bp insertion mutation in the promoter region of PdCYP51B is responsible for the IMZ-R3 type of DMI resistance in P. digitatum. Zhu et al. (2014) reported the identification and functional characterization of PdGcs1, a gene encoding GlcCer synthase (GCS) essential for the biosynthesis of GlcCers, in Penicillium digitatum genome. They demonstrated that the deletion of PdGcs1 in P. digitatum resulted in the complete loss of production of GlcCer (d18:1/18:0 h) and GlcCer (d18:2/18:0 h), a decrease in vegetation growth and sporulation, and a delay in spore germination. The virulence of the PdGcs1 deletion mutant on citrus fruits was also impaired, as evidenced by the delayed occurrence of water soaking lesion and the formation of smaller size of lesion. These results suggest that PdGcs1 plays an important role in regulating cell growth, differentiation, and virulence of P. digitatum by controlling the biosynthesis of GlcCers. In order to identify P. digitatum genes up-regulated during the infection of oranges that may constitute putative virulence factors, López-Pérez et al. (2014) followed a polymerase chain reaction (PCR)-based suppression subtractive hybridization and

81 cDNA macroarray hybridization approach. The origin of expressed sequence tags (ESTs) was determined by comparison against the available genome sequences of both organisms. Genes coding for fungal proteases and plant cell wall-degrading enzymes represent the largest categories in the subtracted cDNA library. Northern blot analysis of a selection of P. digitatum genes, including those coding for proteases, cell wall- related enzymes, redox homoeostasis and detoxification processes, confirmed their up- regulation at varying time points during the infection process. Agrobacterium tumefaciens-mediated transformation was used to generate knockout mutants for two genes encoding a pectin lyase (Pnl1) and a naphthalene dioxygenase (Ndo1). Two independent P. digitatum Δndo1 mutants were as virulent as the wild-type. However, the two Δpnl1 mutants analysed were less virulent than the parental strain or an ectopic transformant. An orthologus gene encoding a putative sterol regulatory element-binding protein (SREBP) was identified in the genome of P. digitatum and named sreA (Liu et al., 2015). It was proved that sreA is a critical transcription factor gene required for prochloraz resistance and full virulence in P. digitatum and is involved in the regulation of cyp51 expression

6.5. Sexual life cycle of P. roqueforti Ropars et al. (2014) reported, for the first time, the induction of the sexual structures of P. roqueforti - ascogonia, cleistothecia and ascospores. The progeny of the sexual cycle displayed clear evidence of recombination. They used the recently published genome sequence for this species to develop microsatellite markers for investigating the footprints of recombination and population structure in a large collection of isolates from around the world and from different environments. They found tremendous genetic diversity within P. roqueforti, even within cheese strains and identified six highly differentiated clusters that probably predate the use of this species for cheese production. Screening for phenotypic and metabolic differences between these populations could guide future development strategies. Böhm et al. (2015) recently discovered mating-type loci and a sexual life cycle in the penicillin-producing fungus, Penicillium chrysogenum, MAT1-2. All industrial penicillin production strains worldwide are derived from a MAT1-1 isolate. Similar to MAT1-1, the MAT1-2 locus has functions beyond sexual development. Unlike MAT1-1, the MAT1-2 locus affects germination and surface properties of conidiospores and controls light-dependent asexual sporulation. Mating of the MAT1-2 wild type with a MAT1-1 high penicillin producer generated sexual spores. The genomic sequences of parental and progeny strains using next-generation sequencing provided an evidence

82 for genome-wide recombination. SNP calling showed that derived industrial strains had an uneven distribution of point mutations compared with the wild type.

6.6. Genome shuffling

Wang et al. (2013) reported a high producer of nuclease P1, Penicillium citrinum G- 16, that was bred by the classical physics-mutagenesis and genome shuffling process. The starting populations were generated by (60)Co γ-irradiation mutagenesis. The derived two protoplast fractions were inactivated by heat-treatment and ultraviolet radiation respectively, then mixed together and subjected to recursive protoplast fusion. Three recombinants, E-16, F-71, and G-16, were roughly obtained from six cycles of genome shuffling. The activity of nuclease P1 by recombinant G-16 was improved up to 1,980.22 U4/ml in a 5-l fermentor, which was 4.7-fold higher than that of the starting strain. The sporulation of recombinant G-16 was distinguished from the starting strain. Random amplified polymorphic DNA assay revealed genotypic differences between the shuffled strains and the wild type strain. The close similarity among the high producers suggested that the genetic basis of high-yield strains was achieved by genome shuffling. 6.7. Mechanism of conidiation in P. decumbens Qin et al. (2013) investigated the mechanism of conidiation in P. decumbens generated mutants defective in two central regulators of conidiation, FluG and BrlA. Deletion of fluG resulted in neither "fluffy" phenotype nor alteration in conidiation, indicating possible different upstream mechanisms activating brlA between P. decumbens and Aspergillus nidulans. Deletion of brlA completely blocked conidiation. Further investigation of brlA expression in different media (nutrient-rich or nutrient- poor) and different culture states (liquid or solid) showed that brlA expression is required but not sufficient for conidiation. The brlA deletion strain exhibited altered hyphal morphology with more branches. Genome-wide expression profiling identified BrlA-dependent genes in P. decumbens, including genes previously reported to be involved in conidiation as well as previously reported chitin synthase genes and acid protease gene (pepB). The expression levels of seven secondary metabolism gene clusters (from a total of 28 clusters) were drastically regulated in the brlA deletion strain, including a downregulated cluster putatively involved in the biosynthesis of the mycotoxins roquefortine C and meleagrin. In addition, the expression levels of most cellulase genes were upregulated in the brlA deletion strain detected by real-time quantitative PCR. The brlA deletion strain also exhibited an 89.1 % increase in cellulase activity compared with the wild-type strain. The results showed that BrlA in P. decumbens not only has a key role in regulating conidiation, but it also regulates secondary metabolism extensively as well as the expression of cellulase genes.

6.8. Expression and characterization of enzymes Cardoso et al. (2010) evaluated an extracellular pectin lyase (PL) production by recombinant P. griseoroseum strain 105 in submerged fermentation system bioreactors BioFloIII and BioFloIV using 2 or 10 L working volumes under different growth conditions and to analyze the production of cellulase, polygalacturonase, pectin methylesterase, and protease. PL overproduction by recombinant P. griseoroseum strain 105 was 112 times higher than that of P. griseoroseum PG63 grown in sugarcane juice. Cellulases and proteases were not detected in the culture filtrate, and evaluation for extracellular proteins in the culture medium by SDS-PAGE showed the presence of a 36 kDa predominant band, similar to the molecular mass estimated from the nucleotide sequence of plg1 gene for PL of P. griseoroseum strain 105. This recombinant strain provides the advantage of PL production, which predominates over other extracellular proteins usually present in most commercial pectinase preparations, using sugarcane juice as a substrate of low cost. Regueira et al. (2011) reported the discovery of a polyketide synthase (PKS), MpaC, which they successfully characterized and identified as responsible for MPA production in Penicillium brevicompactum. mpaC resides in what most likely is a 25-kb gene cluster in the genome of Penicillium brevicompactum. The gene cluster was successfully localized by targeting putative resistance genes, in this case an additional copy of the gene encoding IMP dehydrogenase (IMPDH). They reported the cloning, sequencing, and the functional characterization of the MPA biosynthesis gene cluster by deletion of the polyketide synthase gene mpaC of P. brevicompactum and bioinformatic analyses. As expected, the gene deletion completely abolished MPA production as well as production of several other metabolites derived from the MPA biosynthesis pathway of P. brevicompactum. Our work sets the stage for engineering the production of MPA and analogues through metabolic engineering. Maĭsuradze et al. (2011) have cloned 4r novel genes of the enzymes of the endoxylanase families found in the mycelial fungus Penicillium canescens . The xylB, xylC, and xylD genes encode endoxylanases of glycosyl hydrolase family 11; the xylEgene, those of family 10. In the promoter region of the xylB, xylC, and xylD genes, the binding sequences for the protein activator of xylanolytic gene transcription have been found; the promoter region of the xylB gene contains the binding sequences for the catabolite repression protein. Since the TATAA sequence, which is an element of the minimal eukaryotic promoter, has not been found in the promoter region of the xylC gene, in contrast to those of the xylB and xylD genes, it may be assumed that this gene is silent. Comparative phylogenetic analysis has shown that the cloned genes are highly homologous to some endoxylanase genes of mycelial fungi of the genera Penicillium and Aspergillus. However, within the species P. canescens, they exhibit a low homology both within and between families, and they diverge into

84 different branches of the phylogenetic tree, which suggest divergence of the genes of this group at an early stage of evolution. Teixeira et al. (2011) reported on a strain with high production of pectin lyase (PL) was transformed with the plasmid pAN52pgg2, containing the gene encoding PG of P. griseoroseum, under control of the gpd promoter gene from Aspergillus nidulans. Southern blot analysis demonstrated that all strain had at least one copy of pAN52pgg2 integrated into the genome. The recombinant strain P. griseoroseum T20 produced levels of PL and PG that were 266- and 27-fold greater, respectively, than the wild-type strain. Furthermore, the extracellular protein profile of recombinant T20 showed two protein bands of c. 36 and 38 kDa, associated with PL and PG, respectively. Gandía et al. (2012) carried out the isolation and characterization of chitin synthase genes (CHS) of Penicillium digitatum. Using distinct sets of degenerate primers designed from conserved regions of CHS genes of yeast and filamentous fungi, PCR methods, and a DNA genomic library, five putative CHS genes (PdigCHSI, PdigCHSII, PdigCHSIII, PdigCHSV, and PdigCHSVII) were identified, isolated, sequenced, and characterized. Phylogenetic analyses, sequence identity, and domain conservation support the annotation as CHS. A very high sequence identity and strong synteny were found with corresponding regions from the genome of Penicillium chrysogenum. Gene expression of P. digitatum CHS genes during mycelium axenic growth, under oxidative and osmotic stress conditions, and during infection of citrus fruits was confirmed and quantified using quantitative RT-PCR (qRT-PCR). PdigCHSIII had the highest expression among the five genes by one order of magnitude, while PdigCHSII had the lowest. However, PdigCHSII was strongly induced coincident with conidial production, suggesting a role in conidiogenesis. The expression of PdigCHSI, PdigCHSIII, PdigCHSV, and PdigCHSVII was upregulated during infection of citrus fruit. PdigCHSV and PdigCHSVII coexpressed in most of the experiments carried out, and they are separated by a 1.77 kb intergenic region and arranged in opposite directions. The PDE01641 deletion mutant showed enhanced cellobiohydrolase activity and β- glucosidase activity compared to the parental strain 114-2 (Zhang et al,. 2012). Increased transcription of the main cellulase and hemicellulase genes in ΔPDE01641 gave evidence that PDE01641 might affect the process associated with the regulation of cellulolytic enzymes expression. Furthermore, the deletion of PDE01641 from the genome of hypercellulolytic industrial strain JU-A10-T resulted in 36% and 80% increase in cellulase activity and hemicellulase activity respectively. These results revealed that PDE01641 plays an important role in the regulation of cellulolytic enzyme production in P. decumbens, and the engineering strain constructed in this work could be potentially used in bioenergy production.

The ochratoxin A (OTA) polyketide synthase otapks gene has been cloned from Penicillium verrucosum by Abbas et al. (2013). A P. verrucosum mutant in which the otapksPV gene has been interrupted cannot synthesize ochratoxin A. The protein is most similar to the citrinin polyketide synthase CtnpksMa from Monascus anka (83% identity at the amino acid level). Different nutritional conditions influence OTA production in P. verrucosum, with the addition of glycerol and galactose to MCB resulting in approximately 19 and 32 fold increases in OTA production respectively. These effects are mirrored in increased levels of otapksPV gene transcription. In contrast, the addition of glucose to MCB containing galactose results in an approximate 10 fold repression in OTA production, with this repression again being mirrored in decreased levels of otapksPV gene transcription. Thus the effects of different carbon sources on OTA production in P. verucosum appear to be regulated at the level of gene transcription. Two additional open reading frames, otaE and otaT, were identified in the 5' and 3' flanking regions of otapksPV, respectively. The otaT and otaE genes are coexpressed with P. verrucosum otapksPv, indicating a possible role for these genes in OTA biosynthesis. Furthermore, otaT and otaE were identified as putative homologues of the M. anka citrinin transporter ctnC (72% amino acid identity) and M. anka citrinin oxidoreductase ctnB (83% amino acid identity); suggesting that the genes involved in OTA production in P. verrucosum may be very similar to those involved in citrinin production in M. anka. A gene that encodes a carboxylesterase (carb) in Penicillium expansum GF was cloned, sequenced and overexpressed byPenicillium griseoroseum PG63, and the enzyme was characterized (Corrêa et al., 2013). The recombinant strain, P. griseoroseum T55, obtained upon transformation using the plasmid pAN-52-1-carb, showed integration of the carb gene into at least two heterologous sites of the genome by Southern blotting. Furthermore, the recombinant strain T55 exhibited almost a fourfold increase in carboxylesterase activity compared with PG63 strain when both were cultured without inducers. Based on the secondary structure and multiple sequence alignments with carboxylesterases, cholinesterase and lipase, a three- dimensional model was obtained. The α/β barrel topology, that is typical of esterases and lipases, was indicated for the CARB protein with Ser(213)-Glu(341)-His(456) as the putative catalytic triad. CARB preferentially hydrolysed acyl chains with eight carbon atoms, and its activity was optimal at a pH of 7·0 and a temperature of 25°C. CARB exhibited stability in alkaline pH, high activity under mesophilic conditions and stability in organic solvents. Teixeira et al. (2013) reported the inactivation of the pgg2 gene, a polygalacturonase- encoding gene from Penicillium griseoroseum, reduced the total activity of polygalacturonase (PG) by 90 % in wild-type P. griseoroseum, which indicates that the pgg2 gene is the major gene responsible for PG production in this species. To increase

PG production, the coding region of the pgg2 gene was cloned under the control of the glyceraldehyde 3-phosphate dehydrogenase (gpd) promoter and the terminator region of the tryptophan synthase (trpC) gene from Aspergillus nidulans (pAN52pgg2 vector). This vector was then used to transform P. griseoroseum. The transformed strains were characterized according to PG production using glucose, sucrose, or sugar cane juice as the carbon sources. The recombinant P. griseoroseum T146 strain contained an additional copy of the pgg2 gene, which resulted in a 12-fold increase in PG activity when compared with that detected in the supernatant of the control PG63 strain. The proteins secreted by the recombinant strain T146 showed a strong band at 38 kDa, which corresponds to the molecular weight of PG of the P. griseoroseum. The results demonstrate the significant biotechnological potential of recombinant P. griseoroseum T146 for use in PG production. Eighteen cellulases, including three cellobiohydrolases (CBHs), eleven endo-β-1,4- glucanases (EGs) and four cellulose monooxygenases (CMOs [32], previously known as glycosyl hydrolase (GH) family 61 EGs) were predicted to be encoded in the P. decumbens genome.(Liu et al., 2013) Liu et al. (2013) characterized an endo-acting cellulase PdCel5C from industrial cellulase-producing fungus Penicillium decumbens with distinctive domain composition. In addition to a cellulose-binding domain and a catalytic domain, PdCel5C contains two immunoglobulin (Ig)-like domains near the C-terminal end. Truncated mutation experiment reveals that the two Ig-like domains are important for the hydrolytic activity of PdCel5C. Moreover, PdCel5C releases cello-oligosaccharides from cellulosic substrates, which is different from that of most characterized cellulases in the same glycoside hydrolase family 5. To the best of our knowledge, this is the first report on the characterization of an Ig-like domain-containing cellulase in fungi. Wang et al. (2013) cloned an endoxylanase gene (PoxynA) that belongs to the glycoside hydrolase (GH) family 11 from a xylanolytic strain, Penicillium oxalicum B3-11(2). PoxynA was overexpressed in Trichoderma reesei QM9414 by using a constitutive strong promoter of the encoding pyruvate decarboxylase (pdc). The high extracellular xylanase activities in the fermentation liquid of the transformants were maintained 29~35-fold higher compared with the wild strain. The recombinant POXYNA was purified to homogeneity, and its characters were analyzed. Its optimal temperature and pH value were 50 degrees C and 5.0, respectively. The enzyme was stable at a pH range of 2.0 to 7.0. Using beechwood as the substrate, POXYNA had a high specific activity of 1,856 +/- 53.5 IU/mg. In the presence of metal ions, such as Cu2+, and Mg2+, the activity of the enzyme increased. However, strong inhibition of the enzyme activity was observed in the presence of Mn2+ and Fe2+. The recombinant POXYNA hydrolyzed birchwood xylan, beechwood xylan, and oat spelt xylan to produce short-chain xylooligosaccharides, xylopentaose, xylotriose, and xylobiose as

87 the main products. This is the first report on the expression properties of a recombinant endoxylanase gene from Penicillium oxalicum. The properties of this endoxylanase make it promising for applications in the food and feed industries.

Zhang et al. (2013) isolated the glyceraldehyde-3-phosphate dehydrogenase gene (PeGPD) from P. expansum PE-12 through reverse transcriptase PCR and 5'-3' rapid amplification of cDNA ends (RACE-PCR). The gene is 1266 bp long, including an ORF of 1014 bp, encoding a polypeptide chain of 337 amino acids. A phylogenetic tree based on GPD proteins showed that P. expansum is close to Aspergillus species, but comparatively distant from P. marneffei. Southern blot results revealed a single copy of PeGPD, and expression analysis gave evidence of high expression levels. PeGPD genes have potential for genetic engineering of P. expansum for industrial lipase production.

6.9. Molecular identification Phylogenetic analysis of the most important penicillin producing P. chrysogenum isolates carried out by Houbraken et al. (2011) revealed the presence of two highly supported clades, representing two species, P. chrysogenum and P. rubens. These species are phenotypically similar, but extrolite analysis showed that P. chrysogenum produces secalonic acid D and F and/or a metabolite related to lumpidin, while P. rubens does not produce these metabolites. Fleming's original penicillin producing strain and the full genome sequenced strain of P. chrysogenum were re-identified as P. rubens. Sang et al. (2013) used multigene phylogenetic analyses with the nuclear ribosomal internal transcribed spacer (ITS) region and genes encoding β-tubulin (benA) and calmodulin (cmd), as well as morphological analyses to identify two isolates as members of the P. sclerotiorum complex in Penicillium subgenus Aspergilloides, but different from species of the P. sclerotiorum complex. The isolates were found to be closely related to P. cainii, P. jacksonii, and P. viticola in terms of their multigene phylogeny, but their colony and conidiophore morphologies differ from those of closely related species. The name P. daejeonium was proposed for this unclassified new species belonging to the P. sclerotiorum complex in subgenus Aspergilloides.

7. Industral applications of Penicillium species

7.1. Penicillin

History of discovery of penicillin

1928, Scottish biologist Sir Alexander Fleming noticed a halo of inhibition of bacterial growth around a contaminant blue-green mould on a Staphylococcus plate culture. He concluded that the mould was releasing a substance that capable of killing a wide range of harmful bacteria, such as Streptococcus, Meningococcus and the diphtheria bacillus. He grew a pure culture of the mould and discovered that it was Penicillium notatum. With help from a chemist, he concentrated what he later named "penicillin".

sir_alexanderfleming.jpeg www.thestar.com ouponcw.com

1929, Fleming published his findings in the British Journal of Experimental Pathology, with only a passing reference to penicillin's potential therapeutic benefits.

1938, Florey came across Fleming’s paper on the penicillium mould in The British Journal of Experimental Pathology. Soon after, Florey and his colleagues decided to unravel the science beneath what Fleming called penicillium’s ”antibacterial action.”

1939, Howard Florey, Ernst Chain and their colleagues at Oxford University began growing the Penicillium in a strange array of culture vessels such as baths, bedpans, milk churns and food tins. Later, a customized fermentation vessel was designed for ease of removing and, to save space, renewing the broth beneath the surface of the

89 mould. A team of "penicillin girls" was employed, at £2 a week, to inoculate and generally look after the fermentation. In effect, the Oxford laboratory was being turned into a penicillin factory.

Howard Florey, Ernst Chain en.wikipedia.org Early penicillin culture facility , Museum of the History of Science, Oxford

Norman Heatley, a biochemist, extracted penicillin from huge volumes of filtrate coming off the production line by extracting it into amyl acetate and then back into water, using a countercurrent system.

Edward Abraham, another biochemist who was employed to help step up production, then used the newly discovered technique of alumina column chromatography to remove impurities from the penicillin prior to clinical trials.

Norman Heatley Edward Abraham Andrew Moyer

1940, Florey and his colleagues infected a group of 50 mice with deadly streptococcus. Half the mice died miserable deaths from overwhelming sepsis. The others, which received penicillin injections, survived.

1940, Albert Alexander, became the first recipient of the Oxford penicillin. He had a life-threatening infection with huge abscesses affecting his eyes, face, and lungs. Penicillin was injected and within days he made a remarkable recovery. But supplies of the drug ran out and he died a few days later. Better results followed with other patients though and soon there were plans to make penicillin available for British troops on the battlefield.

1941, Florey and Heatley traveled to the United States. They were referred to Robert Thom, the famous mycologist and authority on the Penicillium mould, and Orville May, Director of the NRRL, who agreed to have the Laboratory undertake a vigorous program to increase penicillin yields under the direction of Robert Coghill, Chief of the Fermentation Division.

Andrew Moyer found that he could significantly increase the yield of penicillin by substituting lactose for the sucrose used by the Oxford team in their culture medium. Shortly thereafter, Moyer made the even more important discovery that the addition of corn-steep liquor to the fermentation medium produced a ten-fold increase in yield.

Kenneth Raper, staff at the NRRL screened various Penicillium strains and found a strain that came from a moldy cantaloupe from a Peoria fruit market that produced acceptable yields of penicillin in submerged culture. When this strain was exposed to ultraviolet radiation at the University of Wisconsin, its productivity was increased still further.

1942 enough penicillin had been produced under the Office of Scientific Research and Development (OSRD) auspices to treat the first patient (Mrs. Ann Miller, in New Haven, Connecticut); a further ten cases were

1943, the War Production Board (WPB) took responsibility for increased production of the drug. The WPB investigated more than 175 companies before selecting 21 to participate in a penicillin program under the direction of Albert Elder; in addition to Lederle, Merck, Pfizer and Squibb, Abbott Laboratories

1944, The United States Army established the value of penicillin in the treatment of surgical and wound infections. Clinical studies also demonstrated its effectiveness against syphilis, it was the primary treatment for this disease in the armed forces of Britain and the United States.

1944, penicillin production began to increase dramatically in the United States it jumped from 21 billion units in 1943, to 1,663 billion units in 1944, to more than 6.8

91 trillion units in 1945, and manufacturing techniques had changed in scale and sophistication from one-liter flasks with less than 1% yield to 10,000-gallon tanks at 80- 90% yield. The American government was eventually able to remove all restrictions on its availability

Photograph courtesy of Merck Archives, ©Merck & Co. Inc. Fermentation unit used in purifying

penicillin in 1945.

1945, penicillin was distributed through the usual channels and was available to the consumer in his or her corner pharmacy.

1945, the unique feature of the structure was finally established, namely the four- membered highly labile beta-lactam ring, fused to a thiazolidine ring.

1945, Alexander Fleming, Howard Florey, and Ernst Chain were awarded the Nobel Prize for their penicillin research.

1949, the annual production of penicillin in the United States was 133,229 billion units, and the price had dropped from twenty dollars per 100,000 units in 1943 to less than ten cents.

Alexander Fleming receiving his Nobel Prize in 1945 British Library Add. MS 56115, f.81

Fleming's penicillin producing strain is not Penicillium chrysogenum but P. rubens. Houbraken J, Frisvad JC, Samson RA. IMA Fungus. 2011 Jun;2(1):87-95.

 As reported by Houbraken et al. (2011), Penicillium chrysogenum was described by Thom (1910), but the name P. chrysogenum is predated by P. griseoroseum, P. citreoroseum, and P. brunneorubrum, which were described by Dierckx (1901).  Since P. chrysogenum is a widely used name, Frisvad et al. (1990) and Kozakiewicz et al. (1992) suggested to preserve the name P. chrysogenum and to reject the older name P. griseoroseum with its synonyms P. citreoroseum, and P. brunneorubrum.  This proposal was accepted by the Committee for Fungi and Lichens and the name P. chrysogenum is currently listed as nomen conservandum

 Phylogenetic analysis of P. chrysogenum isolates revealed the presence of two highly supported clades, which represent two species, P. chrysogenum and P. rubens. These species are phenotypically similar, but extrolite analysis shows that P. chrysogenum produces secalonic acid D and F and/or a metabolite related to lumpidin, while P. rubens does not produce these metabolites. Fleming's original

penicillin producing strain and the full genome sequenced strain of P. chrysogenum are re-identified as P. rubens.

openi.nlm.nih.gov Penicillium rubens (Fleming's original penicillin-producer). Houbraken J, Frisvad JC, Samson RA - (2011)

Penicillium chrysogenum, Mycobank

7. 2. Griseofulvin

Synonyms: Amudane, Biogrisin-fp, Delmofulvina, Fulcin, Fulcine, Fulvina, Fulvinil, Fulvistatin, Fungivin, Greosin Gresfeed, Gricin, Grifulin, Grifulvin, Grisactin, Griscofulvin, Grisefuline, Griseo, Griseofulvin, Griseomix, Grisetin, Grisofulvin, Grisovin, Grizeofulvin, Grysio, Guservin, Lamoryl, Likuden, Likunden, Murfulvin, Neocid, Nufvlin, Poncyl, Spirofulvin, Sporostatin, Xuanjin

Griseofulvin is an antifungal antibiotyic produced by several Penicillium species, namely P. aethiopicum, P. coprophilum , P. dipodomyicola , P. griseofulvum, P. persicinum and P. sclerotigenum. It is fungistatic and acts only against dermatophytes.

P. griseofulvum.www.dehs.umn.edu www.fermentek.co.il. P. scerotiorum synapse.koreamed.org

P. aethiopicum Mycobank P. coprophilum

History of griseofulvin discovery

1939, Oxfordd, Raistrick and Simonart isolated a metabolic product of Penicillium griseofulvum.

1946, Brian Curtis and Hemming isolated an antibiotic from Penicillium janczewskii, which they called “curling factor”

1947, Grove and McGowan showed the curling factor was identical with griseofulvin

1949, Brian showed griseofulvin to be fungistatic to most mycelial fungi

1951, Brian, Wright, Stubbs and Way demonstrated systemic antifungal activity in plants

1957-1958, Williams and Peterkin reported that griseofulvin failed when applied topically to ringworm lesions

1958, Tomich proved that griseofulvin shoed a remarkable absence of acute toxicity, when administered orally to animals, even in very high dosage

1958, James C. Gentles showed in a prelilinary report that oral griseofulvin rapidly cleared artificially induced ringworm in guinea-pigs

1959, Gustav Riehl in Vienna treated the first human case of ringworm by griseofulvin orally

7. 3. Roquefort cheese

Roquefort is a sheep milk blue cheese from the south of France, and together with Bleu d'Auvergne, Stilton, and Gorgonzola is one of the world's best known blue cheeses. Though similar cheeses are produced elsewhere, EU law dictates that only those cheeses aged in the natural Combalou caves of Roquefort-sur-Soulzon may bear the name Roquefort, as it is a recognised geographical indication, or has a protected designation of origin. The cheese is white, tangy, crumbly and slightly moist, with distinctive veins of green mold. It has characteristic odor and flavor with a notable taste of butyric acid; the green veins provide a sharp tang. The overall flavor sensation begins slightly mild, then waxes sweet, then smoky, and fades to a salty finish. It has no rind; the exterior is edible and slightly salty. A typical wheel of Roquefort weighs between 2.5 and 3 kilograms (5.5 and 6.6 pounds), and is about 10 cm (4 inches) thick. Each kilogram of finished cheese requires about 4.5 litres (1.2 US gal) of milk to produce.

Roquefort is still being made exclusively from sheep's milk in caves that tunnel four miles into Mount Combalou in the Massif Central of south-central France. The village of Roquefort (it means ''strong rock''), perched on the mountain, has given its name to the cheese that must be aged in these caves, sections of which often figured importantly in the doweries of young women of the region. By law, only this cheese can be called Roquefort. The name is so protected that if a product such as a dip or a salad dressing uses the word Roquefort in its name, it must contain genuine Roquefort cheese.

A Lacaune flock in France imgarcade.com. Roquefort Village A Lacaune flock in France

Roquefort Caves- Cantobre. www.cantobre-france.eu Close to Milau are the Roquefort Caves; an extraordinary geological feature, formed by nature after the landslide of the Combalou mountain.

History of Roquefort cheese  According to legend (Fabricant, Florence, 1982), Roquefort first occurred when a lovestruck gaulois shepherd left his lunch and bread at the mouth of a cave while he went chasing a shepherdess. When he returned to the spot three months later he found the bread covered with mold, some of which had infected the cheese. the mold (Penicillium roqueforti) had transformed his plain cheese into Roquefort. He must have like what he tasted.

 It is claimed that Roquefort, or a similar cheese, is mentioned in literature as far back as AD 79, when Pliny the Elder remarked upon its rich flavor. In 1411, Charles VI granted a monopoly for the ripening of the cheese to the people of Roquefort-sur-Soulzon as they had been doing for centuries.

 Cheesemaking colanders have been discovered amongst the region's prehistoric relics.[

 In 1925, the cheese was the recipient of France's first Appellation d'Origine Contrôlée (AOC) when regulations controlling its production and naming were first defined.

AOC means that any product labelled AOC is guaranteed to come from a specific geographic region, often very strictly determined. For example, AOC Roquefort cheese can only come from the town Roquefort-sur-Soulzon. More precisely, it can only be produced on a 1.5 mile x 900 foot plot of land in Roquefort-sur-Soulzon.

 In 1961, in a landmark ruling that removed imitation, the Tribunal de Grande Instance at Millau decreed that, although the method for the manufacture of the cheese could be followed across the south of France, only those whose ripening occurred in the natural caves of Mont Combalou in Roquefort-sur-Soulzon were permitted to bear the name Roquefort.

 Roquefort is made entirely from the milk of the Lacaune, Manech and Basco- Béarnaise breeds of sheep. Prior to theAOC regulations of 1925, a small amount of cow’s or goat’s milk was sometimes added. A total of around 4.5 litres of milk is required to make one kilogram of Roquefort. The mould that gives Roquefort its distinctive character (Penicillium roqueforti) is found in the soil of the local caves. Traditionally, the cheesemakers extracted it by leaving bread in the caves for six to eight weeks until it was consumed by the mould. The interior of the bread was then dried to produce a powder.

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In modern times, the mould can be grown in a laboratory, which allows for greater consistency. The mould may either be added to the curd or introduced as an aerosol through holes poked in the rind.

P. roqueforti grown on bread to seed milk, makeitwithme-kell.blogspot.com P. roqueforti colonies on agar

Penicillium roqueforti. esmisab.univ-brest.fr atlas.microumftgm

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Penicillium roqueforti toxins

Vallone et al. (2014) concluded that In order to reduce the presence of secondary metabolites in blue cheeses, one of the methods suggested by several authors is the use of strain with low productive ability (Engel and Teuber, 1989; Moss, 1992; Lafont et al., 1990; Pose et al., 2007). Buzilova et al. (2000) have studied the effect of mutation on synthesis of alkaloid by Penicillium roquefortii VKM F-141. The study has shown that the Penicilli mutants were unable to synthesise alkaloids or they changed the alkaloid composition. Puel et al. (2007) have shown that the ability to produce patulin from Byssochlamys fulva is related to presence/absence of two genes. Hidalgo et al. (2014) have cloned and sequenced a four gene cluster that includes the ari1 gene from P. roqueforti. Gene silencing of each of the four genes (named prx1 to prx4) resulted in a reduction of 65-75% in the production of PR-toxin indicating that the four genes encode enzymes involved in PR-toxin biosynthesis. Interestingly the four silenced mutants overproduce large amounts of mycophenolic acid, an antitumor compound formed by an unrelated pathway suggesting a cross-talk of PR-toxin and mycophenolic acid production.

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7. 4. Camembert cheese

History of Camembert cheese

1791, Marie Harel, a farmer from Normandy, was the first to make Camembert, following advice from a priest who came from Brie.

Marie Harel, the inventor of the Camembert, has in fact made her first ladled cheese in 1791 - See more at: http://fromageriegillot.fr/en/accueil/secrets-daop/#sthash.ejr8hHqx.dpuf fromageriegillot.fr

www.travelandlifestylediaries.com Normandy cow/ Normandy village parisbymouth.com

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1890, an engineer, M. Ridel, devised the wooden box which was used to carry the cheese and helped to send it for longer distances, in particular to America, where it became very popular. These boxes are still used today.

depositphotos.com French cheese of Camembert in t wood box, NikaNovak Before fungi were understood, the colour of camembert rind was a matter of chance, most commonly blue-grey, with brown spots.

P. cememberti www.dgfm-ev.de www.schimmelpilze.de From the early 20th century onwards, the rind has been more commonly pure white, but it was not until the mid-1970s that pure white became standard. 1983, the AOC variety "Camembert de Normandie" is required by law to be made only from raw, unpasteurized milk from "Vaches Normandes" cows. 1992 ,The variety named "Camembert de Normandie" was granted a protected designation of origin

1909, M. Vignoboule created the "Syndicat des Fabricants du Véritable Camembert de Normandie" (Genuine Camembert of Normandy Makers Syndicate). M. Vignoboule defined precisely that was a Norman camembert and how it should be made....

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After World War II, new changes occured. The production of camembert, that was still realized by more than 1400 dairies spread in the whole Auge country, had to evolve to adapt itself to the increase of the consumption as well as the struggle to survive to the new competition. The first giant production factories were built and the first robot to make camembert was successfully tested.

1968, the producers of Camembert cheese have obtained a "Label rouge", a french quality label

1983 the camembert received the "Appellation d'Origine Controlée" (AOC), a special brand given by the french government, to the producers who respect a high number of recommendations. This label was created to protect the original flavour of the camembert.

Later it also received an AOP "Appellation d'Origine Protégée" a special brand given by the european community.

Finaly a ''Syndicat interprofessionnel de défense de l'Appelation d'Origine Controlée" was created to promote that AOC. The future of the camembert cheese in Normandy is still rather uncertain.Some of producers have decided to modify the standard way of producing to increase their productivity. For example, they have installed moulding robots, they treat the crude milk and even want to redefine the notion of crude milk. The others go on using crude

105 milk (That imply to use costfull tests to ensure the quality and the safety of the milk), they keep on moulding manually the cheeses, and they want to preserve the quality and the diversity of the milk by using the traditionnal norman cows

The stele dedicated to Marie Harel: It was built in 1926. It is a monument to the glory of Camembert. Camembert Museum!

Founded in 1986, the Museum of the Camembert was born of an exhibition bringing together objects and documents related to the history and manufacture of Camembertcheese.The Museum of Vimoutiers Camembert is the one and only in France. It’s name is registered and is managed by an association of volunteers.

musee-du-camembert.jpg www.tripadvisor.com

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Through an amazing collection of boxes of Camembert labels, we can reconstruct the history, places of manufacture and the consumption pattern of this prestigious cheese.

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7. 5. Gorgonzola cheese

Gorgonzola cheese is an uncooked cheese made from unskimmed cow's and/or goat's milk. The milk is added to Lactobacillus bulgaricus and Streptococcus thermophilus bacteria along with spores of the mold Penicillium glaucum or Penicillium roqueforti.

Gorgonzola originated in the town of the same name near Milan, Italy in the 8th century. Prior to being called Gorgonzola, the cheese was generically called "green stracchino".

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By 1970, a Consortium for the protection of Gorgonzola cheese was created in order to protect and oversee the production of Gorgonzola.

In 1996, Gorgonzola was granted PDO (Designation of Protected Origin) status, which governs how and where Gorgonzola is produced in either the Piedmont or Lombardy regions of Italy.

Today, it is mainly produced in the northern Italian regions of Piedmont and Lombardy. Whole cow's milk is used, to which starter bacteria is added, along with spores of the mold Penicillium glaucum. Penicillium roqueforti, used in Roquefort cheese, may also be used. The whey is then removed during curdling, and the result aged at low temperatures. During the aging process metal rods are quickly inserted and removed, creating air channels that allow the mold spores to grow into hyphae and cause the cheese's characteristic veining.

Gorgonzola is typically aged for three to four months. The length of the aging process determines the consistency of the cheese, which gets firmer as it ripens. There are two varieties of Gorgonzola, which differ mainly in their age: Gorgonzola Dolce (also called Sweet Gorgonzola) and Gorgonzola Piccante (also called Gorgonzola Naturale, Gorgonzola Montagna, or Mountain Gorgonzola). Under Italian law, Gorgonzola enjoys Protected Geographical Status. Termed DOP in Italy, this means that it can only be produced in the provinces of Novara, Bergamo, Brescia, Como, Cremona, Cuneo, Lecco, Lodi, Milan, Pavia, Vare se,Verbano-Cusio-Ossola and Vercelli, as well as a number of comuni in the area of Casale Monferrato (province of Alessandria).

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7. 6. Cambozola cheese

Cambozola is a cow's milk cheese that is a combination of a French soft-ripened triple cream cheese and ItalianGorgonzola.. It was patented and industrially produced for the world market by the large German company Champignon in the 1970s. The cheese was invented in around 1900 and is still produced by Champignon. In English-speaking countries, Cambozola is often marketed as blue brie. It is made from a combination of Penicillium camemberti and the same blue Penicillium roqueforti mold used to makeGorgonzola, Roquefort, and Stilton. Extra cream is added to the milk, giving Cambozola a rich consistency characteristic of triple crèmes, while the edible bloomy rind is similar to that of Camembert. Cambozola is considerably milder than Gorgonzola and features a smooth, creamy texture with a subdued blue flavour.

7. 7. Penicillium-cured salami Penicillium nalgiovense is a close relative of Penicillium chrysogenum. Similar to other fungi such as Penicillium roqueforti or Penicillium camemberti, the mould Penicillium nalgiovense is used for refining of foods. In particular, for the maturation of certain varieties of salami, Penicillium nalgiovense is important and is applied as a so-called starter culture on the outer skin of the sausages.

Originally this Penicillium was isolated as nalgiovense food destroyer of cheese. Only later, its beneficial properties for flavor formation in salami was realized. In addition, A

110 major metabolic product of Penicillium nalgiovense is penicillin. However, only those strains are used for foods which do not produce penicillin, because the intake of penicillin in some individuals resulting in allergic reactions.

The surface coverage of certain dry fermented sausages such as Italian salami by some species of Penicillium provides their characteristic flavour and other beneficial properties. One of them is the protective effect by means of a uniform film of white mould against undesirable microorganisms.

www.shutterstock.com www.pinterest.com

www.grillsportverein.de\. The surface growth of P. nalgiovense suppresses the growth of other undesirable organisms such as indigenous molds, yeasts and bacteria.. lpoli.50webs.com

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www.wedlinydomowe.pl www.wpthm.com Cured Italian Salami In a recent paper published in the International Journal of Food Microbiology, a group of scientists from Italy, the Netherlands, Denmark, and Slovenia described a new species of the fungus Penicillium from an Italian salami. The fungus that most commonly colonizes salami is Penicillium nalgiovense, a mould that makes the white fluffiness associated with salami. Spores of mould are applied to the surface of the salami right after the meat has been fermented. The fungus rapidly colonizes the surface and prevents contaminating molds from growing and spoiling the salami.

Fungi have an important role in the production of dry-cured meat products, especially during the seasoning period. In general, both industrially and handmade salami are quickly colonized by a composite mycobiota during seasoning, often with a strong predominance of Penicillium species. These species are involved in the improvement of the characteristics and taste, and in the prevention of the growth of pathogenic, toxigenic or spoilage fungi.

As mentioned by Perrone et al. (2015), two Penicillium species were predominantly present on the salami surface and in the air of the seasoning and storage areas of a salami plant (Calabria, Italy). One species was identified as Penicillium nalgiovense, and the other was related to, but distinct from, Penicillium olsonii. Further molecular and biochemical analyses showed that this strain has high homology with the not yet described species named “Penicillium milanense” isolated in Denmark and Slovenia on cured meats. The taxonomic position of these strains in Penicillium was investigated using calmodulin, β tubulin and ITS sequences, phenotypic characters and extrolite patterns, and resulted in the discovery of a new Penicillium species, described here as P. salamii. A literature search showed that this species occurs on (cured) meat products worldwide. In our study, P. salamii predominated the salami and capocollo surface in levels similar to the commonly known starter culture P. nalgiovense, irrespective of the room or age of seasoning. Preliminary inoculation trials with P. salamii showed that it

112 was able to colonize salami during seasoning, indicating that this species could be used as a fungal starter for dry-cured meat.

Penicillium nalgiovense Mycobank

Penicillium olsonii Mycobank

7. 8. Penicillium pigments

Natural colorants are considered to be safer than synthetic ones, and their applications in foods, cosmetics and pharmaceuticals are growing rapidly (Lauro, 1991). There are a number of natural pigments, but only a few are available in sufficient quantities for industrial production. Production of pigments from microorganisms is advantageous over other sources because microorganisms can grow rapidly which may lead to a high productivity of the product (Kim et al., 1999). Monascus red pigments are of polyketide origin and are used commercially in the orient as non-toxic colorants for colouring rice wine, "Koji", soybean, cheese and red meat (Hajjai et al., 1999). The red pigments have

113 attracted worldwide commercial interest , but little information is available on the production of this pigment by other microbial sources. It has been reported that Penicillium may be the potential candidate to produce polyketide structure compounds (Jûzlová et al., 1996). Pigments are also used for dyeing of cotton yarn and. wood spalting

Red pigments It has been already reported by Curtin et al. (1940) that Penicillium phoeniceum, produced a red pigment, phoenicine, which, being watersoluble, diffused almost entirely into the medium. Wang et al. (2004) reported on a unique Penicillium isolate from Chinese soil with terverticillate penicilli and ellipsoidal to cylindrical smooth-walled conidia, produced, in addition to the common metabolite ergosterol, copious amounts of an unknown peach-red pigment . This isolate, CBS 111235, was described as Penicillium persicinum sp. nov., which belongs to subgenus Penicillium section Chrysogena but is morphologically similar to P. italicum. On the basis of the production of secondary metabolites it resembles P. griseofulvum and P. coprophilum.

Jiang et al. (2005) identified a new isolate from medicinal plant endophytes as Penicillium sp. (HKUCC 8070). A water-soluble red pigment was produced by this strain in potato-dextrose broth, maltextract broth and a chemically defined medium containing glutamate as a nitrogen source. The red pigment produced was identified as heat-stable, polyketide Monascus red pigment. The highest yield of the red pigment was 1107 mg l-1 obtained from the culture of Penicillium sp. (HKUCC 8070) grown on the malt-extract medium.

Alejandro Méndez et al., 2011, showed the feasibility of producing and obtaining natural water-soluble pigments by P. purpurogenum GH2 for potential use in food industries. A strong combined effect (p<0.05) of pH and temperature was associated with maximal red pigment production (2.46 g/L). Under optimized conditions, a 78% increase in red colourants production was achieved. The best pH and temperature conditions were obtained at pH 8.0 and 70°C, respectively. In the presence of salts NaCl and Na2 SO4 , both at concentrations of 0.1 and 0.5 M in Mcllvaine buffer (pH 8.0), the red colorants showed good stability. In the presence of both polymers polyethylene glycol and sodium polyacrylate, the red colorants kept their color intensity. A specific self-immobilization biomembrane-surface liquid culture (SIBSLC) was developed to overproduce a potential penicillium red pigment. Statistic analysis showed

114 that both glucose concentration and membrane diameter are important factors influencing the yield of red pigment. After the optimization using central composite experimental design, the maximum yield of red pigment in shake flask reached 4.25 g/l. The growth of strain HSD07B consists of three phases, and the pigment secreted in the decelerated phase, is originated from the interior of biomembrane where glucose exhaustion occurs. In addition, the batch and continuous SIBSLC were conducted for production of the pigment, and the latter was more competitive in consideration of the fact that it not only increased 61.5 % of pigment productivity, but also simplified the production process. (,Hailei et al., 2012 and Méndez et al., 2013)

Penicillium strain (DLR-7) producing red extracellular pigment was isolated from the mangrove soils of Andhra Pradesh, India (Lathadevi Karuna. 2014). A multifactorial and step-wise experiment was designed to study the physical and nutritional conditions that favor red pigment and biomass production. Potato extract prepared in the laboratory produced more pigment than the commercial potato dextrose broth and was therefore used as the basal medium. Culture conditions such as xylose 2% (w/v), glycine 1% (w/v), pH and temperature of 3.0 and 25 ˚C, were observed to be the optimal conditions producing 1050 mg/L of red pigment and 3.1 g/L of mycelia biomass. At pH 2.0, yellow fluorescent pigment was observed instead of red and spores were completely absent. Pigment was not produced when basic amino acids like arginine or lysine were supplemented to the medium, but acidic amino acids such as aspartic acid and glutamic acid enhanced pigment production. Also simple amino acids such as glucose maximized the growth of fungus whereas, amino acids such as xylose, mannose and glycine stimulated pigment yield.

Blue pigments Stodola et al. (1951) observed a curious change in pigmentation induced by variation of the substrate, a phenomenon which had been reported by Bainier and Sartory when the fungus was described in 1912. These workers observed that a yellow pigment was formed on potato, banana, carrot and Raulin's medium; on a medium rich in peptone the pigment was a beautiful blue. Apparently no attempt was made at chemical characterization. Several minor pigments from cultures of Penicillium herquei were described by Narasimhachari et al. (1963). One of these has been identified as the naphthalic anhydride which is formed when atrovenetin is oxidized with alkaline peroxide. Evidence was presented which suggested that the characteristic green pigment found in this fungus was formed by a reaction analogous to that between ninhydrin and amino derivatives. In P. herquei , the 1,2,3-trione is probably generated by mild oxidation of perinaphthenone pigments known to be produced by this organism. These

115 blue pigments were isolated and demonstrated by Robinson et al. (1992) to be a phenalenone derivative. As isolated, the major blue pigment is a zinc complex of elemental formula C76H64O20N2Zn in which two anions of the dimeric phenalenone 2 act as the tridentate ligands. Compounds 2 and the zinc complex have been identified previously as secondary metabolites of two other Ascomycetes. In addition, strong evidence was obtained for the presence in the CH2Cl2 extract of P. herquei of minor amounts of a novel compound, having a structure identical with that of the major blue pigment, but with nickel as the coordinating metal.

Brown pigments Vasanthakumar et al. (2015) isolated a strain of Penicillium chrysogenum from a tomb in Upper Egypt, which was capable of producing brown pigment in vitro when grown in a minimal salts medium containing tyrosine. They present ed evidence that this pigment is a pyomelanin, a compound that is known to assist in the survival of some microorganisms in adverse environments. They tested type strains of Penicillium chrysogenum, which were also able to produce this pigment under similar conditions. Inhibitors of the DHN and DOPA melanin pathways were unable to inhibit the formation of the pigment. FTIR analysis indicated that this brown pigment is similar to pyomelanin. Pyrolysis-GC/MS revealed the presence of phenolic compounds. Using LC/MS, homogentisic acid, the monomeric precursor of pyomelanin, was detected in supernatants of P. chrysogenum cultures growing in tyrosine medium but not in cultures lacking tyrosine. Partial regions of the genes encoding two enzymes in the homogentisic acid pathway of tyrosine degradation were amplified. Data from reverse-transcription PCR demonstrated that hmgA transcription was increased in cultures grown in tyrosine medium, suggesting that tyrosine induced the transcription. Yellow pigments Penicillum sclerotiorum can produce yellow pigments, which structurally are classified as azaphilones. The major constituent in those pigments is sclerotiorin. The main characteristic of azaphilones is its affinity to ammonia. The oxygen atom in azaphilone ring is likely to be substituted by NH group. Azaphilones are therefore called “nitrogen loving” Rugulosin is an intense yellow pigment produced by some species of Penicillium. Rugulosin shows antibacterial and insecticidal activity and has found application as a bioinsecticide, notably as the active secondary metabolite in endophytic fungi of seedlings. Rugulosin (+ form) is an inhibitor of RNA Polymerase and RNase.

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Luteoskyrin, a hepatotoxic pigment, produced by Penicillium islandicum Sopp. Various factors affecting the yields of luteoskyrin and related pigments in the liquid medium were studied. Maximal yields of luteoskyrin (0.13% by isolation) and of other pigments were attained in the late phase of the cultivation. The yield of the pigment was increased by supplying malt extract, malonic acid, glutamic acid, or asparagine. A useful material for preparation of 'IC-labeled luteoskyrin was 2-'4C-malonate. Penicillium chrysogenum IFL1 and IFL2, , and Penicillium vasconiae IFL4 were reported by Lopez et al. (2013) to large producers of water-soluble yellow pigments on agroindustrial residues.

The pigment production by P. chrysogenum was earlier reported. Until today there are few reports on pigments produced by these species, because the pigments were usually used as a chemotaxonomic tool. P. chrysogenum produces the yellow pigments sorbicillin and xanthocillins and chrysogenin was also reported as a yellow pigment produced by this fungus.

The pigment emodin is isolated from the strains of Penicillium citrinum and P. islandicum. As a first commercial product within this chemical family, the natural food colorant Arpink redTM (now Natural RedTM) is manufactured by the Czech company (Ascolor Biotech followed by Natural Red) and has been claimed to be produced by fermentation and bioprocess engineering using the fungal strain Penicillium oxalicum var. Armeniaca CCM 8242, a soil isolate (Fungal pigments for the food industry Dufosse´ et al. Current Opinion in Biotechnology 2014, 26:56–61)

Pigments producing Penicillium

P. atramentosum: Uncharacterized dark brown

P. atrosanguineum: Phoenicin (red)nnUncharacterized yellow and red P. atrovenetum: Atrovenetin (yellow), Norherqueinone (red) P. aurantiogriseum:Uncharacterized P. chrysogenum: Sorbicillins (yellow), Xanthocillins (yellow) P. brevicompactum :Xanthoepocin (yellow) P. citrinum Anthraquinones :(yellow), Citrinin (yellow) P. cyclopium : Viomellein (reddish-brown) ,Xanthomegnin(orange) P. discolor : Uncharacterized

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P. echinulatum: Uncharacterized (yellow)

P. flavigenum: Xanthocillins P. freii: Viomellein (reddish-brown), Vioxanthin, Xanthomegnin(orange) P. herquei :Atrovenetin (yellow), Herqueinones (red and yellow) P.oxalicum: Arpink red™- anthraquinone derivative (red), Secalonic acid D(yellow) P. paneum: Uncharacterized (red) P. persicinum: Uncharacterized (Cherry red) P. viridicatum: Viomellein (reddish-brown), Vioxanthin, Xanthomegnin (orange)

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7. 9. Penicillium enzymes Several Penicillium species produce enzymes that are used in industry 1. Cellulases and xylanases produced by Penicillium species have broad applications in food and feed, the textile industry, and the pulp and paper industries. 2. Cellulases and xylanases produced by Penicillium species are used for hydrolysis of cellulose and xylan, available in large quantities in lignocellulosic residues from agricultural, urban, and industrial solid waste. Syrups of glucose and xylose obtained from the hydrolysis of lignocellulosic biomass could be used for various biotechnological purposes, especially the production of ethanol 3. Plant growth-promoting fungi (PGPF) isolate, Penicillium spp. GP15-1 (Penicillium neoechinulatum or Penicillium viridicatum) stimulates growth and neoechinulatum or Penicillium viridicatum. 4. Peroxidase enzyme of Penicillium crysosporium has potential biodegradable activities that degrade Amaranth dye, Orange G, heterocyclic dyes like, Azure B and Lip dye. 5. Penicillium ochrochloron is used for biodegradation of triphenylmethane dye cotton blue, which is used extensively in the textile industries for dying cotton, wool, silk, nylon, etc. and is generally considered as the xenobiotic compound , which is very recalcitrant to biodegradation. 6. The ability of Penicillium sp. to tolerate oil pollutants and grow on them, suggest that it can be employed as bioremediation agent and can be used in restoring the ecosystem when contaminated by oil. 7. Penicillium species have a role in the removal & recovery of heavy metals from wastewater and industrial effluents. Hg, Cu, Ni, Pb, Cd are extracted at pH 2-5.

7. 10. Production of metal nanoparticles 1. The eco-friendly biosynthesis of gold nanoparticles by P. aurantiogriseum, P. citrinum, and P. waksmanii from a solution of AuCl. Gold nanoparticles are formed fairly uniform with spherical shape with the Z- average diameter of 153.3 nm, 172 nm and 160.1 nm for the 3 species, respectively. 2. Development of ecofriendly and reliable processes for the synthesis of silver nanoparticles by Penicillium species

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8. Health risks and economic losses caused by Penicillium species

 Penicillium is among the most prevalent airborne fungi. Its spores have been identified as potential respiratory allergens. It is a major cause of indoor mould allergy.  Penicillium spp. are commonly considered as contaminants but may cause infections, including pneumonia, particularly in immunocompromised hosts.  Some Penicillium spp. are known to produce mycotoxins. For example, P. verrucosum produces the mycotoxin, ochratoxin A, which is nephrotoxic and carcinogenic. The production of the toxin usually occurs in cereal grains at cold climates. Other mycotoxin-like compounds include patulin, citrinin, citroviridin, emodin, gliotoxin, verraculogen and secalonic acid D.  Infections are usually caused by inhalation of the spores. It starts as a pulmonary disease but may spread into the adjacent blood vessels. It disseminates to the rest of the body. Total invasion occurs in debilitated patients and can occur in immunocompetent (those with healthy immune systems) individuals as well.  Penicillium species causes keratitis (inflammation of the cornea), keratomycosis, penicilliosis, ostomycosis, onychomycosis (infection of the nail) and deep infections.  It has been known to cause external ear, respiratory, and urinary tract infections, and endocarditis after insertion of valve prostheses.

 Penicillium are major causative agents of food spoilage (dairy products, fruits, vegetables and meat) and postharvest decay.  The genus causes significant economic losses to the fruit .  Penicillium is also a huge problem in the wine industry. Its presence in wine and grape juice during the various stages of fermentation is highly detrimental to the quality of the wine due to the production of compounds such as Geosmin (trans- 1,10-dimethyl-trans-9-decalol), an earthy-musty compound which produces off odours and flavours.  Penicillium spp. prefer damp and dark places, but can occur elsewhere. Penicillium grows in dusty green colonies and dominates in temperate soils where spores are easily released into the atmosphere.  Penicillium spp. are widespread and are found in soil, decaying vegetation and compost, and the air, in particularly in temperate zones (forests, grassland and arable soils). It may be found in vineyards and wine cellars, the soil of citrus

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plantations, among all types of stored seeds, barns, damp hay, dried fruit and fruit juice.  Indoors it is the familiar blue-green mould found on stale bread, citrus fruits, apples and as a contaminant in rye flour in industrial bakeries.  The Penicillium species may also be found in foam rubber mattresses, house dust, stuffed furniture, wallpaper, books, refrigerator doors and rubber tubing.  Penicillium may cause the black spots on window-sills.  The Penicillium species can grow on different substrates, such as plants, cloth, leather, paper, wood, tree bark, cork, animal dung, carcasses, ink, syrup, seeds, and virtually any other item that is organic.  P. allii, P. hirsutum or P. viridicatum, are the pathogenic species responsible for garlic crop losses due to blue mould rot.  Penicillium expansum causes blue mould rot, a post-harvest disease of apples and pears. Other Penicillium species associated with Penicillium ear rot: P. chrysogenum, P. glaucum, P.cyclopium, P. italicum P. funiculosu., and P. digitatum are common causes of rot of citrus fruits.

 A large number of Penicillium species are mainly associated with food spoilage. Those covered here include , Penicillium chrysogenum, Penicillium aurantiogriseum, Penicillium digitatum, Penicillium griseofulvum, Penicillium italicum,Penicillium oxalicum and Penicillium viridicatum etc.  Penicillium citrinum is the dominant contaminant in cacao and coffee beans, and cassava in Indonesia.

Penicillium digitatum on lemon...www.insectimages.org Penicillium ulaiense on lemon...www.insectimages.org

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Moldy Lemon dict.space.4goo.net www.reddit.com

Penicillium expansum on pear - www.ipmimages.org

Mouldy Orange Two moldy oranges galleryhip.com mouldy strawberry imgkid.com

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Blue mold of citrus caused by a species of Penicillium oranges attacked by Penicillium italicum.

Three-pears-fresh-mouldywww.theguardian.com gillianscommsblog

www.snapthepix mouldy apples cherisephotography.blogspot.com

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Stock Photo of mouldy rotten old banana csp9766214 -...www.agric.wa.gov.au Penicillium is a secondary disease and characterised by berries covered in a mass of coloured spores

red grapes www.dreamstime.com www.21food.com SUN DRIED APRICOT Dried Figs cutcaster.com

Mouldy nuts. sst-web.tees.ac.uk www. Ars.usda.gov

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www.daff.qld.gov.au ...

. food-hacks.wonderhowto.com www.flickr.com

Mouldy Tomatowww.dreamstime.com flickrhivemind.net www.studyblue.com Blue mould rots – Penicillium gladioli and other Penicillium spp ...www.oakleafgardening.com

CW 09-10-23 Corn Ear Rots due to Penicillium -.cropwatch.unl.edu www.angelfire.com

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chicken littlecolumnist.blogspot.com www.snipview.com

Meat spoilage galleryhip.comSpoiled Steak Color

Green Mold on Dry-Cured Sausages forums.egullet.org waterinfood.com paninihappy.com

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Cakes contaminated with Penicillium, www.shroomery.org Mouldy bread www.wisegeek.com Mouldy sliced wholemeal bread with sunflower seedsyaymicro.com, news.vanderbilt.edu

www.mycolog.com spoki.tvnet.lv parasites.czu.cz cheddar cheese with various species of Penicillium, which can grow even in the refrigerator.

Penicillium funiculosum sur textile (coton) www.moldtesting-moldinspection-nj-newjersey.com Penicillin Mold Under A Cabinett in New Jersey. Penicillium colonizes leather ...

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www.nlm.nih.gov Mold on books: inspectapedia.com

Indoor Air Qulaity Blog

www.moldbacteriaconsulting.com www.blackmould.me.uk

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www.blackmould.me.uk

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9. Penicillium species identification

1. Morphological characteristics

2. Isoenzyme profiling

3. Identification of extrolites

4. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry fingerprinting (MALDI-TOF MS)

5. Genotypic characterization  Amplification of the internal transcribed spacers (ITS1 and ITS2)  Restriction fragment length polymorphism (RFLP)  Random amplified polymorphic DNA (RAPD)  Amplified fragment length polymorphism (AFLP).

DNA barcoding for identification and multilocus sequence typing for phylogenetic species recognition ( Visagie et al. 2014).

 Only recently however, was the internal transcribed spacer rDNA area (ITS) accepted as the official barcode for fungi (Schoch et al. 2012).

 ITS is the most widely sequenced marker for fungi, and universal primers are available (Schoch et al. 2012).  In Penicillium, it works well for placing strains into a species complex or one of the 25 sections, and sometimes provides a species identification ( Visagie et al. 2014).

Primers used for amplification and sequencing ( Visagie et al. 2014).

1. Internal Transcribed Spacer (ITS)

5′-3′) ITS1 Forward TCC GTA GGT GAA CCT GCG G White et al. 1990 ITS4 Reverse TCC TCC GCT TAT TGA TAT GC V9G Forward TTA CGT CCC TGC CCT TTG TA de Hoog et al., 1998

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LS266 Reverse GCA TTC CCA AAC AAC TCG ACT C Masclaux et al. 1995

2. β-tubulin (BenA)

Bt2a Forward GGT AAC CAA ATC GGT GCT GCT TTC Glass & Donaldson 1995 Bt2b Reverse ACC CTC AGT GTA GTG ACC CTT GGC

3. Calmodulin (CaM)

CMD5 Forward CCG AGT ACA AGG ARG CCT TC Hong et al. 2006 CMD6 Reverse CCG ATR GAG GTC ATR ACG TGG CF1 Forward GCC GAC TCT TTG ACY GAR GAR Peterson et al. 2005 CF4 Reverse TTT YTG CAT CAT RAG YTG GAC

4. RNA polymerase II second largest subunit (RPB2)

5F Forward GAY GAY MGW GAT CAY TTY GG Liu et al. 1999 7CR Reverse CCC ATR GCT TGY TTR CCC AT 5Feur Forward GAY GAY CGK GAY CAY TTC GG Houbraken et al. 2012

7CReur Reverse CCC ATR GCY TGY TTR CCC AT

Thermal cycle programs used for amplification.

Profile Initial Final Gene type denaturing Cycles Denaturing Annealing Elongation elongation Rest period General ITS,BenA, CaM standard 94 °C, 5 min 35 94 °C, 45 s 55 °C, 45 s 72 °C, 60 s 72 °C, 10 °C, ∞ 7 min General alternative standard 94 °C, 5 min 35 94 °C, 45 s 52 °C, 45 s 72 °C, 60 s 72 °C, 10 °C, ∞ 7 min RPB2 touch-up 94 °C, 5 min 5 94 °C, 45 s 50 °C, 45 s 72 °C, 60 s 5 94 °C, 45 s 52 °C, 45 s 72 °C, 60 s 30 94 °C, 45 s 55 °C, 45 s 72 °C, 60 s 72 °C, 10 °C, ∞ 7 min RPB2 alt. touch-up 94 °C, 5 min 5 94 °C, 45 s 48 °C, 45 s 72 °C, 60 s 5 94 °C, 45 s 50 °C, 45 s 72 °C, 60 s 30 94 °C, 45 s 52 °C, 45 s 72 °C, 60 s 72 °C, 10 °C, ∞ 7 min

Examples of recently successful application of the above methods

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1. As reported by Boysen et al. (2000) P. roqueforti was recently split into the three species, P. roqueforti, Penicillium carneum, and Penicillium paneum, (collectively referred to as the P. roqueforti group) based on ribosomal DNA sequence comparison, random amplified polymorphic DNA (RAPD) profiles, and secondary metabolite profiles. These three species synthesize different mycotoxins. All three produce roquefortine C, only P. roqueforti produces PR toxin, and both P. carneum and P. paneum produce patulin, which is mutagenic, immunotoxic, and neurotoxic (5, 8). PR toxin is the most acutely toxic metabolite produced, with 50% lethal dose values in mice ranging from 1 to 5.8 mg kg of body weight−1 (intraperitoneally [IP]) . The equivalent 50% lethal dose values for roquefortine C and patulin are 20 and 5 mg kg of body weight−1 (IP), respectively (5, 25). All three Penicillium species can grow on 0.5% acetic acid (2) and have similar microaerophilic capacity with regard to their ability to grow at low oxygen and high carbon dioxide pressures.

RAPD fingerprinting using either NS2 (a) or NS7 (b) as a primer. Lanes: 1, P. roqueforti SVA 3631/1996, having a T in position 180 of the ITS1 region; 2, P. roqueforti SVA 8176/1995, isolate 5 (A in position 180); 3, P. roquefortiSVA 2986/1992 (T in position 180); 4, P. roqueforti type strain (IBT 6754); 5, P. paneum SVA 494/1990; 6, P. paneum SVA 7023/1994, isolate 2; 7, P. paneumtype strain (IBT 12407); 8, P. expansum SVA 2294/1994, isolate 5; 9, P. expansumSVA 6180/1994 isolate 1; 10, P. carneum type strain (IBT 6884); M, molecular weight marker (1-kb DNA ladder; GIBCO BRL, Gaithersburg, Md.), Boysen et al. (2000) 2. As reported by Oliveri et al. (2007), 41 isolates of Penicillium spp. were recovered from rotten fruits (including oranges, grapefruits, pears, lemons, strawberries, apples, loquats, prickly pears) and from air and surfaces of markets and packing-houses. Penicillium isolates were identified as P expansum, R italicum, R digitatum, R olsonii, R chrysogenum or R citrinum. Genetic characterization was performed with ITS4 and ITS5 primers that specifically identified Penicillium isolates by amplification of a 600- bp fragment, with PEF and PER primers used to identify R expansum isolates by amplification of a 404-bp fragment, and with fluorescent amplified fragment length polymorphism analysis (fAFLP). Cluster analysis of fAFLP data divided the isolates

132 into five well-separated R italicum, R digitatum, R citrinum, R chrysogenum and R olsonii clusters, whereas R expansum isolates were divided in three distinct clusters. Within all the eight clusters, isolates were well differentiated. Results obtained with fAFLP analysis confirmed the reliability of the method to characterize and identify strains at intraspecific level.

3. As reported by Demirel et al. (2013), nine terverticillate Penicillium isolates (P. griseofulfum, P. puberulum, P. crustosum, P. aurantiogriseum, P. chrysogenum, P. primulinum, P. expansum, P. viridicatum, Eupenicillium egyptiacum) from 56 soil samples were characterized genetically by a PCR method. The DNAs of the strains were isolated using the glass beads and vortexing extraction method and then used for PCR amplification with the internal transcribed spacer 1 (ITS1) and ITS4 universal fungal specific primers. The ITS regions of fungal ribosomal DNA (rDNA) were sequenced through the CEQ 8000 Genetic Analysis System. ITS-5.8S sequences obtained were compared with those deposited in the GenBank Database. The results indicated that the identification of Penicillium species with PCR based methods provided significant information about the solution to taxonomy

Newly generated ITS sequences with their closest GenBank sequences according to Blast, Demirel et al. (2013),

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PCR products (570 bp) of different terverticillate Penicillium species using the universal fungal primers (ITS1/ITS4), PCR products (570 bp), Demirel et al. (2013),

4. As reported by You et al. (2014), during an investigation of the fungal diversity of Korean soils, four Penicillium strains could not be assigned to any described species. The strains formed monoverticillate conidiophores with occasionally a divaricate branch. The conidia were smooth or finely rough-walled, globose to broadly ellipsoidal and 2.5-3.5 × 2.0-3.0 µm in size. Their taxonomic novelty was determined using partial β-tubulin gene sequences and the ribosomal internal transcribed spacer region. The phylogenetic analysis showed that the isolates belonged to section Lanata-Divaricata and were most closely related to Penicillium raperi. Phenotypically, the strains differed from P. raperi in having longer and thicker stipes and thicker phialides. Strain KACC 47721T from bamboo field soil was designated as the type strain of the new species, and the species was named Penicillium koreense sp. nov., as it was isolated from various regions in Korea.

5. As reported by Park et al. (2015), three strains of an unidentified Penicillium species were isolated during a fungal diversity survey of marine environments in Korea. These strains were described as a new species following a multigene phylogenetic analyses of nuc rDNA internal transcribed spacer barcodes (ITS1-5.8S-ITS2), genes for β-tubulin, calmodulin and RNA polymerase II second largest subunit, and observation of macro-and micromorphological characters. Phylogenetic analyses revealed that the three strains formed a strongly supported monophyletic group distinct from previously reported species of section Aspergilloides. Morphologically this species can be

134 distinguished from its sister species, P. crocicola, by the reverse color on Czapek yeast autolysate agar, abundant production of sclerotia on malt extract agar and colony characters on yeast extract sucrose agar. They named this new species P. jejuense, after the locality where it was discovered.

Examples of Misidentifications in Penicillium

1. Phylogenetic analysis of P. chrysogenum isolates described by Thom in 1010, revealed the presence of two highly supported clades, which represent two species, P. chrysogenum and P. rubens. These species are phenotypically similar, but extrolite analysis shows that P. chrysogenum produces secalonic acid D and F and/or a metabolite related to lumpidin, while P. rubens does not produce these metabolites. Fleming's original penicillin producing strain and the full genome sequenced strain of P. chrysogenum are re-identified as P. rubens (Houbraken et al., 2011).

2. A fungus producing penicillones A and B and chloctanspirones A, B and terrestrols was identified as P. terrestre ( Liu et al. 2005). The name P. terrestre was used by Raper & Thom (1949), but has since been considered a synonym of P. crustosum. However, additional secondary metabolites produced by the isolate, such as sorbicillin and trichodimerol, are never seen in P. crustosum ( Frisvad et al. 2004), suggesting the strain was P. chrysogenum or P. rubens. Liu et al. (2005) did not describe how their isolate was identified.

3. A strain of P. chrysogenum or P. rubens was misidentified as a P. commune ( Shang et al., 2012 and Zhao et al., 2012), based on “100 % sequence identity” with P. commune.The production of chrysogine, sorbicillins and meleagrin indicated that the strain, SD-118, was in fact P. chrysogenum.

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10. Gallery of Penicillia

Penicillium frequentans Mycota Penicillium glabrum

Penicillium lividum Tzean, SS et al. 1994. Penicillium spinulosum Tzean, SS et al. 1994..

Penicillium corylophilum Tzean, SS et al. 1994. Penicillium decumbens Gerald Holmes, California

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Penicillium alexiae Visagie et al.,. 2013. Penicillium adametzioides , Jian Xin Denget al.,

Penicillium arianeae Visagie,et al.,. 2013. Penicillium amaliae Visagie, et al.,. 2013. ,

Penicillium herquei Tzean, SS et al. 1994 Penicillium cainii K.G. Rivera, et al., 2011

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Penicillium johnkrugii K.G. Rivera, et al.,. 2011. P. jacksonii K.G. Rivera, et al.,. 2011.

P. sclerotiorum K.G. Rivera and K.A. Seifert, 2011 Penicillium maximae Visagie, et al.,. 2013

P. vanoranjei Visagie, et al.,2013 P. viticola . Rivera and . Seifert Stud Mycol. Nov 15, 2011.

P. bialowiezense Won Ki Kim et al. Mycobiol. 2007 P. brevicompactum Tzean, SS et al. 1994.

. Penicillium janczewskii Tzean, SS et al. 1994. P. canescens Tzean, SS et al. 1994.

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Penicillium chrysogenum . Penicillium confertum

Penicillium dipodomyis Penicillium flavigenum

P. glycyrrhizacola Chen, Sun & Gao, 2013. Penicillium mononematosum

Penicillium nalgiovense Penicillium chermesinum Tzean, SS et al. 1994.

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Penicillium anatolicum Houbraken et al. (2011) Penicillium argentinense Houbraken et al. (2011)

Penicillium atrofulvum Houbraken et al. (2011) P. aurantiacobrunneum Houbraken et al. (2011)

Penicillium cairnsense Houbraken et al. (2011) Penicillium christenseniae Houbraken et al. (2011)

P. chrzaszczii Houbraken et al. (2011 Penicillium citrinum Tzean, SS et al. 1994

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P/ cosmopolitanum. Houbraken et al. (2011) Penicillium copticola Houbraken et al. (2011)

Penicillium decaturense Houbraken et al. (2011) Penicillium euglaucum. Houbraken et al. (2011)

Penicillium gallaicum Houbraken et al. (2011) Penicillium godlewskii. Houbraken et al. (2011)

P. gorlenkoanum Houbraken et al. (2011) Penicillium hetheringtonii. Houbraken et al. (2011)

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Penicillium manginii. Houbraken et al. (2011) Penicillium miczynskii Tzean, SS. et al. 1994

Penicillium neomiczynskii Houbraken et al. (2011) Penicillium nothofagi. Houbraken et al. (2011)

Penicillium pancosmium Houbraken et al. (2011) Penicillium pasqualense Houbraken et al. (2011)

Penicillium paxilli Tzean, SS et al. 1994. Penicillium quebecense Houbraken et al. (2011)

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Penicillium raphiae Houbraken et al. (2011) Penicillium roseopurpureum. Houbraken et al. (2011)

Penicillium sanguifluum. Houbraken et al. (2011) Penicillium shearii Houbraken et al. (2011)

Penicillium sizovae. Houbraken et al. (2011) Penicillium steckii. Houbraken et al. (2011)

Penicillium westlingii. Houbraken et al. (2011) Penicillium sumatrense. Houbraken et al. (2011)

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Penicillium terrigenum. Houbraken et al. (2011) Penicillium tropicoides. Houbraken et al. (2011)

Penicillium tropicum. Houbraken et al. (2011) Penicillium ubiquetum Houbraken et al. (2011)

Penicillium vancouverense. Houbraken et al. (2011) Penicillium waksmanii. Houbraken et al. (2011)

Penicillium wellingtonense Houbraken et al. (2011) Penicillium digitatum Tzean, SS et al. 1994.

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Penicillium dravuni Janso, . 2005. Penicillium melinii Tzean, SS et al. 1994.

P. albocoremium Won Ki Kim et al., 2006 Penicillium aurantiogriseum Tzean, SS et al. 1994

P. crustosum Tzean, SS et al. 1994 Penicillium echinulatum

P. solitum , Won Ki Kim et al. Mycobiology. 2007 P. tulipae Won Ki Kim Microbiol. 2006

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Penicillium daleae SS et al. 1994 Penicillium javanicum SS et al. 1994

P. rolfsii Tzean, SS et al. 1994. P. simplicissimum Tzean, SS et al. 1994.

Penicillium atramentosum Mycobank Penicillium clavigerum Mycobank

Penicillium concentricum Mycobank Penicillium coprobium Mycobank

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Penicillium coprophilum Mycobank Penicillium dipodomyicola (Mycobank

Penicillium expansum , 1994Tzean P. formosanum H.M. Hsieh, H.J. Su & Tzean, 1987.

Penicillium gladioli L. Mycobank P. glandicola (Oudem.) Seifert & Samson,1985 Mycobank

P. griseofulvum Tzean, SS et al. 1994. Penicillium italicum Tzean, SS et al. 1994.

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P. marinum Frisvad & Samson, Mycobank P. sclerotigenum Mycobiology. Won Ki Kim et al.,2008

P. ulaiense . Hsieh, Su & Tzean,1987. P. vulpinum (Cooke & Massee) Seifert & Samson, 1985 Mycobank

Penicillium roqueforti

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11. References

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