Fungi Eurotiales Aspergillaceae Torres-GarciaDanielGenéJosepaGarcíaDaniaNew and interesting species of Penicillium (Eurotiomycetes, Aspergillaceae) in freshwater sediments from SpainMycoKeys010220228610314510.3897/mycokeys.86.73861 Penicillium vaccaeorum 109999 Quintanilla, Mycopathol. 80: 74. 1982.Figure 16 =Penicilliumlacussarmientei Ramírez, Mycopathol. 96: 29. 1986. Type.

Spain, Valladolid, San Miguel del Arroyo, from sandy soil under pine tree; J.A. Quintanilla (holotypeCBS H-148.83, cultures ex-type CBS 148.83, DTO 9E2, CECT 2753).

10.3897/mycokeys.86.73861.figure16

Morphological characters of Penicilliumvaccaeorum (FMR 17967). A colonies from left to right (top row) CYA, MEA, YES, and OA; (bottom row) CYA reverse, MEA reverse, DG18, and CREAB–E conidiophores on MEAF conidia. Scale Bars: 25 μm (B), 10 μm (C–F).

https://binary.pensoft.net/fig/640615
Subgeneric classification.

Subgenus Aspergilloides, sectionCitrina, series Roseopurpurea

Description.

Mycelium superficial and immersed composed of septate, smooth-walled, hyaline hyphae of 1.5–2.5 μm wide. Conidiophores monoverticillate, rarely biverticillate and divaricate; stipes smooth-walled, vesiculate, 22.5–103 × 1.5–2.5 μm (vesicle up to 4.5 µm); metulae divergent 2–3, unequal in length, 7–37 × 1.5–3 µm; phialides 2–5 per stipe/metula, ampulliform, 6–8.5 × 2–2.5 μm; conidia smooth- or finely roughened, globose, 2–2.5 × 2–2.5 μm.

Culture characteristics (7 d at 25 °C).

Colonies on CYA, 20–22 mm diam., slightly raised, velvety, radially sulcate, dull red (8C3) at center to light yellow (4A5) and white (1A1) towards periphery, margins slightly undulate, sporulation sparse, conidial masses grayish green (28B3); reverse brownish orange (5C6); with reddish soluble pigment. On MEA, 24–27 mm diam., slightly elevated, velvety, light yellow (4A5) and pale orange (5A2) at periphery, margins low and entire, sporulation sparse, conidial masses grayish green (27C4); reverse golden yellow (5B7) and reddish-yellow (4A6) at periphery; soluble pigment absent. On YES, 30–32 mm diam., slightly raised at center, velvety, radially sulcate, pale yellow (3A3) to white (1A1) and brownish orange (5C3) towards periphery, margins slightly undulate, sporulation abundant, conidial masses grayish green (28B3); reverse brownish yellow (5C8); soluble pigment absent. On OA, 18–20 mm diam., flattened, velvety, dark green (28F4) to light gray (25D1) and white (1A1) towards periphery, margins low and entire, sporulation abundant, conidial masses dull green (25D3); reverse brown (6E4) and yellowish gray (4B2) at periphery; soluble pigment absent. On DG18, 12–13 mm, slightly raised at center, velvety, radially sulcate, white (1A1) and yellowish white (1A2) towards periphery, margins regular, sporulation sparse, conidial masses grayish green (27C3); reverse light yellow (4A5) and white (1A1) at periphery; soluble pigment absent. On CREA, 9–11 mm diam., flattened, floccose, yellowish green (29B7) and white (1A1) towards periphery, margins entire, sporulation sparse, conidial masses grayish green (28B3); reverse dark gray (1F1); soluble pigment and production of acid absent.

Colony diameter on CYA after 7d (mm).

On CYA: 5 °C no growth, 15 °C 13–14, 20 °C 16–17, 30 °C 18–20, 35 °C 6–11, 37 °C no growth, 40 °C no growth.

Specimens examined.

Spain, Catalonia, Fogars de Montclús, La Costa de l’Infern, from stream sediments, Oct 2018, D. Torres (FMR 17531); Fogars de Montclús, La Costa de l’Infern, from stream sediments, Oct 2018, D. Torres (FMR 17534); Aitona, Segre River, from sediments, Dec 2020, D. Torres & J. Gené (FMR 18100); La Granja d’Escarp, Segre River, from sediments, Dec 2020, D. Torres & J. Gené (FMR 18123); Balearic Islands, Mallorca, Serra de Tramontana, from stream sediments, Dec 2018, J. F. Cano (FMR 17616); Basque Country, from stream sediments, Aug 2019, J. Gené (FMR 17967).

Distribution.

Argentina, Canada, Chile, Spain, The Netherlands and Turkey.

Notes.

Penicilliumvaccaeorum and P.lacussarmientei, two species described from sandy soils in Spain and Chile (Quintanilla 1982; Ramírez 1986), respectively, were considered synonyms of P.roseopurpureum by Frisvad et al. (1990), noting that both species were fast growing variants of P.roseopurpureum. Later on, based on that criterion and the lack of morphological differences, Houbraken et al. (2011) considered the two former species synonyms of P.sanguifluum despite some sequence variation where P.vaccaeorum and P.lacussarmientei clustered together in a clade sister to that of P.sanguifluum. Our phylogeny correlates with Houbraken et al. (2011) who found the same topology. Having the opportunity to examine specimens from both monophyletic sister clades (Fig. 8), we observed consistent phenotypic features to distinguish them. For instance, isolates of P.vaccaeorum had longer stipes (up to 103 μm; up to 120 μm in the protologue of the species) (Quintanilla 1982), they were able to grow on CYA at 35 °C (6–11 mm diam. after 7 d), had good sporulation and faster growth on YES agar (30–32 mm diam. 7 d) and more restricted on DG18 (12–13 mm diam. 7 d). In contrast, isolates of the P.sanguifluum clade showed considerably shorter conidiophores (15–50 µm long), they were unable to grow above 30 °C, and the colonies on YES and DG18 showed sparsely or absent sporulation and attained 18–28 mm and 16–22 mm diam., respectively (Houbraken et al. 2011). Hence, genetic and phenotypic differences support the reinstatement of P.vaccaeorum as an accepted species, with P.lacussarmientei considered synonym. This species together with P.sanguifluum and P.roseopurpureum are classified in series Roseopurpurea, which differs from almost all series of the sectionCitrina by species’ monoverticillate conidiophores. The only other series in the section with monoverticillate conidiophores is Gallaica, represented exclusively by P.gallaicum, which differs from the former series mainly by the production of sclerotia (Houbraken et al. 2020).

According to the revised data, P.vaccaeorum occurs worldwide, and is commonly isolated from sandy soils of beaches and forests, and even associated with ants (Table 1).

10.3897/mycokeys.86.73861.figure8

Phylogenetic tree of PenicilliumsectionCitrina based on ML analysis obtained by RAxML inferred from the combined tub2, cmdA, and ITS loci. Branch lengths are proportional to phylogenetic distance. Bootstrap support values/Bayesian posterior probability scores above 70%/0.95 are indicated on the nodes. Bold branches indicate bs/pp values 100/1. The tree is rooted to P.cainii DAOM 239914 and P.jacksonii CCFC 239937. The name in green is the resurrected species P.vaccaeorum included in this study. T= Ex-type strain.

https://binary.pensoft.net/fig/640607

Strain information and GenBank/EMBL accession numbers of the Penicillium species investigated in this study.

SpeciesSectionStrain no.1Substrate and OriginGenBank nucleotide accession no.2:Citation
tub2 cmdA ITS rpb2
P.ausonanum LanataDivaricata FMR 16948T Fluvial sediment, stream of the Guilleries National Park, Barcelona, Catalonia, Spain LR655809 LR655810 LR655808 LR655811 This study
P.guarroi Gracilenta FMR 17747T Fluvial sediment, Brugent River, Tarragona, Catalonia, Spain LR814134 LR814140 LR814139 LR814145 This study
P.heteromorphum Exilicaulis CBS 226.89TSoil, China KJ834455 KP016786 KC411702 JN406605 Visagie et al. 2014a
Exilicaulis FMR 18043 Fluvial sediment, stream of the Cadí–Moixerò Natural Park, Lleida, Catalonia, Spain LR861780 LR861782 LR861783 LR861784 This study
P.irregulare Canescentia FMR 17859T Fluvial sediment, Miraflores River, Community of Madrid, Spain LR814144 LR814151 LR814181 LR814182 This study
P.sanguifluum Citrina CBS 127032TSoil, Calahonda, Costa del Sol, Spain JN606819 JN606555 JN617681 Houbraken et al. 2011
Citrina FMR 17617 Fluvial sediment, Mallorca Island, Spain LR861778 LR861781 LR861779 This study
Citrina FMR 17619 Fluvial sediment, Mallorca Island, Spain OU375375 This study
P.sicoris Paradoxa FMR 18076T Fluvial sediment, Segre River, Lleida, Catalonia, Spain LR884494 LR884496 LR884497 LR884495 This study
P.submersum Robsamsonia FMR 17140T Fluvial sediment, stream of the Montsant Natural Park, Tarragona, Catalonia, Spain LR814187 LR814188 LR814194 LR814195 This study
P.tardochrysogenum Chrysogena CBS 132200TSoil, McMurdo Dry Valley, Antarctica JX996898 JX996239 JX997028 JX996634 Houbraken et al. 2012
Chrysogena FMR 17137 Fluvial sediment, stream of the Montsant Natural Park, Tarragona, Catalonia, Spain HG996463 HG996465 HG996464 This study
P.vaccaeorum Citrina CBS 148.83TSandy soil under pine tree, Valladolid, Spain JN606846 JN606543 MH861558 Houbraken et al. 2011
Citrina FMR 17967 Fluvial sediment, Basque Country, Spain LR814226 LR814227 LR814235 This study
Citrina FMR 17531 Fluvial sediment, stream of Montseny National Park, Barcelona, Spain LR814203 LR814204 LR814213 This study
Citrina FMR 17534 Fluvial sediment, stream of Montseny National Park, Barcelona, Spain OU375168 OU375273 OU375272 This study
Citrina FMR 17616 Fluvial sediment, stream of Serra de Tramontana, Mallorca, Spain LR814212 LR814218 LR814217 This study
Citrina FMR 18100 Fluvial sediment, Segre River, Lleida, Spain LR814234 LR814242 LR814241 This study
Citrina FMR 18123 Fluvial sediment, Segre River, Lleida, Spain LR814265 LR814264 LR814273 This study
Citrina CBS 110.64Forest soil, Erzurum, Turkey JN606829 JN606533 MH858377 Houbraken et al. 2011
Citrina CBS 441.88Sandy soil, Chile JN606846 JN606568 Houbraken et al. 2011
Citrina CBS 643.73Sandy soil, Manitoba, Canada JN606576 Houbraken et al. 2011
Citrina CBS 644.73Sandy soil, Manitoba, Canada LR814213 JN606577 Houbraken et al. 2011
Citrina CBS 685.85 (Type of P.lacussarmientei)Sandy soil, Torres del Paine National Park, Tierra del Fuego, Chile JN606855 JN606533 JN617711 Houbraken et al. 2011
Citrina CBS 300.67Sandy greenhouse soil, The Netherlands JN606561 Houbraken et al. 2011
Citrina CBS 127029Forest soil, Los Alerces National Park, Argentina JN606814 MH864309 Houbraken et al. 2011
Citrina CBS 118024Ants (Camponotus spp.), New Brunswick, Canada JN606833 JN606537 Houbraken et al. 2011

CBS: Culture collection of the Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands; FMR: Facultat de Medicina i Ciències de la Salut, Reus, Spain. T Indicate ex-type strains. 1 Strain no.: strain number. 2tub2: β-tubulin; cmdA: calmodulin; ITS: Internal transcribed spacer regions of the rDNA and 5.8S region; rpb2: the DNA dependent RNA polymerase II largest subunit. Novelties and sequences generated in this study are in bold.

QuintanillaJA (1982) Three new species of Penicillium isolated from soil.Mycopathologia80: 7382. https://doi.org/10.1007/BF00641181RamírezC (1986) A new species of Penicillium from the Chilean Tierra del Fuego.Mycopathologia96: 2932. https://doi.org/10.1007/BF00467682FrisvadJCSamsonRAStolkAC (1990) Disposition of recently described species of Penicillium.Persoonia14: 209232.HoubrakenJFrisvadJCSamsonRA (2011) Taxonomy of PenicilliumsectionCitrina.Studies in Mycology70: 53138. https://doi.org/10.3114/sim.2011.70.02HoubrakenJKocsubéSVisagieCMYilmazNWangXCMeijerMKraakBHubkaVBenschKSamsonRAFrisvadJC (2020) Classification of Aspergillus, Penicillium, Talaromyces and related genera (Eurotiales): an overview of families, genera, subgenera, sections, series and species.Studies in Mycology95: 5169. https://doi.org/10.1016/j.simyco.2020.05.002VisagieCMHoubrakenJFrisvadJCHongSBKlaassenCHWPerroneGSeifertKAVargaJYaguchiTSamsonRA (2014a) Identification and nomenclature of the genus Penicillium.Studies in Mycology78: 343371. https://doi.org/10.1016/j.simyco.2014.09.001HoubrakenJFrisvadJCSeifertKAOveryDPTuthillDMValdezJGSamsonRA (2012) New penicillin-producing Penicillium species and an overview of sectionChrysogena.Persoonia29: 78100. https://doi.org/10.3767/003158512X660571