Short Communication
A new species of Neofavolus (Polyporales, Basidiomycota) from Brazil
2023 Volume 64 Issue 2 Pages 69-73
Details
2023 Volume 64 Issue 2 Pages 69-73
Neofavolus teixeirae sp. nov. (Basidiomycota) is described and illustrated based on specimens collected from a reforestation area in southeastern Brazil. This new species is characterized by a lateral stipe up to 1.3 cm long, lacerate and angular pores measuring 0.5-2 (-2.5) per mm, and cylindrical to subcylindrical basidiospores. Phylogenetic analyses of the ITS and LSU regions confirmed its phylogenetic placement and taxonomic identity. A key to Neofavolus species is presented.
Neofavolus species were previously considered members of Polyporus P. Micheli ex Adans. Based on morphological and phylogenetic studies, three species with a reduced stipe, glabrous upper surface covered with flattened scales, cuticle composed of agglutinated generative hyphae covering the context, and poroid hymenophore were segregated from the “Favolus group” (Sotome, Akagi, Lee, Ishikawa, & Hattori, 2013). Subsequently, Seelan et al. (2015, 2016) transferred to this genus a species of Lentinus (L. suavissimus Fr.) with decurrent lamellae, sub-poroid only at the stipe apex. Although Favolus and Neofavolus are morphologically similar, they are phylogenetically unrelated, confirming generic segregation (Sotome et al., 2013; Seelan et al., 2015; Zhou & Cui, 2017). Neofavolus has currently eight valid species. Up to date, Neofavolus subpurpurascens (Murrill) Palacio & Robledo is the only species recorded in the Neotropics (Jamaica - type locality and South America) (Palacio, Silveira, & Robledo, 2019; Alcantara, Gugliotta, & Barbosa, 2019). Neofavolus alveolaris (DC.) Sotome & T. Hatt., type species of genus Neofavolus, have been found in southern Brazil, but not in the tropics (Sotome et al., 2013; Coelho & Silveira, 2014; Palacio et al., 2019).
During field samplings of polypores in a reforestation area with native semi-deciduous forest in the municipality of Mogi Guaçu (São Paulo state, Brazil), we collected specimens morphologically similar to Polyporus. We described and illustrated a new species of Neofavolus based on morphological studies and molecular analyses of the ITS and partial LSU regions. In addition, we presented an identification key for Neofavolus species. Specimens were collected at “Reserva Particular de Patrimônio Natural (RPPN) São Marcelo”, municipality of Mogi Guaçu, São Paulo state, Brazil, in 2015 and 2016. This reserve can be considered a reforestation area after 14 years of planting (Colmanetti & Barbosa, 2013). The RPPN is inserted in a mosaic of seasonal semideciduous forest remnants, eucalyptus plantations, other anthropogenic land covers, and heterogeneous reforested areas and is surrounded by remnants of the Cerrado biome (Neotropical savannah) (Colmanetti, Barbosa, Shirasuna, & Couto, 2016). For the morphological analysis, macroscopic basidiome characteristics, such as pileus size, color, and shape, and pore size and shape, were observed using an S6D stereo microscope (Leica, Wetzlar). The color of basidiomes was described according to Küppers (2002). Cross-sections of the basidiomes were mounted on slides with 5% (w/v) KOH solution for microscopy analysis (Teixeira, 1995). Amyloid or dextrinoid reactions were tested using Melzer’s reagent following Gilbertson and Ryvarden (1987). Observations and measurements of microscopic structures were performed using a DM 1000 optical microscope (Leica), and illustrations were drawn with a camera lucida. Abbreviations and codes used for the measurements were as follows: Xm = length mean × width mean, Q = range of length/width ratios, Qm = length/width mean, and n = x/y (x = number of measurements of a given number (y) of specimens) (Coelho, 2005). The specimens were deposited in the SP herbarium (Thiers, 2019, continuously updated).
DNA extraction was performed from basidiomes using Sigma-Aldrich Gen EluteTM Plant Genomic DNA Miniprep Kit (Sigma-Aldrich Corporation, St. Louis, MO, USA). The ITS region (including ITS1, 5.8S, and ITS2) was amplified with ITS1 and ITS4 primers (White, Bruns, Lee, & Taylor, 1990) and the LSU region with LROR primer (Rehner & Samuels, 1994) and LR7 primer (Vilgalys & Hester, 1990) using the Sigma-Aldrich ReadMixTM Taq PCR P4600 kit (Sigma-Aldrich Corporation, St. Louis, MO, USA). Amplification of the ITS region followed these parameters: one cycle at 95 °C for 2 min, five cycles at 95 °C for 45 s, 60 °C for 50 s, and 72 °C for 1 min and 20 s, where the annealing temperature decreased 1 °C each cycle until it reached 56 °C under the touch-down technique (Korbie & Mattick, 2008), 30 cycles at 95 °C for 45 s, 55 °C for 50 s, 72 °C for 1 min and 20 s, and 72 °C for 10 min. Before gene sequencing, PCR products were checked on a 2% agarose gel and then sequenced in both directions using the same amplification primers. For the LSU region, the following cycle parameters were used: one cycle at 94 °C for 2 min, five cycles at 94 °C for 45 s, 54 °C for 50 s, and 72 °C for 1 min and 20 s, where the annealing temperature decreased 1 °C each cycle until it reached 50 °C under the touch-down technique (Korbie & Mattick, 2008); 30 cycles at 95 °C for 45 s, 50 °C for 50 s, 72 °C for 80 s, and 72 °C for 10 min. Nucleotide sequences were obtained using an Applied Biosystems 3730xl DNA Analyzer at the DNA Synthesis and Sequencing Facility (Macrogen, South Korea). Edited sequences were deposited in GenBank.
Sequences included in this study were selected from GenBank (NCBI). Datronia mollis (Sommerf.) Donk and Trametes conchifer (Schwein.) Pilát were chosen as outgroups for ITS and LSU analyses (Palacio et al., 2019). Nucleotide sequences (Table 1) were aligned with MAFFT v.7 (Katoh & Standley, 2013) and then optimized manually with BioEdit 7.2.0 (Hall, 1999). Subsequently, ITS and LSU regions were concatenated in a single file for joint analysis in SequenceMatrix 1.7.8 (Vaidya, Lohman, & Meier, 2011). The nucleotide substitution model was estimated with JModeltest 2.1.4 (Darriba, Taboada, Doallo, & Posada, 2012). As a result, the substitution models selected were TIM2 + G (partition = 010232) for the ITS region and GTR+I+G (partition = 012345) for the LSU region. Both Bayesian analysis (BA) and Maximum Likelihood analysis (ML) were run on the CIPRES Science Gateway (Miller, Pfeiffer, & Schwartz, 2010). The alignments and tree have been submitted to TreeBase (http://purl.org/phylo/treebase/phylows/study/TB2:S29812).
| Species | Reference | Origin | Voucher | GenBank accession numbers | |
| ITS | nLSU | ||||
| Neofavolus alveolaris | Zhou et al. (2016) | China | Dai11290 | KU189768 | KU189799 |
| N. alveolaris | Zhou e Cui (2017) | China | Cui9900 | KX548974 | KX548996 |
| Neofavolus sp. | Seelan et al. (2015) | USA | SV10 | KP283507 | KP283526 |
| Neofavolus sp. | Seelan et al. (2015) | USA | M672 | KP283506 | KP283524 |
| N. cremeoalbidus | Sotome et al. (2013) | Japan | TUMH50006 | AB735979 | AB735956 |
| N. cremeoalbidus | Sotome et al. (2013) | Japan | TUMH50008 | AB735981 | AB735958 |
| N. cremeoalbidus | Sotome et al. (2013) | Japan | TUMH50009 | AB735980 | AB735957 |
| N. suavissimus | Seelan et al. (2015) | USA | DSH2011 | KP283496 | KP283525 |
| N. suavissimus | Zmitrovich e Kovalenko (2016) | USA | LE202237 | KM411460 | KM411476 |
| N. yunnanensis | Luo et al. (2019) | China | CLZhao 1633 | MK834523 | MK834521 |
| N. yunnanensis | Luo et al. (2019) | China | CLZhao 1639 | MK834524 | MK834522 |
| N. mikawae | Zhou et al. (2016) | China | Cui11152 | KU189773 | KU189804 |
| N. mikawae | Zhou e Cui (2017) | China | Dai12361 | KX548975 | KX548997 |
| N. subpurpurascens | Palacio et al. (2019) | Brazil | CG6241 | MH544274 | MH544276 |
| N. subpurpurascens | Palacio et al. (2019) | Brazil | CG6242 | MH544275 | MH544277 |
| N. subpurpurascens | Palacio et al. (2019) | Argentina | Robledo383 | NS | MH544278 |
| N. subpurpurascens | Palacio et al. (2019) | Argentina | Robledo390 | NS | MH544280 |
| N. teixeirae | This study | Brazil | AL60 | MG675058 | MG675059 |
| N. teixeirae | This study | Brazil | AL106 | MT218014 | MT218015 |
| Favolus niveus | Zhou e Cui (2017) | China | Cui11129 | KX548955 | KX548981 |
| F. spatulatus | Zhou et al. (2016) | China | Dai13615 | KU189775 | KU189806 |
| F. acervatus | Zhou et al. (2016) | China | Cui11053 | KU189774 | KU189805 |
| F. pseudobetulinus | Sotome et al. (2011) | Canada | TRTC51022 | AB587629 | AB587620 |
| F. subtropicus | Zhou e Cui (2017) | China | Cui4292 | KX548970 | KX548992 |
| F. emerici | Zhou e Cui (2017) | China | Yuan4410 | KX548954 | KX548980 |
| F. philippinensis | Zhou e Cui (2017) | China | Cui10941 | KX548976 | KX548998 |
| F. pseudoemerici | Zhou e Cui (2017) | China | Cui11079 | KX548958 | KX548984 |
| F. roseus | Sotome et al. (2008) | Malaysia | PEN33 | AB735975 | AB368099 |
| F. brasiliensis | Sotome et al. (2013) | Costa Rica | TENN10242 | AB735976 | AB368097 |
| F. brasiliensis | Sotome et al. (2013) | Brazil | INPA241452 | AB735977 | AB735953 |
| Polyporus arcularius | Zhou et al. (2016) | China | Cui10998 | KX548973 | KX548995 |
| P. brumalis | Zhou et al. (2016) | China | Cui10750 | KU189765 | KU189796 |
| Lentinus badius | Seelan et al. (2015) | Thailand | DED07668 | KP283480 | KP283518 |
| L. tigrinus | Zmitrovich e Kovalenko (2016) | Russia | LE214778 | KM411459 | KM411475 |
| Datronia mollis | Justo e Hibbett (2011) | USA | RLG6304sp | JN165002 | JN164791 |
| Trametes conchifer | Justo e Hibbett (2011) | USA | FP106793sp | JN164924 | JN164797 |
“NS” = no data in GenBank.
Bayesian analysis (BA) was conducted using MrBayes v.3.2.7a (Ronquist et al., 2012) and performed with Monte Carlo independent Markov chain (MCMC). The runs performed ten million generations, two independent races, and four independent chains. The sampling frequency was every 500 generations. The first 10% of the analysis (Burn-in) was discarded. Posterior probabilities (PP) were determined to test branch support. Values of PP ≥ 0.95 were considered statistically significant. Maximum Likelihood (ML) analysis was performed with RAxML v.8.1.4 (Stamatakis, 2014), with 1,000 fast bootstrap repetitions. Bootstrap values > 50% were used to support the final node topologies. This analysis was performed with 100 ML searches and a GTRGAMMA substitution model with no invariant site proportions. All other parameters were standards estimated by the software.
The combined data matrix consisted of 34 ITS sequences with 603 characters and 36 LSU sequences with 1,061 characters. The combined array had a total alignment of 1,664 characters. The combined matrix was analyzed with Bayesian analysis (BA) and Maximum likelihood (ML) inference methods. Trees of BA and ML were compared and resulted in a similar topology, with no inconsistency in any supported clades. Figure 1 shows the BA topology. Phylogenetic analyses were consistent with previous results (Sotome et al., 2013; Seelan et al., 2015; Zhou & Cui, 2017; Luo, Ma, & Zhao, 2019; Xing, Zhou, & Cui, 2020), indicating that the new taxon is within the clade Neofavolus and is sister to Neofavolus subpurpurascens, another Neotropical species, which demonstrate biogeographic correlation. In addition, clades with Favolus Fr. were recovered with high support. Morphological and phylogenetic evidence showed that the new taxon is a new species, segregated from all previously known Neofavolus species.
Taxonomy
Neofavolus teixeirae Alcantara, R. M. Pires & Gugliotta, sp. nov. Figs. 2, 3.
MycoBank no.: MB 824233.
Diagnosis: it differs from other Neofavolus species by the small basidiome, pileus 1 to 1.5 cm long, angular and lacerate pores measuring 0.5-2 (-2.5) per mm, and the stipe up to 1.3 cm long and 3 mm diam.
Type: BRAZIL. São Paulo: Mogi-Guaçu, Reserva Particular de Patrimônio Natural Parque Florestal São Marcelo, Dec 09, 2015, A.A. Alcantara AL60 (holotype, SP467109); ibid, Mar 03, 2016, A.A. Alcantara AL106 (paratype, SP467119).
Etymology: teixeirae is in honor to Dr. Alcides Ribeiro Teixeira, a Brazilian expert in polypores.
Basidiomes annual, stipitate, fleshy when fresh and corky when dry; solitary to cespitose. Pileus flabelliform to infundibuliform, convex, 1-1.5 cm from the base to margin, 0.8-1.5 cm wide, 0.1-0.7 cm thick; surface glabrous, azonate, radially striate, cream color (N00 A40 M10) in fresh condition, pale brown (N10 A40 M30) in dried condition; margin entire, smooth, involute, concolorous with the pileus surface. Stipe lateral to eccentric, cylindrical, smooth, cream color (N00 A40 M10), up to 1.3 cm long, 2-3 mm diam. Pore surface concolorous with the stipe; pores angular, decurrent, 0.5-2 (-2.5) per mm, dissepiments thin, lacerate; tubes cream (N00 A40 M10), up to 1.5 mm deep. Context homogeneous, white to cream (N00 A30 M00), tough when dry, up to 0.7 cm thick. Hyphal system dimitic, with generative hyphae and skeletal-binding hyphae; hyphae non-amyloid and non-dextrinoid. Generative hyphae with clamps, hyaline, thin to thick-walled, 3.3-3.6 μm wide. Skeletal-binding hyphae thick-walled to solid, 2.1-3 (-3.3) μm wide. Hyphal pegs scattered on the hymenium, conic. Pileipellis present as cutis composed of hyaline to brown, parallel, agglutinated, thick-walled generative hyphae, distinct from the contextual hyphae. Basidia clavate, 4-sterigmate, 12-18 × 4.4-5.8 μm, sterigmata 5-6 μm long. Basidiospores cylindrical to subcylindrical, slightly curved, hyaline, thin-walled, smooth, non-amyloid and non-dextrinoid, 7.8-9.5 (-9.8) × 3.1-3.9 (-4) μm (Xm = 8.7 × 3.4 μm), Q = 2.3-2.9 (Qm = 2.6, n = 60/2).
Ecology and distribution: found on dead angiosperm branches, causing white rot. To date, it has only been recorded from seasonal semideciduous forests in São Paulo state, southeastern Brazil.
Comments: Neofavolus teixeirae has a smaller basidiome than other Neofavolus species, with a flabelliform to infundibuliform pileus measuring 1-1.5 cm long, angular and lacerate pores measuring 0.5-2 (-2.5) per mm and stipe up to 1.3 cm long and 3 mm diam. This species is similar to N. mikawae, with which it shares a leathery basidiome when dry and a glabrous pilear surface covered by a cutis, but N. mikawae differs in the size of the stipe (5 mm long), pileus (1.7-6 × 2-8 cm, and 5 mm thick), angular pores (3-5 per mm) and its distribution in temperate areas of China and Japan (Sotome et al., 2013). N. alveolaris and N. subpurpurascens, also found in Brazil, can be distinguished from N. teixeirae by their large (0.5-) 0.7-7 × 0.5-3 mm pores and purplish to vinaceus pileus surfaces, respectively (Sotome et al., 2013; Coelho & Silveira, 2014; Alcantara et al., 2019).
Identification Key to species of Neofavolus
1. Hymenophore with decurrent lamellae, sub-poroid only around the stipe apex ...... Neofavolus suavissimus
1- Hymenophore poroid ...... 2
2. Pilear surface purplish to vinaceus ...... N. subpurpurascens
2- Pilear surface white, cream to brownish orange ...... 3
3. Pilear surface scaled ...... 4
3- Pilear surface glabrous ...... 6
4. Basidiospores small, (5-) 5.5-7.5 (-8) × 2-3 (-3.5) μm ...... N. yunnanensis
4- Basidiospores larger ...... 5
5. Pilei reniform to semicircular, circular in centrally stipitate basidiocarps; contex up to 6 mm thick; pores radially elongated, (0.5-) 0.7-7 × 0.5-3 mm; basidiospores cylindrical, (6.5-) 7-10 (−10.5) × 2.5-4 μm, L= 8.29 μm, W= 3.01 μm, ...... N. alveolaris
5- Pilei suborbicular; context up to 2.5 mm thick; pores angular, 0.7-3 mm × 0.5-1.5 mm; basidiospores cylindrical to navicular, (7.8−) 8.9−12 (−14.5) × 3.1−4.1 (−4.3) μm, L = 10.34 μm, W= 3.63 μm ...... N. squamatus
6. Basidiospores up to 9.5 μm long ...... 7
6- Basidiospores up to 12 μm long ...... 8
7. Pores 0.5-2 (-2.5) per mm; basidiospores 7.8-9.5 (-9.8) × 3.1-3.9 (-4) μm ...... N. teixeirae
7- Pores 3-5 per mm; basidiospores 6-9.5 × 2.3-3.6 μm ...... N. mikawae
8. Pores 2-4 (-5) per mm; basidiospores (7.7-) 8-12 × 3-4 μm ...... N. cremeoalbidus
8- Pores larger, 1-3 mm long × 0.5-1 mm wide; basidiospores 10.4-12 × 3.8-4.5 μm ...... N. americanus
Authors declare no conflicts of interest. All the experiments undertaken in the study comply with the current laws of the country where they were performed.
We thank to Sylvamo of Brazil (formerly International Paper of Brazil) for the support, especially to Miguel Magela and João Machado. To Dr. Luiz Mauro Barbosa (“Coordenação Especial de Restauração de Áreas Degradadas”/CERAD, Instituto de Pesquisas Ambientais) members for their assistance, especially to Fernando Cirilo de Lima and Marcia Regina Ângelo. We thank Viviana Motato-Vásquez for the assistance in the molecular analysis. CNPq and the “Programa de Pós-Graduação em Biodiversidade Vegetal e Meio Ambiente”/Instituto de Pesquisas Ambientais for the scholarship awarded to the first author.
