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. 2025 Jun 12:16:e144731.
doi: 10.3897/imafungus.16.144731. eCollection 2025.

Morphological and molecular re-assessment of European and Levantine species of the genus Hortiboletus (Boletaceae)

Alona Yu Biketova  1   2   3   4 , Tatyana Yu Svetasheva  5 , Andy F S Taylor  6 , Giampaolo Simonini  7 , Matteo Gelardi  1   8 , Olga V Morozova  9 , Elias Polemis  10 , José A Muñoz  11 , László Albert  12 , Salvatore Saitta  13 , Solomon P Wasser  4   14 , Eviatar Nevo  4 , Georgios I Zervakis  10 , Alfredo Vizzini  15 , Bálint Dima  16
Affiliations

Morphological and molecular re-assessment of European and Levantine species of the genus Hortiboletus (Boletaceae)

Alona Yu Biketova et al. IMA Fungus. .

Abstract

Hortiboletus (the former Xerocomusrubellus species complex) is one of the most taxonomically critical and difficult genera for species identification in the family Boletaceae. Here, we provide a detailed morphological and molecular re-assessment of European and Levantine species of Hortiboletus. A new species, H.hershenzoniae, is described from Israel. It is sister to H.engelii and associated with the evergreen oak Quercuscalliprinos and potentially also with Q.ithaburensis. Based on the sequence retrieved from INSDC, this species is also found in Lebanon. Accurate morphological descriptions, comprehensive sampling, type studies, biogeography, macro- and microphotographs and a historical overview on the nomenclatural issues surrounding H.rubellus, H.bubalinus, H.engelii, and H.hershenzoniae are given. An epitype collection is designated for H.rubellus. A key is provided for identification of the European and Levantine taxa. In addition, we propose a novel taxonomic combination Hortiboletusflavorubellus, which is conspecific with Boletusrubellusvar.flammeus, based on the DNA barcoding and phylogenetic analysis of type material. Boletusharrisonii is also shown to be conspecific with H.campestris. A multilocus phylogenetic analysis of four markers (ITS, LSU, tef1-α, and rpb2) reveals that Hortiboletus is a sister genus to Xerocomellus. Using the Genealogical Concordance Phylogenetic Species Recognition method, at least 19 phylogenetic species and eight putative phylogenetic species of the genus Hortiboletus can be delimited. Based on multilocus analysis, it contains from 24 to 25 species-level clades worldwide, 17 out of which represent known species, one newly described and potentially six to seven undescribed species. Tandem repeat insertions within the ITS region (both in ITS1 and ITS2) are reported for the first time, not only in the genus Hortiboletus, but in the entire subfamily Boletoideae. Their identification and characterisation were based on Tandem Repeat Finder analysis and visual assessment of the ITS alignment.

Keywords: Agaricomycetes; Boletales taxonomy; biogeography; diversity; phylogeny; xerocomoid fungi.

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Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Figure 1.
Figure 1.
ML phylogenetic tree of Hortiboletus generated from a multilocus (ITS + LSU + tef1-a + rpb2) dataset. BS values ≥ 50% and PP values ≥ 0.8 indicated at nodes. Thickened branches indicate high statistical support (BS ≥ 70% and PP ≥ 0.95). Species names of collections follow current identification, except original names of types of Boletusharrisonii and B.rubellusvar.flammeus. Hortiboletus species present in Europe and Levant are indicated by light red-coloured fields. Collections with newly-generated sequences are indicated in bold. Two-letter country codes (ISO 3166-1 alpha-2) denote the origin of specimens. At the end of the annotations to some collections, abbreviations are indicated in brackets: e – epitype, h – holotype, p – paratype, env – environmental sample.
Figure 2.
Figure 2.
Phylogenetic trees of Hortiboletus from separate ML analyses of each locus: ITS, LSU, tef1-a, and rpb2. BS values ≥ 50% and PP values ≥ 0.8 are indicated at nodes. Thickened branches indicate high statistical support (BS ≥ 70% and PP ≥ 0.95). Pie charts underneath each label are the percent of total characters in the alignment that are parsimony-informative.
Figure 3.
Figure 3.
Scanning electron micrographs of Hortiboletus basidiospores: aH.rubellus (MCVE31743, epitype); bH.engelii (K-M000170238); c, dH.hershenzoniae (K-M001435594, holotype). Photos: (a) A. Yu. Biketova and Y. Pollak; (b) A. Yu. Biketova and B. Dobrić; (c, d) A. Yu. Biketova and M. Kalina.
Figure 4.
Figure 4.
Distribution map of aH.rubellus and bH.bubalinus, based on the current study and sequence data from public repositories and associated publications.
Figure 5.
Figure 5.
Distribution map of aH.engelii and bH.hershenzoniae, based on the current study and sequence data from public repositories and associated publications.
Figure 6.
Figure 6.
Hortiboletusrubellus basidiomes: a epitype collection MCVE31743; b MG781; c DB-2022-06-29-3; d AL 13-125, e LE F-316026; f K-M001435677; g basal mycelium of Vgy-2018-07-21-1; h orange-red dots in the stipe base context of GK6705. Photos: (a) G. Simonini; (b) M. Gelardi; (c) B. Dima; (d) L. Albert; (e) A. I. Ivanov; (f) A. Yu. Biketova; (g) G. Vrba; (h) G. Konstantinidis.
Figure 7.
Figure 7.
Microscopical features of H.rubellus: a basidiospores of the epitype collection MCVE31743 in L4; b basidia in Congo red, left collection K-M001437348, right – MCVE31743; c pileipellis in Congo red, left collection MCVE31743, centre – GS10226, right – GS11020; d pleurocystidia in Congo red (K-M001437348); e cheilocystidia in Congo red (GS11020); f end cells of pileipellis in Congo red (GS10226). Scale bar: 10 μm. Photos: G. Simonini.
Figure 8.
Figure 8.
Size distributions of a spores and b terminal cells of pileipellis in Hortiboletus using the “isoprobability ellipse” method. The distributions of average spore sizes in collections are shown with one standard deviation.
Figure 9.
Figure 9.
Hortiboletusbubalinus basidiomes: a holotype collection L 0053449; b AL12-28; c AL22-75; d MCVE25582; e TUR-A209256; f LE F-315962; g GS10842; h LE F-332081; i LE F-332077. Photos: (a) B. Dima; (b-c) L. Albert; (d) M. Gelardi; (e) S. Saitta; (f) O. V. Morozova; (g) E. Bizio; (h) T. Yu. Svetasheva; (i) E. S. Popov.
Figure 10.
Figure 10.
Microscopical features of H.bubalinus: a basidiospores of collection MCVE18373 in L4; b basidia in Congo red (GS10414); c pileipellis in Congo red; d pleurocystidia in Congo red, left collection GS10072, others – GS10414; e tuft of cheilocystidia in Congo red (GS10414). Scale bar: 10 μm. Photos: G. Simonini.
Figure 11.
Figure 11.
Hortiboletusengelii basidiomes: a JAM0547; b epitype collection MCVE18563; c MCVE18267; d GS10273; e GS10848; f AL21-73; g TUR-A209577; h orange-red dots in the stipe base context of GK6124. Photos: (a) J. A. Muñoz; (b) A. F. S. Taylor; (c) G. Simonini; (d) A. Mua; (e) M. Raumi; (f) L. Albert; (g) S. Saitta; (h) G. Konstantinidis.
Figure 12.
Figure 12.
Microscopical features of H.engelii: a basidiospores in L4; b basidia in Congo red; c pileipellis in Congo red; d pleurocystidia in Congo red; e tuft of cheilocystidia in Congo red. All photos from collection GS10409. Scale bar: 10 μm. Photos: G. Simonini.
Figure 13.
Figure 13.
Hortiboletushershenzoniae basidiomes: a, b holotype collection K-M001435594; c K-M001435695; d K-M001435706; e, f pileus surface of collections: e K-M001435695 and f K-M001435706. Scale bars: 2 mm. Photos: (a, b) R. Kuznetsov; (c–f) A. Yu. Biketova.
Figure 14.
Figure 14.
Details of macroscopical features of H.hershenzoniae: a uneven, denticulate hymenophoral surface of collection K-M001436710; b–d context of the stipe base: b without orange-red dots in a young basidiome (K-M001436710), c with sporadic orange-red dots in young basidiome (K-M001435695), and d collapsed and dispersed orange-red dots throughout the whole stipe base context of a mature basidiome (K-M001435695); e stipe surface with reddish fibrils on light-yellow ground in a brown-pileus basidiome of K-M001436710; f stipe surface with light-yellow ground in a red-pileus basidiome of K-M001435695. Scale bars: 2 mm (a, e, f); 1 mm (b–d). Photos: A. Yu. Biketova.
Figure 15.
Figure 15.
Microscopical features of H.hershenzoniae: a basidiospores of collection K-M001435693 in L4; b basidia in Congo red, left collection K-M001435710, right – K-M001435695; c pileipellis in Congo red, left collection K-M001435693, centre – K-M001435686, right – K-M001435594 (holotype); d pleurocystidia in Congo red, from left to right collections K-M001435710, K-M001435695 – twice, and K-M001435710; e cheilocystidia in Congo red of K-M001435710. Scale bar: 10 μm. Photos: G. Simonini.
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