Mycologia Iranica
Volume:10 Issue: 1, Winter and Spring 2023

Phytopythium is a cosmopolitan genus found in different regions of the world from various substrates. This genus is a newly described taxon that was once a member of the genus Pythium sensu lato. Phytopythium is an intermediate genus between Pythium and Phytophthora with 34 formally described species. Recently, some studies focused on the phylogeny of this genus in Iran. Although not many studies concentrated on isolating species assigned to this genus from Iran, some comprehensive studies showed that Phytopythium is an important genus with vast distribution in this part of the world. Accurate identification and classification of Phytopythium species are quite challenging. Morphological identification of Phytopythium is troublesome due to the lack of identification keys, overlapping of some morphological features, the presence of species complexes, pleomorphism, and the absence of certain structures in some species. Besides, most species cannot be separated using only one or two loci for the phylogenetic analyses. In addition, some studies in Iran do not include molecular investigations to support their morphological identification or make it possible to reidentify the reported species. Having no accurate and current checklist of the country’s species also adds to the problem. This review focuses on the current systematics of Phytopythium species in Iran, discussing the challenges in the morphological and molecular identification of the species in the country. It also proposes some approaches to address the problem of characterizing the species in the genus Phytopythium.

The walnut (Juglans regia L.) tree is considered one of the most important fruit trees in Iran. Decline disease of walnut trees is one of the significant problems that have been noticed in different regions of Kerman province (South-eastern Iran) during the last ten years. Therefore, a study was conducted to identify the fungal species associated with walnut trees showing trunk diseases between May 2015 and September 2016. Collected symptomatic wood samples were cut and surface disinfested with NaClO (Sodium hypochlorite) and then plated onto potato dextrose agar (PDA) plates. In this study, 21 isolates of a fungus were isolated from trees showing canker and dieback, and internal wood necrosis in the Bidkhun region of Kerman province. The obtained fungal isolates were first identified and grouped based on the most important morphological characteristics. Total genomic DNA was extracted and identification of the isolates was confirmed by sequence analysis of ITS (the internal transcribed spacer) using the primers ITS1 and ITS4. Based on the most important morphological characteristics and molecular data, these isolates were identified as Parasporendocladia bactrospora. This study is the first report of this species associated with canker and dieback symptoms on walnut trees worldwide and also the first report of this fungus for the funga of Iran.

Apple (Malus domestica) is considered one of the most important economic products in Iran. Its cultivation is common in various regions and so far, more than 200 different domestic and foreign apple cultivars are identified, which Golden and Red Delicious cultivars are the most important cultivars in terms of cultivated area. Various fungal, bacterial, nematode, and viral agents cause weakness and decline of the apple tree and the quantitative and qualitative reduction of the crop. Apple root and crown rot is one of the important diseases of apple root, due to soil-born fungal pathogens, which lead to the decline of apple trees. The disease has a global spread and can occur at all ages of apple production. Different fungal species have been identified in infected regions so far, and it may be due to climate change. Sampling was performed from June to August 2019 from apple orchards with decline symptoms in Semirom and Padena in Isfahan province, Iran. Fungal species, including Fusarium solani, F. oxysporum, F. equiseti, F. acuminatum, and F. redolens, were morphologically identified. To confirm morphological identification, EF1/EF2 and ITS1/ITS4 primers were used in the PCR reaction. In vitro and greenhouse tests confirmed the pathogenicity of the identified species. As a result, it was proved that F. solani, F. oxysporum, F. equiseti, F. acuminatum, and F. redolens could be the causal agents of apple trees decline in Isfahan province, Iran, that F. solani, and F. oxysporum were more abundant.

Exserohilum species are plant or human pathogens, saprobes or endophytes mostly associated with grasses including cultivated cereals. At present, 11 phylogenetic species have been accepted in this genus worldwide. In this study, we introduce a novel species, Exserohilum persianum from Festuca sp. (Poaceae, Poales) showing necrotic leaf lesions based on morphological characteristics and sequence data obtained from the internal transcribed spacer (ITS‒rDNA) region, parts of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and translation elongation factor-1 alpha (TEF1) genes. A detailed morphological description, illustration and comparison with closely allied species are provided.

In the summer of 2022, sansevieria plants with symptoms of root and crown rots were detected in greenhouses of Isfahan province, Iran. Symptomatic tissues of the root were cultured on CMA-PARPH. Recovered oomycete isolates were purified by single-zoospore technique on WA. The baiting method using citrus leaf disc was also used to isolate Phytophthora spp. isolates from the soil around roots. Four isolates were obtained directly from infected root tissues and two isolates from soil. Fungal isolates were identified based on morphological characteristics and molecular data of β-tubulin (βtub) and translation elongation factor 1-alpha gene (tef1-α) genes. According to the morphological and phylogenetic analysis, all isolates were recognized as Phytophthora occultans. Koch's postulates were completed and confirmed that Phytophthora occultans isolates were responsible for sansevieria root and crown rots. To our knowledge, this is the first report that Phytophthora occultans associated with root rot of sansevieria in Iran and probably in the world.

Endophytic fungi are a group of host-associated fungal communities that benefit their hosts. According to the conditions of their specific living environment, plant endophytic fungi produce many bioactive metabolites with different structural features. The bioactive compounds isolated from endophytic fungi, have significant effects on increasing the compatibility of both endophytic fungi and their host plants, such as the tolerance to biotic and abiotic stresses. In addition, some of these metabolites have indicated medicinal and ecological importance. In the present investigation, five fungal endophytes were isolated from the seeds of some medicinal plants. These endophytic isolates were characterized by sequencing of Internal Transcribed Spacer (ITS) regions as Acremonium sp. isolated from Echium amoenum, Epicoccum nigrum from Rosa canina, Fusarium sp. and Fusarium equiseti from Calendula officinalis and Lecanicillium aphanocladii from Physalis peruviana. To screen the phytochemical derivatives of ethyl acetate of these endophytic fungal isolates, the extracts were subjected to phytochemical analysis by gas chromatography-mass spectrometry (GC-MS). GC-MS spectrum of the compounds found in the extracts of the endophytic fungi was matched with the standard compounds present in the WILEY8 library and the National Institute of Standards and Technology (NIST14) library. The GC-MS analysis of extracts from these endophytic fungi revealed the presence of 49 phytocompounds such as 2,4-Di-tert-butylphenol, Hexadecane, Eicosane, Octadecane, Docosane, Nonadecane, Isopropyl myristate, Hexadecanoic acid, Undecane, Methyl stearate and so on. The results of the present study acknowledge that the endophytic fungi of these medicinal plants are the potential source of biologically active compounds and envisage the possible drug discovery using them. In addition, the compounds 14-Beta-H-Pregna and Cyclohexane,1,1'-(2-methyl-1,3-propanediyl) bis are reported here for the first time as fungal metabolites.

In order to contribute to the knowledge of endophytic fungi of Phragmites australis (Poaceae) in Iran, seventeen isolates were recovered from the leaves, stems and roots of healthy common reed plants collected from five different sites around Lake Urmia in the East and West Azarbaijan provinces, Iran. Five different species viz Achroiostachys betulicola, Ac. humicola, Cephalotrichum tenuissimum, Myrmecridium schulzeri, and Priconia igniaria were identified based on morphological characteristics and phylogeny inferred from nuclear internal ribosomal transcribed spacer sequence (ITS-rDNA). To the best of our knowledge, this is the first report of Ac. betulicola and Ac. humicola for the Funga of Iran. In addition, this study provided new insights into the distribution and host range of the identified species.

In a survey on species diversity and phylogeny of ophiostomatoid fungi associated with bark beetle galleries and decline symptoms on woody hosts in the northwestern region of Iran, ninesynnematous fungal isolates with similar growth patterns were isolated from the bark beetle galleries on declining elm trees in Mazandaran, East, and West Azerbaijan provinces. Based on the integration of molecular phylogeny (ITS-rDNA region and tef1-αgene sequences) with cultural and morphological characteristics, the isolates were identified as Graphium carbonarium. Pathogenicity test using anexcised shoot assay revealed that the isolates are pathogenic on Ulmus carpinifolia. Our study is the first occurrence and pathogenicity of G. carbonariumfrom bark beetlae galleries on declining elm in the world.

In a survey of xylarialean fungi in the Northern part of Iran, some specimens that showed affinities to the rosellinoid Xylariaceae were collected. Morphological evidence, phylogenetic analyses of a combined ITS and ACT1 genes dataset confirmed the presence of Dematophora buxi on Buxus sempervirens in Iran for the first time. In addition, to our knowledge, this is the first report of this species in Asia. Dematophora buxi is illustrated, described, and discussed.

Powdery mildew is a common disease of Spiraea spp. worldwide; however, there are no reports of this destructive disease in Spiraea species from Iran.  In recent years, severe powdery mildew symptoms have been observed in Spiraea spp. throughout the Guilan Province. In this study, we collected infected plant specimens and used morphological and molecular approaches to identify the powdery mildew fungus involved in this disease. We did not observe the teleomorphic state of the fungus in this area, and it appears that the fungus occurred only in the anamorphic state. At least six species of Podosphaera occur worldwide, and there is a considerable overlap between the anamorphic characteristics of some species. Hence, the ITS sequence was used to identify common powdery mildew species occurring in Spiraea in the Guilan Province. The results showed that the ITS sequence is useful for Podosphaera species in Spiraea. Podosphaera minor Howe. was identified as the causal agent of powdery mildew disease in this plant. To our knowledge, this is the first report of Podosphaera minor as a fungus in Iran.

Specimens of phalloid fungi, collected from Kerman province, south of Iran, were characterized morphologically and identified as Itajahya rosea (Delile) E. Fisch (Phallaceae, Phallales, Agaricomycetes, Agaricomycotina, Basidiomycota). The main characteristic features of specimens were as have been described as follow:Young fruit-bodies were egg shaped, ovate to sub-globose, 3-6.5 cm in diameter, white to greyish white in color. The preliminary stages of the stipe and gleba can be seen inside the young fruit-bodies via cross-sectioning. Splitting the eggs (which usually happens during the night time), resulted in appearance of fruit bodies in the form of a stipe and a cap-like spore bearing gleba at the apex. Stipe was cylindrical, spongy, pinkish white to pinkish, hollow, tapering at the base and top, reached up to 10 cm in height and up to 3 cm in width in base, enclosed by volva.Receptaculum up to 5.5 cm diameters, fully covered by gleba. Gleba spongy, greenish-brown to black brown, turning blackish with age, soft, mucilaginous and sticky, with strong odor. The top surface of the gleba covers by “calyptra” that is the remnant of fruit body splitting. A fully developed volva, completely surrounds the stipe base (Fig.1). Basidiospores ellipsoid, hyaline, smooth, 3-4 × 1.5-2 µm in dimension. Phalloid form of fruit-body, color and texture of the gleba, presence of the calyptra and pinkish color of stipe were the main morphological characteristics of the specimens for differentiation. This is the first report of species of the genus Itajahya in Iran.Specimen examined: Iran, Kerman province, Jiroft, Jiroft Agricultural Research Center campus (near a felled tree of Ziziphus spina-christi), 25/3/ 2019, Najafiniya, (IRAN 18218 F). Fungi of the family Phallaceae or stinkhorns are interesting and a particular group of mushrooms in the order Phallales, phylum Basidiomycota. The genus Itajahya, is a rare and lesser-known genus of the family Phallaceae that was characterized by Möller in 1895 for a fungus discovered in Brazil. The main characteristic that distinguishes Itajahya from other taxa of stinkhorn fungi is the presence of calyptra at the apex of the gleba (Ottoni et. al. 2010). roseus was originally described from Egypt in 1813 (Delile 1813). Fischer in 1929 placed the species in the genus Itajahya Möller,1895, based on morphological characteristics and then, Kreisel (1996) included Itajahya as a subgenus of Phallus (Ottoni et al. 2010). Cabral et al. (2012) using the DNA sequence and phylogenetic analyses of Phallus roseus, moved species from Phallus and placed it as a species of the genus Itajahya. Later, Marincowitz et al. (2015) sequenced the DNA of the Itajahya galericulata and showed that it is phylogenetically separate from the Phallus and Dictyophora species. Their study also confirmed that Itajahya rosea and I. galericulata (type of the genus) are phylogenetically related and indeed both of them belonged to the genus Itajahya (Cabral et al. 2012, Marincowitz et al. 2015, Patel et. al. 2018).Four species have currently been reported from rare genus Itajahya in the world: Itajahya galericulata, I. rosea, I. hornseyi and I. argentina (Hansford 1954, Patel et. al. 2018). Itajahya rosea is a rare fungal species in desert and semiarid regions recorded from Argentina, Brazil, Bolivia, Egypt, France, Ghana, India, Morocco, Pakistan, Paraguay, South Africa and Yemen (Borde et al. 2021, Campi Gaona et. al. 2017, Kreisel & Al-Fatimi 2008, Moreno et al. 2013, Ottoni et al. 2010)

The genus Toxicocladosporium (Cladosporiaceae, Cladosporiales, and Dothideomycetes) was first introduced by Crous et al. (2007) based on Toxicocladosporium irritans Crous & U. Braun. To date, 20 species namely T. aquimarinum R. Fotedar, Sand.-Den., Kolecka & Boekhout, T. banksiae Crous, R.G. Shivas & McTaggart, T. cacti J.D.P. Bezerra, Souza-Motta & Crous, T. crousianum M. Bakhshi, T. chlamydosporum Crous & M.J. Wingf., T. eucalyptorum M. Bakhshi, Zare & Jafary, T. ficiniae Crous & A.R. Wood, T. glendoranum Crous & Jurjević, T. hominis Sand.-Den., Gené & Deanna A. Sutton, T. immaculatum J.D.P. Bezerra, Souza-Motta & Crous, T. irritans, T. losalamitosense Crous & Jurjević, T. pini Crous & Y. Zhang, T. posoqueriae Crous & R.G. Shivas, T. protearum Crous & Roets, T. pseudovelox Crous, T. pterocarpi Crous, T. qatarense R. Fotedar, Sand.-Den., Kolecka & Boekhout, T. rubrigenum Crous & M.J. Wingf., and T. strelitziae Crous have been described in this genus based on a combination of morphological and molecular data (Crous et al. 2007, 2021a, 2021b; Bezerra et al. 2017; Fotedar et al. 2019; Bakhshi et al. 2022b). Species of this genus are cosmopolitan and occur in a wide range of host plants, different ecosystems, and extreme habitats (Bezerra et al. 2017; Fotedar et al. 2019). In addition, they display diverse lifestyles, from plant pathogens to epiphytes, endophytes, and saprobes (Bezerra et al. 2017; Bakhshi et al. 2022b).Loquat or Japanese medlar (Eriobotrya japonica Lindl.) belongs to the family Rosaceae, is predominantly adapted to tropical and subtropical climates worldwide, and is a highly nutritious fruit tree of economic relevance (Wang & Paterson 2021). In an investigation aimed at revealing the diversity of foliicolous fungi associated with fruit orchards in northern Iran, one isolate of Toxicocladosporium was isolated from the leaf surface of a loquat tree in Golestan Province. The living culture was deposited in the culture collection of the Iranian Research Institute of Plant Protection (IRAN) in Tehran, Iran. To extract genomic DNA using the protocol described by Möller et al. (1992), the fungal isolate was transferred onto a plate containing malt-extract agar (MEA; Merck, Darmstadt, Germany) and incubated at 25 °C for 10 days. Amplification and sequencing of two nuclear loci, DNA-directed RNA polymerase II second largest subunit (RPB2), and internal transcribed spacer regions and intervening 5.8S nrRNA gene of the nrDNA operon (ITS), were carried out with primers RPB2-5F2 + fRPB2-7cR and V9G + ITS4, respectively, as described by Bakhshi et al. (2021). The new sequences generated in this study were further edited and concatenated using the MEGA v. X (Kumar et al. 2018) software in the sense and antisense directions, submitted to GenBank, and accession numbers were obtained. The Bayesian method was performed using MrBayes v. 3.2.6 (Ronquist et al. 2012) for the two-locus DNA phylogenetic analysis, as elucidated by Bakhshi et al. (2021). Neocladosporium leucadendri (Crous) J.D.P. Bezerra, Sand.-Den., Souza-Motta & Crous (CBS 131317, Cladosporiaceae) was used as the outgroup taxon. According to phylogenetic analyses (Fig. 1), the Iranian strain obtained from Japanese medlar clustered together with the T. irritans reference culture (CBS 185.58). To the best of our knowledge, this is the first report of T. irritans for funga of Iran (Bakhshi et al. 2022a). The fungus exhibited the following morphology:Toxicocladosporium irritans Crous & U. Braun, Stud. Mycol. 58: 39 (2007)Description in vitro: on SNA: Mycelium consisting of subhyaline to brown, smooth to finely verruculose branched, septate, 2–3.5 µm wide hyphae. Conidiophores solitary, brown, smooth to finely verruculose, dimorphic. Macronematous conidiophores arising from hyphae, erect, unbranched or branched, subcylindrical, straight to geniculous-sinuous, or irregularly curved, walls thick, septa dark brown, 20–75 × 3–5.5 µm, 1–6-septate. Micronematous conidiophores aseptate, reduced to conidiogenous cells on hyphae, erect, subcylindrical, 8–25 × 2.5–4 µm. Conidiogenous cells integrated, lateral or terminal, smooth to slightly verruculose, brown, subcylindrical with slight taper towards apex, 6–11 × 2.5–4 µm, proliferating sympodially; loci thickened, darkened and refractive, 1–2 µm wide. Conidia in general catenulate in branched or unbranched chains, medium brown to brown, thick-walled, with dark, thick septa, smooth to finely verruculose. Ramoconidia, subcylindrical to broadly ellipsoid, straight or slightly flexuous, polyblastic, with sympodial proliferation, 0–3-septate, 8–18 × 3–4(–5) µm. Conidia ellipsoid to ovoid, 0-1-septate, 6–10 × 3–5 µm; hila darkened and thickened, 1–1.5 µm wide.Culture characteristics: (in the dark, at 25 °C after 20 days): Colonies on MEA, surface folded, velvety, olivaceous grey to olivaceous, with an even to irregularly undulating margin, with moderate aerial mycelium, reverse dark grey, slow-growing, reaching 12 mm. On SNA, surface flat, grey-olivaceous, with sparse aerial mycelium, with even, smooth margins, reaching 11 mm.Specimens examined: Iran: Golestan province, Fazel Abad, on leaves of Eriobotrya japonica (Rosaceae), 27 April 2019, M. Bakhshi, IRAN 4788C (ITS = OQ625992, RPB2 = OQ632795).