Endophytic Fungi in Iranian Ulva sp.: First Report and Diversity Analysis

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Ali Moosawi-Jorf This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6196330/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 31 Oct, 2025 Read the published version in Scientific Reports → Version 1 posted 11 You are reading this latest preprint version Abstract Endophytic fungi are diverse microorganisms that colonize plants symbiotically without causing overt infections. While numerous studies have focused on endophytes in terrestrial plants, there are no prior reports of endophytes associated with algae in Iran. Samples of Ulva sp. were collected during the fall of 2022 from the Bandar Abbas Fishery Coast, Iran, and transported to the laboratory. Following surface sterilization, the samples were cultured on potato dextrose agar (PDA) medium and incubated at 25°C for 3 weeks. The resulting isolates were purified using the hyphal tip method. This study identified 33 fungal isolates from Ulva sp. collected at the Bandar Abbas Fishery Coast, Iran. Morphological and molecular analyses classified these isolates into 8 species across 6 genera: Aspergillus , Penicillium , Chaetomium , Cladosporium , Alternaria , and Syncephalastrum . Aspergillus was the most abundant genus (34% of isolates), while Alternaria and Syncephalastrum were the least frequent (9% each). Phylogenetic analyses of ITS, beta-tubulin, GAPDH, TEF, and LSU gene sequences supported the morphological identification of the isolates. Species identified included Aspergillus caespitosus , Aspergillus terreus , Alternaria tenuissima , Alternaria alternata , Cladosporium cladosporioides , Penicillium digitatum , Chaetomium globosum , and Syncephalastrum racemosum . All species are reported here for the first time as endophytes of Ulva sp. in Iran. Furthermore, this study represents the first documentation of endophytic fungi associated with the marine alga Ulva sp. in Iranian waters. This research enhances understanding of the ecological interactions between fungal endophytes and marine algae in Iranian ecosystems, emphasizing the diversity of symbiotic relationships in marine environments. Biological sciences/Microbiology/Fungi/Fungal biology Biological sciences/Microbiology/Fungi/Fungal ecology Endophyte fungus Ulva sp. Bandar Abbas Iran Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction The southern coasts of Iran, bordering the Persian Gulf and the Sea of Oman, are recognized for their rich biological resources, particularly marine macroalgae (seaweeds). Seaweed communities rank among the most productive and biodiverse marine ecosystems, functioning as primary producers and contributing significantly to global oxygen production. Investigating endophytic fungi associated with these macroalgae is essential to understanding the ecological dynamics and biodiversity of Iran’s coastal regions. Globally, over 6,000 seaweed species have been identified, with approximately 300 species documented in the Persian Gulf and the Sea of Oman 1 . The southern coasts of Iran, bordering the Persian Gulf and the Sea of Oman, exhibit remarkable biodiversity, particularly in marine macroalgae (seaweeds). Seaweed communities rank among the most productive marine ecosystems globally, with primary production rates even those of tropical rainforests 2 . The high productivity of multicellular seaweeds and eukaryotic organisms 3 is critical to marine ecosystems. They act as primary producers in marine food webs and contribute significantly to global atmospheric oxygen levels 4 . Endophytic fungi, a diverse group of microorganisms, colonize plants symbiotically without causing overt harm 5 – 6 . Over recent decades, these fungi have attracted considerable research interest due to their functional traits, including enhancing plant growth and tolerance to biotic and abiotic stresses 7–8−9 . They offer promising strategies for mitigating yield losses caused by abiotic stress 10 and can protect host plants from pathogens 6 . Additionally, endophytes produce bioactive metabolites with applications in medicine, agriculture, and industry 11 – 12 . Endophytic colonization has been documented across diverse plant ecosystems, from tropical rainforests to temperate herbaceous communities. However, studies on endophytic fungi associated with marine macroalgae, such as Ulva sp. (family: Ulvaceae), remain limited. The coastal regions of Bandar Abbas, Iran, host abundant Ulva populations, yet their fungal endophytes remain understudied. This study addresses this gap by investigating endophytic fungi in Ulva sp. from the Shilat-Bandar Abbas coastline. Insights from this research could advance understanding of fungal-algal symbioses and their potential applications in agriculture and ecology. Materials and Methods Collection of Plant Material and Isolation of Endophytes In autumn 2022, samples of Ulva lactuca ( Ulva sp.) were collected from the Fishery Coast in Bandar Abbas, Iran. Samples were rinsed with seawater to remove sand and epiphytes, stored in sterile plastic bags, and transported to the laboratory. In the lab, samples were washed with sterile distilled water and refrigerated at 4°C. A multi-step surface sterilization protocol was applied: Immersion in 70% ethanol for 1 minute. Rinsing with sterile distilled water. Immersion in 70% ethanol for 15 seconds. Triple rinsing with sterile distilled water. To validate sterilization efficacy, aliquots from the final rinse water were cultured on potato dextrose agar (PDA). No fungal growth in these controls confirmed successful surface disinfection. Samples were air-dried, cut into 1 cm² fragments, and placed on three culture media: PDA (Potato Dextrose Agar), PDA-SW (PDA supplemented with 200 mL L⁻¹ sterilized seawater), PDA-SLE (PDA supplemented with 200 mL L⁻¹ sterilized Ulva extract). Petri dishes were incubated at 25°C for 3 weeks. Emerging fungal colonies were isolated via the hyphal tip method and identified morphologically using standard taxonomic keys. Initial genus-level identification was based on macroscopic (colony morphology, pigmentation) and microscopic (spore structure, hyphal characteristics) traits. Isolates were further cultured on Czapek’s Yeast Extract Agar (CYA), Malt Extract Agar (MEA), and Potato Carrot Agar (PCA) for refined species-level identification. Molecular Identification Morphological analysis classified the isolates into six genera: Aspergillus , Penicillium , Chaetomium , Cladosporium , Alternaria , and Syncephalastrum . For molecular identification, genomic DNA was extracted from eight isolates (U01–U08). Fungi were cultured on PDA at 28°C for 7 days, after which mycelia were harvested, frozen in liquid nitrogen, and disrupted using a mortar and pestle. Genomic DNA was isolated using the cetyltrimethylammonium bromide (CTAB) method. PCR Amplification Gene targets for amplification included the ITS rDNA region, β-tubulin (BenA), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), selected based on taxonomic requirements (Table 1 ). PCR reactions (25 µL total volume) were prepared with the following components: Table 1 Panel of the primers used for amplifying the ITS, TUB2, TEF-1 alpha, GAPDH and D1/D2 genetic regions. Primers Sequences References ITS1 CTTGGTCATTTAGAGGAAGTAA 13 ITS4 TCCTCCGCTTATTGATATGC 13 T1 AACATGCGTGAGATTGTAAGT 14 T22 TCTGGATGTTGTTGGGAATCC 14 EF1-728F CATCGAGAAGTTCGAGAAGG 15 EF1-986R TACTTGAAGGAACCCTTACC 15 gpd 1 CAACGGCTTCGGTCGCATTG 16 gpd 2 GCCAAGCAGTTGGTTGTGC 16 D1 AACTTAAGCATATCAATAAGCGGAGGA 17 D2 GGTCCGTGTTTCAAGACGG 17 2.5 µL 10× PCR buffer, 1.5 mM MgCl 2 , 200 µM dNTPs, 0.1 µM forward and reverse primers, 0.04 U/µL Taq DNA polymerase (Cinagene, Iran), 10 ng template DNA. The thermal cycling protocol was as follows: Initial denaturation: 95°C for 15 min, 39 cycles of: Denaturation: 95°C for 1 min, Annealing: 56°C for 30 s, Extension: 72°C for 1 min, Final extension: 72°C for 5 min. Table 1 The panel of the primers used for amplifying the ITS, TUB2, TEF-1 alpha, GAPDH, and D1/D2 genetic regions. Primers Sequences References ITS1 CTTGGTCATTTAGAGGAAGTAA 13 ITS4 TCCTCCGCTTATTGATATGC 13 T1 AACATGCGTGAGATTGTAAGT 14 T22 TCTGGATGTTGTTGGGAATCC 14 EF1-728F CATCGAGAAGTTCGAGAAGG 15 EF1-986R TACTTGAAGGAACCCTTACC 15 gpd 1 CAACGGCTTCGGTCGCATTG 16 gpd 2 GCCAAGCAGTTGGTTGTGC 16 D1 AACTTAAGCATATCAATAAGCGGAGGA 17 D2 GGTCCGTGTTTCAAGACGG 17 Data Availability All the sequence data supporting the findings of this study have been deposited in GenBank, with the accession numbers listed in the Table 2 . Sequencing and Phylogenetic Analysis PCR products were visualized via agarose gel electrophoresis under UV light. Amplicons were sequenced bidirectionally (Codon Genetics, Iran) and edited using BioEdit v7.2.5. Sequences were aligned with reference data from GenBank using MAFFT v7.4 and curated in Mesquite v3.6. Phylogenetic trees were constructed via Bayesian inference (MrBayes v3.2) through the CIPRES Science Gateway, with final visualizations prepared in Adobe Illustrator 2019. Table 2 Taxa of the endophytic fungi and the GenBank accession numbers of the nucleotide sequences used in the phylogenetic analysis. Taxa GenBank Accession Numbers S. racemosum PP176476.1 S. racemosum PQ665121.1 P. digitatum PQ619432.1 P. digitatum PQ766534.1 A. caespitosus PQ619416.1 A. caespitosus PQ766532.1 A. tenuissima PQ614905.1 A. tenuissima PQ766531.1 A. alternata PQ614901.1 A. alternata PQ766530.1 C. cladosporioides PQ614852.1 C. cladosporioides PQ766529.1 C. globosum PQ588476.1 C.globosum PQ766535.1 A. terreus PQ588088.1 A. terreus PQ766533.1 Table 3 Identification code, Taxa, and Phylum of the endophytic fungi associated with Ulva sp. from Iran. Code Taxa Phylum U01 A. terreus Ascomycota U02 A.caespitosus Ascomycota U03 A. alternata Ascomycota U04 A. tenuissima Ascomycota U05 P.digitatum Ascomycota U06 C. cladosporioides Ascomycota U07 C.globosum Ascomycota U08 S. racemosum Mucoromycota Results This study aimed to isolate and identify endophytic fungi associated with the genus Ulva in Iran. Morphological characterization identified the seaweed as Ulva sp., characterized by vivid grass-green, tubular fronds with unbranched thalli 18 – 19 . A total of 33 fungal isolates were obtained from Ulva sp. collected at the Bandar Abbas Fishery Coast, Iran. Based on cultural and morphological characteristics, all isolates were classified into 8 species across 6 genera: Aspergillus, Penicillium , Cladosporium , Alternaria , Chaetomium (Ascomycota), and Syncephalastrum (Mucoromycota) (Table 3 ). Taxonomic Distribution: Aspergillus (11 isolates, 34% of total), Penicillium (7 isolates, 21%), Chaetomium (5 isolates, 15%), Cladosporium (4 isolates, 12%), Alternaria (3 isolates, 9%), Syncephalastrum (3 isolates, 9%). Aspergillus was the most abundant genus, while Alternaria and Syncephalastrum were the least frequent (Fig. 1 ). Molecular Validation: Seven isolates (U01–U08) were selected for molecular analysis. Sequences of the ITS rDNA region, β-tubulin, and GAPDH genes were compared to GenBank databases using BLAST. Morphological identifications were confirmed for all isolates. Species Descriptions: Aspergillus terreus (U01) : Morphology : Colonies on MEA reached 55 mm in 7 days at 25°C, with pale orange-buff surfaces and yellow reverse. Conidial heads were dense, and double-columned; conidiophores were smooth to slightly rough (148–247 µm), with globose vesicles (14.5–20 µm). Conidia were globose (2–2.5 µm). Molecular : ITS and β-tubulin sequences matched A. terreus . Novelty : First report as an endophyte of Ulva sp. in Iran (10 isolates). Aspergillus caespitosus (U02) : Morphology : Colonies grew 50 mm in 7 days; green/olive conidia with light yellow reverse. Conidiophores were smooth (150–250 × 5–6 µm), with hemispherical vesicles (9–15 µm). Conidia were globose, rough-walled (3.5–4.5 µm). Molecular : ITS and β-tubulin sequences matched A. caespitosus . Novelty : First global report as an endophyte of Ulva sp. (1 isolate). Alternaria alternata (U03) : Morphology : Grayish-green colonies (50 mm) on PCA. Conidiophores were vertical (40–70 × 3–4 µm), with ellipsoidal conidia (24.5–35 × 5.5–9 µm). Molecular : ITS and GAPDH sequences matched A. alternata . Novelty : First global report as an endophyte of Ulva sp. (1 isolate). Alternaria tenuissima (U04) : Morphology : Gray-to-dark-gray colonies (50 mm) on PCA. Conidiophores were branched (80–120 × 4–6 µm), with obclavate conidia (22–48 × 8–14 µm). Molecular : ITS and GAPDH sequences matched A. tenuissima . Novelty : First global report as an endophyte of Ulva sp. (2 isolates). Penicillium digitatum (U05) : Morphology : Green colonies (41 mm) on MEA. Conidiophores were thin (62–154 × 5–6.5 µm), with cylindrical phialides and oval conidia (6.3–9.5 × 3–6 µm). Molecular : ITS and β-tubulin sequences matched P. digitatum . Novelty : First global report as an endophyte of Ulva sp. (7 isolates). Cladosporium cladosporioides (U06) : Morphology : Olive colonies (41 mm) on SNA. Conidiophores were cylindrical (27–160.5 × 2.9–4 µm), with ovoid conidia (3.5–5 × 2–3 µm). Molecular : ITS and TEF sequences matched C. cladosporioides . Novelty : First global report as an endophyte of Ulva sp. (4 isolates). Chaetomium globosum (U07) : Morphology : Olive-green colonies (8 cm) on PDA. Perithecia were spherical with club-shaped asci (45–57 × 10–12 µm) and lemon-shaped ascospores. Molecular : ITS and β-tubulin sequences matched C. globosum . Novelty : First report as an endophyte of Ulva sp. in Iran (5 isolates). Syncephalastrum racemosum (U08) : Morphology : Rapid growth on PDA (white to black colonies). Sporangiophores were branched (3–4 µm), with spherical vesicles (11–16 µm) and ovoid sporangiospores (2–6 µm). Molecular : ITS and LSU sequences matched S. racemosum . Novelty : First global report as an endophyte of Ulva sp. (3 isolates). In this study, phylogenetic analyses were performed using gene regions selected for their taxonomic utility in fungal classification. Phylogenetic trees for Aspergillus , Penicillium , and Chaetomium were constructed using sequences from the ITS rDNA region and β-tubulin gene (Fig. 2 ). For Cladosporium , the ITS region and translation elongation factor (TEF) gene were analyzed (Fig. 3 ). Alternaria phylogenies were resolved using the ITS region and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene (Fig. 4 ). Finally, Syncephalastrum was analyzed using the ITS region and large subunit (LSU) rDNA (Fig. 5). For Aspergillus , Penicillium , and Chaetomium , a dataset comprising 22 ingroup taxa and the outgroup Xylaria wallichii (MZ648863) was analyzed. BLAST comparisons revealed: Isolate U01 (ITS and β-tubulin sequences) clustered with Aspergillus terreus strain AG466. Isolate U02 grouped with A. caespitosus strain v313-04. Isolate U05 aligned with Penicillium digitatum isolate PD038. Isolate U07 clustered with Chaetomium globosum strain CGMCC39454. Molecular results corroborated morphological identifications (Fig. 2 ). For Alternaria , isolates U03 ( A. alternata ) and U04 ( A. tenuissima ) were analyzed using ITS and GAPDH sequences. A dataset of 29 ingroup taxa and outgroup Stemphylium solani (OQ642110.1) was analyzed via MrBayes. Both isolates grouped within the Alternata clade (Fig. 3 ). However, species boundaries within Alternaria remained unresolved, suggesting additional genetic markers (e.g., TEF, CAL) to improve phylogenetic resolution. Phylogenetic Validation of Cladosporium cladosporioides (Isolate U06) The morphological analysis identified isolate U06 as Cladosporium cladosporioides . To confirm this, a phylogenetic analysis was performed using ITS-rDNA and translation elongation factor (TEF) gene sequences. The dataset included 21 ingroup taxa and the outgroup Cercospora beticola (JX143561). BLAST comparisons revealed high sequence similarity between U06 and reference strains of C. cladosporioides . Phylogenetic analysis (Fig. 4 ) placed isolate U06 within the C. cladosporioides clade, corroborating morphological observations. Phylogenetic Analysis of Syncephalastrum racemosum (Isolate U08) BLAST analysis revealed that the ITS-rDNA and LSU sequences of isolate U08 closely matched Syncephalastrum racemosum reference sequences in the NCBI GenBank. To resolve species boundaries, a multigene phylogenetic analysis was performed using MrBayes, incorporating 12 ingroup taxa and the outgroup Rhizopus americanus (HM999967). Phylogenetic results placed isolate U08 within the S. racemosum clade, clustering with reference strains CBS 441.59 and CBS 302.65. Morphological traits (e.g., rapid sporulation, spherical vesicles, ovoid sporangiospores) further supported this identification. This marks the first global report of S. racemosum as an endophyte of Ulva sp. Discussion Fungi represent one of Earth’s most diverse organisms, with approximately 156,000 documented species globally (Species Fungorum, 2024). Marine fungi, despite their ecological significance in nutrient cycling and symbiotic interactions, remain understudied. This study offers the first report of endophytic fungi associated with Ulva sp. in Iran, identifying eight species across six genera ( Aspergillus , Penicillium , Cladosporium , Alternaria , Chaetomium , Syncephalastrum ), thereby expanding knowledge of fungal diversity in Iran’s coastal ecosystems and revealing novel host-fungus associations. Aspergillus dominated isolates (34%), aligning with its widespread occurrence in marine algae 20 – 21 , while Aspergillus caespitosus and Syncephalastrum racemosum were documented as endophytes of Ulva sp. for the first time globally. Although previously isolated from soil and terrestrial plants, respectively 22 , 23 , their presence in marine algae underscores their adaptability to diverse environments. Additionally, Alternaria alternata and A. tenuissima were recorded in Iranian waters for the first time, suggesting biogeographic variability in algal-fungal partnerships 24 . The identification of Cladosporium cladosporioides , a common marine endophyte 25 , further highlights its ecological versatility. Robust species delimitation was achieved through combined morphological and multi-gene phylogenetic analyses (ITS, β-tubulin, GAPDH, TEF, LSU). For instance, A. terreus (isolate U01) was characterized by pale orange-buff colonies and globose conidia (2–2.5 µm), confirmed by ITS/β-tubulin sequencing, while S. racemosum (isolate U08) exhibited rapid sporulation and ovoid sporangiospores (2–6 µm), validated by LSU/ITS data. Comparisons with global studies reveal consistency with prior records of Penicillium digitatum and Chaetomium globosum as marine endophytes 26 – 27 , though the absence of A. niger or Curvularia spp. , common in other regions 28 , emphasizes the distinctiveness of Iran’s marine mycobiota. This study addresses a critical gap in regional research, as previous work in Iran focused on mangrove or fish-associated fungi, leaving Ulva endophytes unexplored. Globally, few studies have examined Ulva endophytes (e.g., in Bangladesh 29 ), making this dataset valuable for biogeographic comparisons. The functional roles of these endophytes, such as nutrient exchange or stress tolerance, remain unexplored but warrant investigation. For example, A. terreus is known for bioactive compound production 30 , while S. racemosum synthesizes antifungal agents 31 , which could enhance algal resilience in polluted waters. Expanding sampling to other Iranian coastal regions (e.g., Persian Gulf, Sea of Oman) and macroalgal hosts (e.g., Gracilaria , and Sargassum ) could uncover broader biogeographic patterns and co-evolutionary dynamics. Additionally, endophytes like A. caespitosus may aid algal adaptation to climate change-induced stressors, such as warming oceans, a critical avenue for future research. This study underscores the ecological and biotechnological potential of marine endophytes, advocating for further exploration of their roles in sustaining marine ecosystems under environmental change. Conclusion This study provides the first report of endophytic fungi in Iranian Ulva sp., revealing 8 novel species and emphasizing the need for integrative approaches (morphology + genomics) in mycological research. Future research should focus on their functional roles and biotechnological potential (e.g., novel enzymes, bioactive compounds) and inform marine conservation strategies. Declarations Acknowledgments The authors acknowledge Tarbiat Modares University, Tehran, Iran, for its financial support. Author contributions The research was designed, executed, and finalized through the collective efforts of Maryam Besharati-Fardand S. Ali Moosawi-Jorf, who jointly contributed to conceptualization, experimentation, data analysis, and manuscript preparation. Competing interests The authors declare no competing interests. Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have influenced the work reported in this paper. Human and animal participants This study did not involve any human or animal participants. References Hafezieh, M., Abkenar, A. M., Jadgal, S. & Ajdari, A. Biochemical composition and investigation on the economic feasibility of sodium alginate production of brown seaweed Sargassum illicifolium (Turner) C. Agardh, 1820 from Chabahar Bay (Gulf of Oman, Iran). Iran. J. Fisheries Sci. 20 (1), 1–12. 10.22092/IJFS.2021.351036.0 (2021). Whittaker, R. H. & Margulis, L. 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Cite Share Download PDF Status: Published Journal Publication published 31 Oct, 2025 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 05 Jun, 2025 Reviews received at journal 04 Jun, 2025 Reviewers agreed at journal 30 May, 2025 Reviews received at journal 27 Mar, 2025 Reviewers agreed at journal 27 Mar, 2025 Reviewers agreed at journal 27 Mar, 2025 Reviewers invited by journal 27 Mar, 2025 Editor assigned by journal 27 Mar, 2025 Editor invited by journal 23 Mar, 2025 Submission checks completed at journal 20 Mar, 2025 First submitted to journal 10 Mar, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6196330","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":435012647,"identity":"02ca8eb3-218a-46a6-b893-97a22dfec3ca","order_by":0,"name":"Maryam Besharati-Fard","email":"","orcid":"","institution":"Tarbiat Modares University,","correspondingAuthor":false,"prefix":"","firstName":"Maryam","middleName":"","lastName":"Besharati-Fard","suffix":""},{"id":435012648,"identity":"3b47b608-14b6-44a9-9e7b-e7a58c819cbc","order_by":1,"name":"S. Ali Moosawi-Jorf","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABCUlEQVRIiWNgGAWjYBACgwNgSoKfgYHx4QMGBma4DA8uLZZQLUClzMYGRGmxh2gBKWU2k0DWghOYHW8+/OFnmwUzv/RhtqobFdby5u3tDxh+1DDImDfg0HLmWJpkb5sEs2RfMtvtnDPphnPOnDFg7DnGwCNzAIeWGzlmDLzbJJgNzvAfu53bdphxhkQOAwNvAwOPBA6HGdzIMf74F6yFma0YqMV+hvzzB4x/8WsxkAbaogzSwgzUkjhDgsGAGa8tQL9Iy/6TMJPsYWaWBvoleQZPjsFhmWMSuLUAQ+zjmzN1Mvw8zIyfcyqsbWewH3/48E2NjT0uLdjBAWDkkqRhFIyCUTAKRgEqAAD8plGmHA+pTgAAAABJRU5ErkJggg==","orcid":"","institution":"Tarbiat Modares University,","correspondingAuthor":true,"prefix":"","firstName":"S.","middleName":"Ali","lastName":"Moosawi-Jorf","suffix":""}],"badges":[],"createdAt":"2025-03-10 14:08:29","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6196330/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6196330/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41598-025-22071-6","type":"published","date":"2025-10-31T15:57:12+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":80560409,"identity":"69a52711-9033-40f4-acc4-e64479da9b4c","added_by":"auto","created_at":"2025-04-14 16:25:51","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":126590,"visible":true,"origin":"","legend":"\u003cp\u003eThe percentage of Fungal Endophytes Isolated from \u003cem\u003eUlva\u003c/em\u003e sp.\u003c/p\u003e","description":"","filename":"Picture1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/977f27116b94abf8dafa0506.jpg"},{"id":80560411,"identity":"a57b5604-f780-401f-bee1-58af7332306e","added_by":"auto","created_at":"2025-04-14 16:25:51","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":16235,"visible":true,"origin":"","legend":"\u003cp\u003eThe MrBayesphylogenetic tree was constructed for various species within the Aspergillus, Penicillium, and Chaetomium genera by integrating the ITS-rDNA region and the Beta tubulin gene, utilizing the SIPRES site. The number of bootstrap replicates was 1000, and the minimum bootstrap value was above 50%. \u003cem\u003eXylaria wallichii\u003c/em\u003e (MZ648863) was used as out group. Bolded taxa represent isolates of this study identified on \u003cem\u003eUlva\u003c/em\u003e sp.\u003c/p\u003e","description":"","filename":"Picture2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/4dff3114cb2bf922d303c7ee.jpg"},{"id":80560410,"identity":"686b2487-f5fe-4135-a070-92b3d077c1f4","added_by":"auto","created_at":"2025-04-14 16:25:51","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":29944,"visible":true,"origin":"","legend":"\u003cp\u003eThe MrBayes phylogenetic tree was constructed for various species within the genera of \u003cem\u003eAlternaria\u003c/em\u003e by integrating the ITS-rDNA region and the GAPDH gene, utilizing SIPRES site. The number of bootstrap replicates was 1,000, and the minimum bootstrap value was above 50%. \u003cem\u003eStemphylium. solani \u003c/em\u003e(OQ642110.1) was used as an out group. Bolded taxa represent isolates of this study identified on \u003cem\u003eUlva sp\u003c/em\u003e.\u003c/p\u003e","description":"","filename":"Picture3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/4b55294bcfddc008fe8ae06b.jpg"},{"id":80561069,"identity":"408c9c54-2ddf-4dd3-8cfe-f6441a7a256c","added_by":"auto","created_at":"2025-04-14 16:33:51","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":27352,"visible":true,"origin":"","legend":"\u003cp\u003eThe MrBayesphylogenetic tree was constructed for various species within the genera of \u003cem\u003eCladosporium\u003c/em\u003eby integrating the ITS-rDNA region and the TEF gene, utilizing SIPRES site. The number of bootstrap replicates was 1,000, and the minimum bootstrap value was above 50%. \u003cem\u003eCercospora beticola\u003c/em\u003e(JX14356.1) was used as out group. Bolded taxa represent isolates of this study identified on \u003cem\u003eUlva\u003c/em\u003e sp.\u003c/p\u003e","description":"","filename":"Picture4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/0ca8a8363bd67268f96b6ddb.jpg"},{"id":80561068,"identity":"329b3513-3639-4e76-b269-738d3e991d31","added_by":"auto","created_at":"2025-04-14 16:33:51","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":16782,"visible":true,"origin":"","legend":"\u003cp\u003eThe MrBayes phylogenetic tree was constructed for various species within the genera of Syncephalastrum by integrating the ITS-rDNA region and the D1/D2 gene, utilizing SIPRES site. The number of bootstrap replicates was 1,000, and the minimum bootstrap value was above 50%. \u003cem\u003eRhizopus americanus \u003c/em\u003e(HM999967) was used as out group. Bolded taxa represent isolates of this study identified on \u003cem\u003eUlva sp.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Picture5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/0ed187edac45c756f49e2ac0.jpg"},{"id":95039860,"identity":"fbdc01ed-f43b-4045-9ff1-ffed4e91159c","added_by":"auto","created_at":"2025-11-03 16:04:54","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1233382,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6196330/v1/737890e2-d90a-4b90-852d-1a30f9d5ba5a.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Endophytic Fungi in Iranian Ulva sp.: First Report and Diversity Analysis","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe southern coasts of Iran, bordering the Persian Gulf and the Sea of Oman, are recognized for their rich biological resources, particularly marine macroalgae (seaweeds). Seaweed communities rank among the most productive and biodiverse marine ecosystems, functioning as primary producers and contributing significantly to global oxygen production. Investigating endophytic fungi associated with these macroalgae is essential to understanding the ecological dynamics and biodiversity of Iran\u0026rsquo;s coastal regions. Globally, over 6,000 seaweed species have been identified, with approximately 300 species documented in the Persian Gulf and the Sea of Oman\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. The southern coasts of Iran, bordering the Persian Gulf and the Sea of Oman, exhibit remarkable biodiversity, particularly in marine macroalgae (seaweeds). Seaweed communities rank among the most productive marine ecosystems globally, with primary production rates even those of tropical rainforests\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. The high productivity of multicellular seaweeds and eukaryotic organisms\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e is critical to marine ecosystems. They act as primary producers in marine food webs and contribute significantly to global atmospheric oxygen levels\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. Endophytic fungi, a diverse group of microorganisms, colonize plants symbiotically without causing overt harm\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e. Over recent decades, these fungi have attracted considerable research interest due to their functional traits, including enhancing plant growth and tolerance to biotic and abiotic stresses\u003csup\u003e7\u0026ndash;8\u0026minus;9\u003c/sup\u003e. They offer promising strategies for mitigating yield losses caused by abiotic stress\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e and can protect host plants from pathogens\u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e. Additionally, endophytes produce bioactive metabolites with applications in medicine, agriculture, and industry\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eEndophytic colonization has been documented across diverse plant ecosystems, from tropical rainforests to temperate herbaceous communities. However, studies on endophytic fungi associated with marine macroalgae, such as \u003cem\u003eUlva\u003c/em\u003e sp. (family: Ulvaceae), remain limited. The coastal regions of Bandar Abbas, Iran, host abundant \u003cem\u003eUlva\u003c/em\u003e populations, yet their fungal endophytes remain understudied. This study addresses this gap by investigating endophytic fungi in \u003cem\u003eUlva\u003c/em\u003e sp. from the Shilat-Bandar Abbas coastline. Insights from this research could advance understanding of fungal-algal symbioses and their potential applications in agriculture and ecology.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eCollection of Plant Material and Isolation of Endophytes\u003c/h2\u003e \u003cp\u003eIn autumn 2022, samples of \u003cem\u003eUlva lactuca\u003c/em\u003e (\u003cem\u003eUlva\u003c/em\u003e sp.) were collected from the Fishery Coast in Bandar Abbas, Iran. Samples were rinsed with seawater to remove sand and epiphytes, stored in sterile plastic bags, and transported to the laboratory. In the lab, samples were washed with sterile distilled water and refrigerated at 4\u0026deg;C. A multi-step surface sterilization protocol was applied:\u003c/p\u003e \u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eImmersion in 70% ethanol for 1 minute.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eRinsing with sterile distilled water.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eImmersion in 70% ethanol for 15 seconds.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eTriple rinsing with sterile distilled water.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e \u003cp\u003eTo validate sterilization efficacy, aliquots from the final rinse water were cultured on potato dextrose agar (PDA). No fungal growth in these controls confirmed successful surface disinfection.\u003c/p\u003e \u003cp\u003eSamples were air-dried, cut into 1 cm\u0026sup2; fragments, and placed on three culture media:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003ePDA (Potato Dextrose Agar),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003ePDA-SW (PDA supplemented with 200 mL L⁻\u0026sup1; sterilized seawater),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003ePDA-SLE (PDA supplemented with 200 mL L⁻\u0026sup1; sterilized \u003cem\u003eUlva\u003c/em\u003e extract).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003ePetri dishes were incubated at 25\u0026deg;C for 3 weeks. Emerging fungal colonies were isolated via the hyphal tip method and identified morphologically using standard taxonomic keys. Initial genus-level identification was based on macroscopic (colony morphology, pigmentation) and microscopic (spore structure, hyphal characteristics) traits. Isolates were further cultured on Czapek\u0026rsquo;s Yeast Extract Agar (CYA), Malt Extract Agar (MEA), and Potato Carrot Agar (PCA) for refined species-level identification.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eMolecular Identification\u003c/h3\u003e\n\u003cp\u003eMorphological analysis classified the isolates into six genera: \u003cem\u003eAspergillus\u003c/em\u003e, \u003cem\u003ePenicillium\u003c/em\u003e, \u003cem\u003eChaetomium\u003c/em\u003e, \u003cem\u003eCladosporium\u003c/em\u003e, \u003cem\u003eAlternaria\u003c/em\u003e, and \u003cem\u003eSyncephalastrum\u003c/em\u003e. For molecular identification, genomic DNA was extracted from eight isolates (U01\u0026ndash;U08). Fungi were cultured on PDA at 28\u0026deg;C for 7 days, after which mycelia were harvested, frozen in liquid nitrogen, and disrupted using a mortar and pestle. Genomic DNA was isolated using the cetyltrimethylammonium bromide (CTAB) method.\u003c/p\u003e\n\u003ch3\u003ePCR Amplification\u003c/h3\u003e\n\u003cp\u003eGene targets for amplification included the ITS rDNA region, β-tubulin (BenA), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), selected based on taxonomic requirements (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e1\u003c/span\u003e). PCR reactions (25 \u0026micro;L total volume) were prepared with the following components:\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePanel of the primers used for amplifying the ITS, TUB2, TEF-1 alpha, GAPDH and D1/D2 genetic regions.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePrimers\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSequences\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eReferences\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eITS1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCTTGGTCATTTAGAGGAAGTAA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eITS4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTCCTCCGCTTATTGATATGC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR13\" 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\u003cp\u003eCAACGGCTTCGGTCGCATTG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egpd 2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGCCAAGCAGTTGGTTGTGC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAACTTAAGCATATCAATAAGCGGAGGA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGGTCCGTGTTTCAAGACGG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e2.5 \u0026micro;L 10\u0026times; PCR buffer,\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e1.5 mM MgCl\u003csub\u003e2\u003c/sub\u003e,\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e200 \u0026micro;M dNTPs,\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e0.1 \u0026micro;M forward and reverse primers,\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e0.04 U/\u0026micro;L Taq DNA polymerase (Cinagene, Iran),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e10 ng template DNA.\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe thermal cycling protocol was as follows:\u003c/p\u003e \u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eInitial denaturation: 95\u0026deg;C for 15 min,\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003e39 cycles of:\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eDenaturation: 95\u0026deg;C for 1 min,\u003c/p\u003e \u003cp\u003eAnnealing: 56\u0026deg;C for 30 s,\u003c/p\u003e \u003cp\u003eExtension: 72\u0026deg;C for 1 min,\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003col start=\"3\"\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eFinal extension: 72\u0026deg;C for 5 min.\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe panel of the primers used for amplifying the ITS, TUB2, TEF-1 alpha, GAPDH, and D1/D2 genetic regions.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePrimers\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSequences\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eReferences\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eITS1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCTTGGTCATTTAGAGGAAGTAA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eITS4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTCCTCCGCTTATTGATATGC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAACATGCGTGAGATTGTAAGT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTCTGGATGTTGTTGGGAATCC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eEF1-728F\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCATCGAGAAGTTCGAGAAGG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eEF1-986R\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTACTTGAAGGAACCCTTACC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egpd 1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCAACGGCTTCGGTCGCATTG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egpd 2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGCCAAGCAGTTGGTTGTGC\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAACTTAAGCATATCAATAAGCGGAGGA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGGTCCGTGTTTCAAGACGG\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e\n\u003ch3\u003eData Availability\u003c/h3\u003e\n\u003cp\u003eAll the sequence data supporting the findings of this study have been deposited in GenBank, with the accession numbers listed in the Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e\n\u003ch3\u003eSequencing and Phylogenetic Analysis\u003c/h3\u003e\n\u003cp\u003ePCR products were visualized via agarose gel electrophoresis under UV light. Amplicons were sequenced bidirectionally (Codon Genetics, Iran) and edited using BioEdit v7.2.5. Sequences were aligned with reference data from GenBank using MAFFT v7.4 and curated in Mesquite v3.6. Phylogenetic trees were constructed via Bayesian inference (MrBayes v3.2) through the CIPRES Science Gateway, with final visualizations prepared in Adobe Illustrator 2019.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eTaxa of the endophytic fungi and the GenBank accession numbers of the nucleotide sequences used in the phylogenetic analysis.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"2\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTaxa\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGenBank Accession Numbers\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eS. racemosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePP176476.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eS. racemosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ665121.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP. digitatum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ619432.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eP. digitatum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766534.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. caespitosus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ619416.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. caespitosus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766532.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. tenuissima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ614905.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. tenuissima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766531.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. alternata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ614901.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. alternata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766530.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eC. cladosporioides\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ614852.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eC. cladosporioides\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766529.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eC. globosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ588476.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eC.globosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766535.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. terreus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ588088.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eA. terreus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePQ766533.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eIdentification code, Taxa, and Phylum of the endophytic fungi associated with \u003cem\u003eUlva\u003c/em\u003e sp. from Iran.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCode\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTaxa\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePhylum\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU01\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eA. terreus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU02\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eA.caespitosus\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU03\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eA. alternata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eA. tenuissima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP.digitatum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eC. cladosporioides\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU07\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eC.globosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscomycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eU08\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eS. racemosum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMucoromycota\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eThis study aimed to isolate and identify endophytic fungi associated with the genus \u003cem\u003eUlva\u003c/em\u003e in Iran. Morphological characterization identified the seaweed as \u003cem\u003eUlva\u003c/em\u003e sp., characterized by vivid grass-green, tubular fronds with unbranched thalli\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e. A total of 33 fungal isolates were obtained from \u003cem\u003eUlva\u003c/em\u003e sp. collected at the Bandar Abbas Fishery Coast, Iran. Based on cultural and morphological characteristics, all isolates were classified into 8 species across 6 genera: \u003cem\u003eAspergillus, Penicillium\u003c/em\u003e, \u003cem\u003eCladosporium\u003c/em\u003e, \u003cem\u003eAlternaria\u003c/em\u003e, \u003cem\u003eChaetomium\u003c/em\u003e (Ascomycota), and \u003cem\u003eSyncephalastrum\u003c/em\u003e (Mucoromycota) (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\n\u003ch3\u003eTaxonomic Distribution:\u003c/h3\u003e\n\u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003eAspergillus\u003c/em\u003e (11 isolates, 34% of total),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003ePenicillium\u003c/em\u003e (7 isolates, 21%),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003eChaetomium\u003c/em\u003e (5 isolates, 15%),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003eCladosporium\u003c/em\u003e (4 isolates, 12%),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003eAlternaria\u003c/em\u003e (3 isolates, 9%),\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cem\u003eSyncephalastrum\u003c/em\u003e (3 isolates, 9%).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003eAspergillus\u003c/em\u003e was the most abundant genus, while \u003cem\u003eAlternaria\u003c/em\u003e and \u003cem\u003eSyncephalastrum\u003c/em\u003e were the least frequent (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eMolecular Validation:\u003c/h3\u003e\n\u003cp\u003eSeven isolates (U01\u0026ndash;U08) were selected for molecular analysis. Sequences of the ITS rDNA region, β-tubulin, and GAPDH genes were compared to GenBank databases using BLAST. Morphological identifications were confirmed for all isolates.\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eSpecies Descriptions:\u003c/h2\u003e \u003cp\u003e \u003cb\u003eAspergillus terreus\u003c/b\u003e \u003cb\u003e(U01)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Colonies on MEA reached 55 mm in 7 days at 25\u0026deg;C, with pale orange-buff surfaces and yellow reverse. Conidial heads were dense, and double-columned; conidiophores were smooth to slightly rough (148\u0026ndash;247 \u0026micro;m), with globose vesicles (14.5\u0026ndash;20 \u0026micro;m). Conidia were globose (2\u0026ndash;2.5 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and β-tubulin sequences matched \u003cem\u003eA. terreus\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. in Iran (10 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eAspergillus caespitosus\u003c/b\u003e \u003cb\u003e(U02)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Colonies grew 50 mm in 7 days; green/olive conidia with light yellow reverse. Conidiophores were smooth (150\u0026ndash;250 \u0026times; 5\u0026ndash;6 \u0026micro;m), with hemispherical vesicles (9\u0026ndash;15 \u0026micro;m). Conidia were globose, rough-walled (3.5\u0026ndash;4.5 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and β-tubulin sequences matched \u003cem\u003eA. caespitosus\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (1 isolate).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eAlternaria alternata\u003c/b\u003e \u003cb\u003e(U03)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Grayish-green colonies (50 mm) on PCA. Conidiophores were vertical (40\u0026ndash;70 \u0026times; 3\u0026ndash;4 \u0026micro;m), with ellipsoidal conidia (24.5\u0026ndash;35 \u0026times; 5.5\u0026ndash;9 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and GAPDH sequences matched \u003cem\u003eA. alternata\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (1 isolate).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eAlternaria tenuissima\u003c/b\u003e \u003cb\u003e(U04)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Gray-to-dark-gray colonies (50 mm) on PCA. Conidiophores were branched (80\u0026ndash;120 \u0026times; 4\u0026ndash;6 \u0026micro;m), with obclavate conidia (22\u0026ndash;48 \u0026times; 8\u0026ndash;14 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and GAPDH sequences matched \u003cem\u003eA. tenuissima\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (2 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003ePenicillium digitatum\u003c/b\u003e \u003cb\u003e(U05)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Green colonies (41 mm) on MEA. Conidiophores were thin (62\u0026ndash;154 \u0026times; 5\u0026ndash;6.5 \u0026micro;m), with cylindrical phialides and oval conidia (6.3\u0026ndash;9.5 \u0026times; 3\u0026ndash;6 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and β-tubulin sequences matched \u003cem\u003eP. digitatum\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (7 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eCladosporium cladosporioides\u003c/b\u003e \u003cb\u003e(U06)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Olive colonies (41 mm) on SNA. Conidiophores were cylindrical (27\u0026ndash;160.5 \u0026times; 2.9\u0026ndash;4 \u0026micro;m), with ovoid conidia (3.5\u0026ndash;5 \u0026times; 2\u0026ndash;3 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and TEF sequences matched \u003cem\u003eC. cladosporioides\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (4 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eChaetomium globosum\u003c/b\u003e \u003cb\u003e(U07)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Olive-green colonies (8 cm) on PDA. Perithecia were spherical with club-shaped asci (45\u0026ndash;57 \u0026times; 10\u0026ndash;12 \u0026micro;m) and lemon-shaped ascospores.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and β-tubulin sequences matched \u003cem\u003eC. globosum\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. in Iran (5 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eSyncephalastrum racemosum\u003c/b\u003e \u003cb\u003e(U08)\u003c/b\u003e:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMorphology\u003c/b\u003e: Rapid growth on PDA (white to black colonies). Sporangiophores were branched (3\u0026ndash;4 \u0026micro;m), with spherical vesicles (11\u0026ndash;16 \u0026micro;m) and ovoid sporangiospores (2\u0026ndash;6 \u0026micro;m).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMolecular\u003c/b\u003e: ITS and LSU sequences matched \u003cem\u003eS. racemosum\u003c/em\u003e.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eNovelty\u003c/b\u003e: First global report as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp. (3 isolates).\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eIn this study, phylogenetic analyses were performed using gene regions selected for their taxonomic utility in fungal classification. Phylogenetic trees for \u003cem\u003eAspergillus\u003c/em\u003e, \u003cem\u003ePenicillium\u003c/em\u003e, and \u003cem\u003eChaetomium\u003c/em\u003e were constructed using sequences from the ITS rDNA region and β-tubulin gene (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). For \u003cem\u003eCladosporium\u003c/em\u003e, the ITS region and translation elongation factor (TEF) gene were analyzed (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). \u003cem\u003eAlternaria\u003c/em\u003e phylogenies were resolved using the ITS region and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). Finally, \u003cem\u003eSyncephalastrum\u003c/em\u003e was analyzed using the ITS region and large subunit (LSU) rDNA (Fig.\u0026nbsp;5).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eFor \u003cem\u003eAspergillus\u003c/em\u003e, \u003cem\u003ePenicillium\u003c/em\u003e, and \u003cem\u003eChaetomium\u003c/em\u003e, a dataset comprising 22 ingroup taxa and the outgroup \u003cem\u003eXylaria wallichii\u003c/em\u003e (MZ648863) was analyzed. BLAST comparisons revealed:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003eIsolate U01 (ITS and β-tubulin sequences) clustered with \u003cem\u003eAspergillus terreus\u003c/em\u003e strain AG466.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eIsolate U02 grouped with \u003cem\u003eA. caespitosus\u003c/em\u003e strain v313-04.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eIsolate U05 aligned with \u003cem\u003ePenicillium digitatum\u003c/em\u003e isolate PD038.\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eIsolate U07 clustered with \u003cem\u003eChaetomium globosum\u003c/em\u003e strain CGMCC39454.\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eMolecular results corroborated morphological identifications (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eFor \u003cem\u003eAlternaria\u003c/em\u003e, isolates U03 (\u003cem\u003eA. alternata\u003c/em\u003e) and U04 (\u003cem\u003eA. tenuissima\u003c/em\u003e) were analyzed using ITS and GAPDH sequences. A dataset of 29 ingroup taxa and outgroup \u003cem\u003eStemphylium solani\u003c/em\u003e (OQ642110.1) was analyzed via MrBayes. Both isolates grouped within the \u003cem\u003eAlternata\u003c/em\u003e clade (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). However, species boundaries within \u003cem\u003eAlternaria\u003c/em\u003e remained unresolved, suggesting additional genetic markers (e.g., TEF, CAL) to improve phylogenetic resolution.\u003c/p\u003e \u003cp\u003e \u003cb\u003ePhylogenetic Validation of\u003c/b\u003e \u003cb\u003eCladosporium cladosporioides\u003c/b\u003e \u003cb\u003e(Isolate U06)\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe morphological analysis identified isolate U06 as \u003cem\u003eCladosporium cladosporioides\u003c/em\u003e. To confirm this, a phylogenetic analysis was performed using ITS-rDNA and translation elongation factor (TEF) gene sequences. The dataset included 21 ingroup taxa and the outgroup \u003cem\u003eCercospora beticola\u003c/em\u003e (JX143561). BLAST comparisons revealed high sequence similarity between U06 and reference strains of \u003cem\u003eC. cladosporioides\u003c/em\u003e. Phylogenetic analysis (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e) placed isolate U06 within the \u003cem\u003eC. cladosporioides\u003c/em\u003e clade, corroborating morphological observations.\u003c/p\u003e \u003cp\u003e \u003cb\u003ePhylogenetic Analysis of\u003c/b\u003e \u003cb\u003eSyncephalastrum racemosum\u003c/b\u003e \u003cb\u003e(Isolate U08)\u003c/b\u003e\u003c/p\u003e \u003cp\u003eBLAST analysis revealed that the ITS-rDNA and LSU sequences of isolate U08 closely matched \u003cem\u003eSyncephalastrum racemosum\u003c/em\u003e reference sequences in the NCBI GenBank. To resolve species boundaries, a multigene phylogenetic analysis was performed using MrBayes, incorporating 12 ingroup taxa and the outgroup \u003cem\u003eRhizopus americanus\u003c/em\u003e (HM999967). Phylogenetic results placed isolate U08 within the \u003cem\u003eS. racemosum\u003c/em\u003e clade, clustering with reference strains CBS 441.59 and CBS 302.65. Morphological traits (e.g., rapid sporulation, spherical vesicles, ovoid sporangiospores) further supported this identification. This marks the first global report of \u003cem\u003eS. racemosum\u003c/em\u003e as an endophyte of \u003cem\u003eUlva\u003c/em\u003e sp.\u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eFungi represent one of Earth\u0026rsquo;s most diverse organisms, with approximately 156,000 documented species globally (Species Fungorum, 2024). Marine fungi, despite their ecological significance in nutrient cycling and symbiotic interactions, remain understudied. This study offers the first report of endophytic fungi associated with \u003cem\u003eUlva sp.\u003c/em\u003e in Iran, identifying eight species across six genera (\u003cem\u003eAspergillus\u003c/em\u003e, \u003cem\u003ePenicillium\u003c/em\u003e, \u003cem\u003eCladosporium\u003c/em\u003e, \u003cem\u003eAlternaria\u003c/em\u003e, \u003cem\u003eChaetomium\u003c/em\u003e, \u003cem\u003eSyncephalastrum\u003c/em\u003e), thereby expanding knowledge of fungal diversity in Iran\u0026rsquo;s coastal ecosystems and revealing novel host-fungus associations. \u003cem\u003eAspergillus\u003c/em\u003e dominated isolates (34%), aligning with its widespread occurrence in marine algae \u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e, while \u003cem\u003eAspergillus caespitosus\u003c/em\u003e and \u003cem\u003eSyncephalastrum racemosum\u003c/em\u003e were documented as endophytes of \u003cem\u003eUlva sp.\u003c/em\u003e for the first time globally. Although previously isolated from soil and terrestrial plants, respectively \u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e,\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e, their presence in marine algae underscores their adaptability to diverse environments. Additionally, \u003cem\u003eAlternaria alternata\u003c/em\u003e and \u003cem\u003eA. tenuissima\u003c/em\u003e were recorded in Iranian waters for the first time, suggesting biogeographic variability in algal-fungal partnerships \u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e. The identification of \u003cem\u003eCladosporium cladosporioides\u003c/em\u003e, a common marine endophyte \u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e, further highlights its ecological versatility.\u003c/p\u003e \u003cp\u003eRobust species delimitation was achieved through combined morphological and multi-gene phylogenetic analyses (ITS, β-tubulin, GAPDH, TEF, LSU). For instance, \u003cem\u003eA. terreus\u003c/em\u003e (isolate U01) was characterized by pale orange-buff colonies and globose conidia (2\u0026ndash;2.5 \u0026micro;m), confirmed by ITS/β-tubulin sequencing, while \u003cem\u003eS. racemosum\u003c/em\u003e (isolate U08) exhibited rapid sporulation and ovoid sporangiospores (2\u0026ndash;6 \u0026micro;m), validated by LSU/ITS data. Comparisons with global studies reveal consistency with prior records of \u003cem\u003ePenicillium digitatum\u003c/em\u003e and \u003cem\u003eChaetomium globosum\u003c/em\u003e as marine endophytes \u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e, though the absence of \u003cem\u003eA. niger\u003c/em\u003e or \u003cem\u003eCurvularia spp.\u003c/em\u003e, common in other regions \u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e, emphasizes the distinctiveness of Iran\u0026rsquo;s marine mycobiota. This study addresses a critical gap in regional research, as previous work in Iran focused on mangrove or fish-associated fungi, leaving \u003cem\u003eUlva\u003c/em\u003e endophytes unexplored. Globally, few studies have examined \u003cem\u003eUlva\u003c/em\u003e endophytes (e.g., in Bangladesh \u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e), making this dataset valuable for biogeographic comparisons.\u003c/p\u003e \u003cp\u003eThe functional roles of these endophytes, such as nutrient exchange or stress tolerance, remain unexplored but warrant investigation. For example, \u003cem\u003eA. terreus\u003c/em\u003e is known for bioactive compound production \u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e, while \u003cem\u003eS. racemosum\u003c/em\u003e synthesizes antifungal agents \u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e, which could enhance algal resilience in polluted waters. Expanding sampling to other Iranian coastal regions (e.g., Persian Gulf, Sea of Oman) and macroalgal hosts (e.g., \u003cem\u003eGracilaria\u003c/em\u003e, and \u003cem\u003eSargassum\u003c/em\u003e) could uncover broader biogeographic patterns and co-evolutionary dynamics. Additionally, endophytes like \u003cem\u003eA. caespitosus\u003c/em\u003e may aid algal adaptation to climate change-induced stressors, such as warming oceans, a critical avenue for future research. This study underscores the ecological and biotechnological potential of marine endophytes, advocating for further exploration of their roles in sustaining marine ecosystems under environmental change.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study provides the first report of endophytic fungi in Iranian Ulva sp., revealing 8 novel species and emphasizing the need for integrative approaches (morphology\u0026thinsp;+\u0026thinsp;genomics) in mycological research. Future research should focus on their functional roles and biotechnological potential (e.g., novel enzymes, bioactive compounds) and inform marine conservation strategies.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors acknowledge Tarbiat Modares University, Tehran, Iran, for its financial support.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe research was designed, executed, and finalized through the collective efforts of Maryam Besharati-Fardand S. Ali Moosawi-Jorf, who jointly contributed to conceptualization, experimentation, data analysis, and manuscript preparation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of competing interest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no known competing financial interests or personal relationships that could have influenced the work reported in this paper.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHuman and animal participants\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study did not involve any human or animal participants.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eHafezieh, M., Abkenar, A. M., Jadgal, S. \u0026amp; Ajdari, A. 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Sci.\u003c/em\u003e \u003cb\u003e3\u003c/b\u003e (5), 780\u0026ndash;783 (2013).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Endophyte fungus, Ulva sp., Bandar Abbas, Iran","lastPublishedDoi":"10.21203/rs.3.rs-6196330/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6196330/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eEndophytic fungi are diverse microorganisms that colonize plants symbiotically without causing overt infections. While numerous studies have focused on endophytes in terrestrial plants, there are no prior reports of endophytes associated with algae in Iran. Samples of \u003cem\u003eUlva\u003c/em\u003e sp. were collected during the fall of 2022 from the Bandar Abbas Fishery Coast, Iran, and transported to the laboratory. Following surface sterilization, the samples were cultured on potato dextrose agar (PDA) medium and incubated at 25\u0026deg;C for 3 weeks. The resulting isolates were purified using the hyphal tip method. This study identified 33 fungal isolates from \u003cem\u003eUlva\u003c/em\u003e sp. collected at the Bandar Abbas Fishery Coast, Iran. Morphological and molecular analyses classified these isolates into 8 species across 6 genera: \u003cem\u003eAspergillus\u003c/em\u003e, \u003cem\u003ePenicillium\u003c/em\u003e, \u003cem\u003eChaetomium\u003c/em\u003e, \u003cem\u003eCladosporium\u003c/em\u003e, \u003cem\u003eAlternaria\u003c/em\u003e, and \u003cem\u003eSyncephalastrum\u003c/em\u003e. \u003cem\u003eAspergillus\u003c/em\u003e was the most abundant genus (34% of isolates), while \u003cem\u003eAlternaria\u003c/em\u003e and \u003cem\u003eSyncephalastrum\u003c/em\u003e were the least frequent (9% each). Phylogenetic analyses of ITS, beta-tubulin, GAPDH, TEF, and LSU gene sequences supported the morphological identification of the isolates. Species identified included \u003cem\u003eAspergillus caespitosus\u003c/em\u003e, \u003cem\u003eAspergillus terreus\u003c/em\u003e, \u003cem\u003eAlternaria tenuissima\u003c/em\u003e, \u003cem\u003eAlternaria alternata\u003c/em\u003e, \u003cem\u003eCladosporium cladosporioides\u003c/em\u003e, \u003cem\u003ePenicillium digitatum\u003c/em\u003e, \u003cem\u003eChaetomium globosum\u003c/em\u003e, and \u003cem\u003eSyncephalastrum racemosum\u003c/em\u003e. All species are reported here for the first time as endophytes of \u003cem\u003eUlva\u003c/em\u003e sp. in Iran. Furthermore, this study represents the first documentation of endophytic fungi associated with the marine alga \u003cem\u003eUlva\u003c/em\u003e sp. in Iranian waters. This research enhances understanding of the ecological interactions between fungal endophytes and marine algae in Iranian ecosystems, emphasizing the diversity of symbiotic relationships in marine environments.\u003c/p\u003e","manuscriptTitle":"Endophytic Fungi in Iranian Ulva sp.: First Report and Diversity Analysis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-14 16:25:47","doi":"10.21203/rs.3.rs-6196330/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-06-05T05:43:07+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-06-04T13:01:01+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"145074855384140530245473472002707485622","date":"2025-05-30T10:35:09+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-03-27T19:47:11+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"292828871449603950556860353373143096525","date":"2025-03-27T16:58:49+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"199854196704872834287154480642721566631","date":"2025-03-27T09:17:33+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-03-27T07:58:01+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-03-27T07:57:03+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-03-24T03:47:29+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-03-20T11:30:56+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-03-10T14:02:49+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"79a8bcd4-48e1-4dfc-a537-33f24b57b0d5","owner":[],"postedDate":"April 14th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":46323089,"name":"Biological sciences/Microbiology/Fungi/Fungal biology"},{"id":46323090,"name":"Biological sciences/Microbiology/Fungi/Fungal ecology"}],"tags":[],"updatedAt":"2025-11-03T15:59:57+00:00","versionOfRecord":{"articleIdentity":"rs-6196330","link":"https://doi.org/10.1038/s41598-025-22071-6","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2025-10-31 15:57:12","publishedOnDateReadable":"October 31st, 2025"},"versionCreatedAt":"2025-04-14 16:25:47","video":"","vorDoi":"10.1038/s41598-025-22071-6","vorDoiUrl":"https://doi.org/10.1038/s41598-025-22071-6","workflowStages":[]},"version":"v1","identity":"rs-6196330","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6196330","identity":"rs-6196330","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}