Mostafa Rahnama

Alternaria leaf spot is an important foliar disease of cotton ( Gossypium hirsutum L.) and has historically been associated primarily with Alternaria macrospora in the United States. In November 2025, symptomatic Upland cotton leaves from a research greenhouse at Tennessee Technological University’s Oakley Farm in Livingston, Tennessee, yielded two Alternaria isolates, 2-2 and 9-1. Quantitative conidial characterization showed substantial overlap in dimensions, septation, and shape. Concatenated ITS- RPB2 phylogenetic analysis supported different broad affinities, placing isolate 2-2 within an A. alternata -dominated group and isolate 9-1 within a group associated with the A. arborescens species complex (AASC), although their immediate relationships were weakly resolved. Phylogenomic analysis of 1,501 single-copy BUSCO orthologs from 63 genomes placed isolate 2-2 within the A. alternata lineage, sister to BMP2343, and isolate 9-1 within the AASC, sister to MOD1-FUNGI6. In controlled pathogenicity assays, isolate 9-1 reproduced foliar lesions on cotton, whereas mock-inoculated plants remained symptomless, and molecularly confirmed re-isolation fulfilled Koch’s postulates. Isolate 2-2 produced insufficient conidia under the culture conditions used to prepare a standardized inoculum, and its pathogenicity was therefore not determined. These results reveal phylogenetically distinct Alternaria lineages associated with symptomatic cotton and establish an AASC isolate as a causal agent of cotton leaf spot in Tennessee.

Motivation: Centromeres are essential chromosomal loci, yet their computational identification remains challenging due to rapid sequence evolution, high repeat content, and the absence of conserved defining motifs. This challenge is particularly pronounced in fungi, where centromere architectures vary widely in size, sequence composition, and chromatin organization, limiting the effectiveness of single-feature or motif-based prediction approaches. Results: We present CentroFinder, a fungal-specific computational framework for de novo centromere prediction from long-read sequencing–based genome assemblies. CentroFinder integrates multiple genomic and long-read–derived features into a weighted scoring model to identify loci where centromere-associated signals converge. Benchmarking against experimentally mapped centromeres in Cryptococcus deuterogattii, Magnaporthe oryzae , and Neurospora crassa demonstrates that CentroFinder consistently predicts a single centromeric region per chromosome, fully nested within CENP-A–defined domains despite substantial diversity in centromere size, sequence composition, and chromatin context. Availability and Implementation: CentroFinder is freely available as open-source software at https://github.com/RahnamaLab/CentroFinder . The pipeline is designed for high-performance computing environments and leverages features derived from long-read sequencing data.

Motivation: Centromeres are essential chromosomal loci, yet their computational identification remains challenging due to rapid sequence evolution, high repeat content, and the absence of conserved defining motifs. This challenge is particularly pronounced in fungi, where centromere architectures vary widely in size, sequence composition, and chromatin organization, limiting the effectiveness of single-feature or motif-based prediction approaches. Results: We present CentroFinder, a fungal-specific computational framework for de novo centromere prediction from long-read sequencing–based genome assemblies. CentroFinder integrates multiple genomic and long-read–derived features into a weighted scoring model to identify loci where centromere-associated signals converge. Benchmarking against experimentally mapped centromeres in Cryptococcus deuterogattii, Magnaporthe oryzae, and Neurospora crassa showed that 27 of 28 predicted intervals overlapped the corresponding experimental domains, yielding 96.4% chromosome-level detection sensitivity. Application to 11 additional fungal genomes produced one contiguous predicted centromeric region per chromosome, supporting the transferability of the workflow for chromosome-level centromere prediction. Availability and implementation: CentroFinder is freely available as open-source software at https://github.com/RahnamaLab/CentroFinder. The pipeline is designed for high-performance computing environments and leverages features derived from long-read sequencing data.

Transposable elements (TEs) are dynamic components of eukaryotic genomes, playing a crucial role in genome evolution and plasticity, particularly in unstable regions such as chromosome ends. In the globally significant fungal pathogen Fusarium oxysporum , we identified and characterized a novel family of non-LTR retrotransposons named FoTeRs ( F. oxysporum Telomeric Retrotransposons). These elements are consistently and uniquely localized at chromosome ends, representing a rare example of site-specific TE integration. Phylogenetic analysis confirmed that FoTeRs form a distinct clade with other telomere-targeting retrotransposons, suggesting a shared evolutionary history and a convergent mechanism for telomeric integration. We found that individual FoTeR elements exhibit a duality in their evolutionary status. Putatively functional elements are under strong purifying selection, indicating that their protein-coding regions are highly conserved. This contrasts with the presence of other, non-functional copies that exhibit signs of mutational decay, a process accelerated by Repeat-Induced Point (RIP) mutations —a fungal-specific defense mechanism. The high density of upstream variable number tandem repeats (VNTRs) also contributes to their genomic plasticity. Furthermore, FoTeRs frequently co-localize with host Telomere-Linked Helicases (TLHs), suggesting a potential functional link in telomere maintenance. This study provides crucial insights into the role of TEs in shaping the genome architecture and adaptive potential of this important fungal pathogen.

SUMMARY: Functional genomic studies rely on controlled gene expression, which is challenging for essential genes or those active only under specific conditions. We provide a detailed protocol for generating Magnaporthe oryzae (synonym of Pyricularia oryzae ) with conditional BUF1 gene expression. This method uses targeted promoter replacement to substitute the native BUF1 promoter with the nitrogen-responsive promoter. This approach enables nitrogen source-dependent control of expression in M. oryzae , leading to observable pigmentation changes that facilitate easy phenotypic screening.

Epichloë species are systemic, often seed-transmissible symbionts (endophytes) of cool-season grasses (Poaceae subfam. Poöideae) that produce up to four classes of bioprotective alkaloids. Whereas haploid Epichloë species may reproduce sexually and transmit between host plants (horizontally), many Epichloë species are polyploid hybrids that are exclusively transmitted via seeds (vertically). Therefore, the generation of, and selection on, chemotypic (alkaloid) profiles and diversity should differ between haploids and hybrids. We undertook a genome-level analysis of haploids and polyploid hybrids, emphasizing hybrids that produce lolines, which are potent broad-spectrum anti-invertebrate alkaloids that can accumulate to levels up to 2% of plant dry mass. Prior phylogenetic analysis had indicated that loline alkaloid gene clusters (LOL) in many hybrids are from the haploid species Epichloë bromicola, but no LOL-containing E. bromicola strains were previously identified. We discovered LOL-containing E. bromicola from host grasses Bromus tomentellus and Melica persica in a Mediterranean-climate region (MCR) in Isfahan Province, Iran, and from Thinopyrum intermedium in Poland. The isolates from B. tomentellus and M. persica were closely related and had nearly identical alkaloid gene profiles, and their LOL clusters were most closely related to those of several Epichloë hybrids. In contrast, several LOL genes in the isolate from T. intermedium were phylogenetically more basal in genus Epichloë, indicating trans-species polymorphism. While identifying likely hybrid ancestors, this study also revealed novel host ranges in central Iran, with the first observation of E. bromicola in host tribe Meliceae and of Epichloë festucae in host tribe Bromeae. We discuss the possibility that MCRs may be hotspots for diversification of grass-Epichloë symbioses via extended host ranges and interspecific hybridization of the symbionts.

Telomere-linked RecQ helicase (tlh) genes have been reported in several fungi and a choanoflagellate in the regions adjacent to the terminal telomere repeats. In this study, we explored the Telomere-linked RecQ helicase (tlh) genes in four strains of Fusarium oxysporum, offering new insights into their genomic structure, functional motifs, and impact on chromosomal ends. We conducted a comprehensive analysis, comparing the tlh genes of F. oxysporum with those previously identified in other organisms and uncovering significant similarities. Through comparative genomics, we identified conserved protein motifs across these genes, including a TLH domain, C₂H₂, and RecQ helicase motifs. Our phylogenetic analysis positions the F. oxysporum tlh genes in a cluster with other known tlhs, suggesting a shared evolutionary origin. Mutation analysis revealed a relatively low level of deleterious mutations in tlh gene paralogs, with a notable proportion of full-size structures maintained across strains. Analysis of subtelomeric sequences indicates that a region with almost identical sequences flanks the majority of chromosome ends, termed tlh-containing region (TLHcr), across these strains. The presence of TLHcrs at chromosome ends, either as single entities or in arrays, underscores their potential role in telomere function and genome stability. Our findings provide a detailed examination of tlh genes in four strains of F. oxysporum, laying the groundwork for future studies on their biological significance and evolutionary history in fungal genomes.

Telomere-linked RecQ helicase (TLH) genes have been reported in several fungi and a choanoflagellate in the regions adjacent to the terminal telomere repeats. In this study, we explore the Telomere-linked RecQ helicase (TLH) genes in four strains of Fusarium oxysporum , offering new insights into their genomic structure, functional motifs, and impact on chromosomal ends. We conducted a comprehensive analysis, comparing the TLH genes of F. oxysporum with those previously identified in other organisms and uncovering significant similarities. Through comparative genomics, we identified conserved protein motifs across these genes, including a TLH domain, C ₂ H ₂ , and RecQ helicase motifs. Our phylogenetic analysis positions the F. oxysporum TLH genes in a cluster with other known TLHs, suggesting a shared evolutionary origin. Mutation analysis revealed a relatively low level of deleterious mutations in TLH gene paralogs, with a notable proportion of full-size structures maintained across strains. Analysis of subtelomeric sequences indicates that a region with almost identical sequences flanks the majority of chromosome ends, termed TLH-containing region (TLHcr), across these strains. The presence of TLHcrs at chromosome ends, either as single entities or in arrays, underscores their potential role in telomere function and genome stability. Our findings provide a detailed examination of TLH genes in four strains of F. oxysporum , laying the groundwork for future studies on their biological significance and evolutionary history in fungal genomes.

Wheat blast, caused by the Pyricularia oryzae Triticum lineage (PoT), first emerged in Brazil and quickly spread to neighboring countries. Its recent appearance in Bangladesh and Zambia highlights a need to understand the disease's population biology and epidemiology so as to mitigate pandemic outbreaks. Current knowledge is mostly based on characterizations of Brazilian wheat blast isolates and comparison with isolates from non-wheat, endemic grasses. These foregoing studies concluded that the wheat blast population lacks host specificity and, as a result, undergoes extensive gene flow with populations infecting non-wheat hosts. Additionally, based on genetic similarity between wheat blast and isolates infecting Urochloa species, it was proposed that the disease originally emerged via a host jump from this grass and that Urochloa likely plays a central role in wheat blast epidemiology owing to its widespread use as a pasture grass. However, due to inconsistencies with broader phylogenetic studies, we suspected that these seminal studies had not actually sampled the populations normally found on endemic grasses and, instead, had repeatedly isolated members of PoT and the related Lolium pathogen lineage (PoL1). Re-analysis of the Brazilian data as part of a comprehensive, global, phylogenomic dataset that included a small number of South American isolates sampled away from wheat confirmed our suspicion and identified four new P. oryzae lineages on grass hosts. As a result, the conclusions underpinning current understanding in wheat blast's evolution, population biology, and epidemiology are unsubstantiated and could be equivocal.