Biology subjects

Lovett, B.

Publications and source records attributed to Lovett, B..

4 recordsLinked to original sources

bioRxiv2023

Prevalence and diversity of TAL effector-like proteins in fungal endosymbiotic Mycetohabitans spp.

Endofungal Mycetohabitans (formerly Burkholderia) spp. rely on a type III secretion system to deliver mostly unidentified effector proteins when colonizing their host fungus, Rhizopus microsporus. The one known secreted effector family from Mycetohabitans consists of homologs of transcription activator-like (TAL) effectors, which are used by plant pathogenic Xanthomonas and Ralstonia spp. to activate host genes that promote disease. These Burkholderia TAL-like (Btl) proteins bind corresponding specific DNA sequences in a predictable manner, but their genomic target(s) and impact on transcription in the fungus are unknown. Recent phenotyping of Btl mutants of two Mycetohabitans strains revealed that the single Btl in one M. endofungorum strain enhances fungal membrane stress tolerance, while others in a M. rhizoxinica strain promote bacterial colonization of the fungus. The phenotypic diversity underscores the need to assess the sequence diversity and, given that sequence diversity translates to DNA targeting specificity, the functional diversity of Btl proteins. Using a dual approach to maximize capture of Btl protein sequences for our analysis, we sequenced and assembled nine Mycetohabitans spp. genomes using long-read PacBio technology and also mined available short-read Illumina fungal-bacterial metagenomes. We show that btl genes are present across diverse Mycetohabitans strains from Mucoromycota fungal hosts yet vary in sequences and predicted DNA binding specificity. Phylogenetic analysis revealed distinct clades of Btl proteins and suggested that Mycetohabitans might contain more species than previously recognized. Within our data set, Btl proteins were more conserved across Mycetohabitans rhizoxinica strains than across Mycetohabitans endofungorum, but there was also evidence of greater overall strain diversity within the latter clade. Overall, the results suggest that Btl proteins contribute to bacterial-fungal symbioses in myriad ways. Impact StatementMany Mucoromycota fungi harbor endosymbiotic bacteria, including Rhizopus spp. that are food fermenters and pathogens of plants and immunocompromised people. Rhizopus microsporus has endofungal Mycetohabitans (formerly Burkholderia) spp. that deploy proteins related to DNA-binding transcription activator-like effectors of plant pathogens, which enter plant nuclei and activate disease susceptibility genes. By sequencing isolated bacteria and mining fungal holobiont sequences, we found Btl proteins in diverse Mycetohabitans strains, varying in predicted DNA binding specificity, thus in potential host targets. Btl proteins were more conserved within M. rhizoxinica, suggesting distinctions among the two named species. The results in the context of phenotypic differences observed in other studies suggest that Btl proteins contribute to symbiosis in diverse ways, providing insight into effector evolution and arguing for functional characterization of additional Btl proteins to understand establishment and maintenance of these important fungal-bacterial interactions.

microbiology↗

bioRxiv2023

Signatures of transposon-mediated genome inflation, host specialization, and photoentrainment in Entomophthora muscae and allied entomophthoralean fungi

Despite over a century of observations, the obligate insect parasites within the order Entomophthorales remain poorly characterized at the genetic level. This is in part due to their large genome sizes and difficulty in obtaining sequenceable material. In this manuscript, we leveraged a recently-isolated, laboratory-tractable Entomophthora muscae isolate and improved long-read sequencing to obtain a largely-complete entomophthoralean genome. Our E. muscae assembly is 1.03 Gb, consists of 7,810 contigs and contains 81.3% complete fungal BUSCOs. Using a comparative approach with other available (transcriptomic and genomic) datasets from entomophthoralean fungi, we provide new insight into the biology of these understudied pathogens. We offer a head-to-head comparison of morphological and molecular data for species within the E. muscae species complex. Our findings suggest that substantial taxonomic revision is needed to define species within this group and we provide recommendations for differentiating strains and species in the context of the existing body of E. muscae scientific literature. We show that giant genomes are the norm within Entomophthoraceae owing to extensive, but not recent, Ty3 retrotransposon activity, despite the presence of machinery to defend against transposable elements(RNAi). In addition, we find that E. muscae and its closest allies are enriched for M16A peptidases and possess genes that are likely homologs to the blue-light sensor white-collar 1, a Neurospora crassa gene that has a well-established role in maintaining circadian rhythms. We find that E. muscae has an expanded group of acid-trehalases, consistent with trehalose being the primary sugar component of fly (and insect) hemolymph. We uncover evidence that E. muscae diverged from other entomophthoralean fungi by expansion of existing families, rather than loss of particular domains, and possesses a potentially unique suite of secreted catabolic enzymes, consistent with E. muscaes species-specific, biotrophic lifestyle. Altogether, we provide a genetic and molecular foundation that we hope will provide a platform for the continued study of the unique biology of entomophthoralean fungi.

genomics↗

bioRxiv2022

Conoideocrella luteorostrata (Hypocreales: Claviciptaceae), a potential biocontrol fungus for elongate hemlock scale in United States Christmas tree production areas

The entomopathogenic fungus Conoideocrella luteorostrata (Hypocreales: Clavicipitaceae) has recently been implicated in natural epizootics among exotic elongate hemlock scale (EHS) insects in Fraser fir Christmas tree farms in the eastern U.S. Since 1913, asexual populations of C. luteorostrata have been reported from various plant-feeding Hemiptera in the southeastern U.S., but a thorough morphological and phylogenetic examination of the species, particularly detailed characterization of populations involved in recent epizootics in EHS, are lacking. The recovery of multiple strains of C. luteorostrata from mycosed EHS cadavers collected in Ashe County North Carolina provided an opportunity to conduct pathogenicity assays and morphological and phylogenetic studies to investigate genus- and species-level boundaries among members of the Clavicipitaceae. Pathogenicity assays confirmed C. luteorostrata causes mortality of EHS first instar crawlers, an essential first step in developing C. luteorostrata as a biocontrol. The results of the morphological study failed to recover a sexual stage from EHS cadavers or pure cultures, but revealed conidia aligned with previous measurements of the paecilomyces-like asexual state of C. luteorostrata (6.9 {micro}m x 2.6 {micro}m average), with colony and conidiophore morphology consistent with previously reported observations. Additionally, a hirsutella-like synanamorph of C. luteorostrata was observed for the first time under specific lab conditions. In both a four-locus, 54-taxa Clavicipitaceae-wide phylogenetic analysis including the D1-D2 domains of the nuclear 28S rRNA gene (28S), elongation factor 1 alpha (EF1-), DNA-directed RNA polymerase II subunit 1 (RPB1) and DNA-directed RNA polymerase II subunit 2 (RPB2) and a two-locus, 38-taxa (28S & EF1-) phylogenetic analysis, C. luteorostrata, C. tenuis and C. krungchingensis were resolved as strongly supported monophyletic lineages across all loci and both methods (maximum likelihood & Bayesian inference) of phylogenetic inference with the exception of 28S for C. tenuis. Despite the strong support for individual Conoideocrella species, none of the analyses supported the monophyly of the genus, with the inclusion of Dussiella. Due to the paucity of publicly available RPB1 and RPB2 sequence data for Conoideocrella, EF1- provided superior delimitation of intraspecies groupings for C. luteorostrata and C. tenuis and should be used in future studies. Further development of C. luteorostrata as a biocontrol agent against EHS both in Christmas tree farms and surrounding hemlock forests will require additional surveys across diverse Hemiptera and expanded pathogenicity testing to better understand host range and efficacy of this fungus.

microbiology↗

bioRxiv2022

Ecology drives the observed spectrum of hydrophobin protein diversity across Kingdom Fungi

1.Hydrophobins mediate the interactions between fungi and the elements of their ecosystem via assembly at interfaces serving a wide range of diverse functions. As such, these proteins can be seen as a means by which fungi not only adapt to a pre-existing environment, but also actively participate in the construction of their own ecological niches. Through this lens, we provide an expansive hydrophobin survey across the ecological breadth of Kingdom Fungi and advance the view that hydrophobins are best defined as a generic molecular structure with shared core structural features that accommodate a remarkable diversity of amino acid sequences. We examine the relationship between hydrophobin sequences, fungus phylogeny, and associated ecology from 45 fungal proteomes predicted from genomes spanning eight phyla and more than 25 orders. To capture the full spectrum of the hydrophobin amino acid sequence space mapped by our study, we describe the family as a continuum of overlapping hidden Markov models (HMMs), each HMM representing clusters of sequence similarity spanning existing hydrophobin classes. Overall, our approach uncovered ecology as a major driver of hydrophobin diversification, further expanded the known hydrophobins beyond Dikarya, and uncovered evidence extending the possibilities for their function from exclusively extracellular to include intracellular. In addition, we identified novel core groups of cysteine-rich proteins whose conservation across fungi suggest they play key ecological roles. Together, our work offers an ontological framework that captures the diversity of hydrophobin amino acid sequences and highlights the need to revisit challenging fundamental questions regarding hydrophobins to achieve a mechanistic understanding of their function as emerging from assembly within an ecosystem.

microbiology↗