Endophytic species of Colletotrichum associated with cashew tree in northeastern Brazil.

Anthracnose caused by Colletotrichum is the most severe and widely occurring cashew disease in Brazil. Colletotrichum species are commonly found as pathogens, endophytes and occasionally as saprophytes in a wide range of hosts. The endophytic species associated with cashew trees are poorly studied. In this study, we report the Colletotrichum endophytic species associated with cashew trees in two locations in the state of Pernambuco, their prevalence in different plant organs (leaves, veins, branches and inflorescences), and compare the species in terms of pathogenicity and aggressiveness using different inoculation methods (wounded × unwounded). Six species of Colletotrichum were identified according to multilocus phylogenetic analyses, including Colletotrichum asianum, Colletotrichum chrysophilum, Colletotrichum karsti, Colletotrichum siamense, Colletotrichum theobromicola, and Colletotrichum tropicale. There were differences in the percentage of isolation in relation to the prevalence of colonized tissues and collection locations. C. tropicale was the prevalent species in both geographic areas and plant tissues collected, with no pattern of distribution of species between areas and plant tissues. All isolates were pathogenic in injured tissues of cashew plants. The best method to test the pathogenicity of Colletotrichum species was utilizing the combination of leaves + presence of wounds + conidial suspension, as it better represents the natural infection process. C. siamense was the most aggressive species.

Selection of stable reference genes for qPCR expression of Colletotrichum lindemuthianum, the bean anthracnose pathogen.

Phaseolus vulgaris L., commonly known as the common bean, is a highly nutritious crop often called the "poor man's meat". However, it is susceptible to various diseases throughout the cropping season, with anthracnose caused by Colletotrichum lindemuthianum being a significant threat that leads to substantial losses. There is still a lack of understanding about the molecular basis of C. lindemuthianum pathogenicity. The first step in understanding this is to identify pathogenicity genes that express more during infection of common beans. A reverse transcription quantitative real-time PCR (qPCR) method can be used for virulence gene expression. However, this approach requires selecting appropriate reference genes to normalize relative gene expression data. Currently, there is no reference gene available for C. lindemuthianum. In this study, we selected eight candidate reference genes from the available genome of C. lindemuthianum to bridge the gap. These genes were ACT (Actin), ß-tub (ß-tubulin), EF (Elongation Factor), Cyt C (Cytochrome C), His H3 (Histone H3), CHS1 (Chitin synthetase), GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) and abfA (Alpha-l-Arabinofuranosidase A). The primers for these candidate reference genes were able to amplify cDNA only from the pathogen, demonstrating their specificity. The qPCR efficiency of the primers ranged from 80% to 103%. We analyzed the stability of gene expression in C. lindemuthianum by exposing the mycelium to nine different stress conditions. We employed algorithms, such as GeNorm, NormFinder, BestKeeper, and RefFinder tools, to identify the most stable gene. The analysis using these tools revealed that EF, GAPDH, and ß-tub most stable genes, while ACT and CHS1 showed relatively low expression stability. A large number of potential effector genes have been identified through bioinformatics analysis in C. lindemuthianum. The stable genes for qPCR (EF and GAPDH) discovered in this study will aid the scientific community in determining the relative expression of C. lindemuthianum effector genes.

Unveiling novel Neocosmospora species from Thai mangroves as potent biocontrol agents against Colletotrichum species.

AIMS: Neocosmospora species are saprobes, endophytes, and pathogens belonging to the family Nectriaceae. This study aims to investigate the taxonomy, biosynthetic potential, and application of three newly isolated Neocosmospora species from mangrove habitats in the southern part of Thailand using phylogeny, bioactivity screening, genome sequencing, and bioinformatics analysis. METHODS AND RESULTS: Detailed descriptions, illustrations, and a multi-locus phylogenetic tree with large subunit ribosomal DNA (LSU), internal transcribed spacer (ITS), translation elongation factor 1-alpha (ef1-α), and RNA polymerase II second largest subunit (RPB2) regions showing the placement of three fungal strains, MFLUCC 17-0253, MFLUCC 17-0257, and MFLUCC 17-0259 clustered within the Neocosmospora clade with strong statistical support. Fungal crude extracts of the new species N. mangrovei MFLUCC 17-0253 exhibited strong antifungal activity to control Colletotrichum truncatum CG-0064, while N. ferruginea MFLUCC 17-0259 exhibited only moderate antifungal activity toward C. acutatum CC-0036. Thus, N. mangrovei MFLUCC 17-0253 was sequenced by Oxford nanopore technology. The bioinformatics analysis revealed that 49.17 Mb genome of this fungus harbors 41 potential biosynthetic gene clusters. CONCLUSION: Two fungal isolates of Neocosmospora and a new species of N. mangrovei were reported in this study. These fungal strains showed activity against pathogenic fungi causing anthracnose in chili. In addition, full genome sequencing and bioinformatics analysis of N. mangrovei MFLUCC 17-0253 were obtained.

Genome comparisons reveal accessory genes crucial for the evolution of apple Glomerella leaf spot pathogenicity in Colletotrichum fungi.

Apple Glomerella leaf spot (GLS) is an emerging fungal disease caused by Colletotrichum fructicola and other Colletotrichum species. These species are polyphyletic and it is currently unknown how these pathogens convergently evolved to infect apple. We generated chromosome-level genome assemblies of a GLS-adapted isolate and a non-adapted isolate in C. fructicola using long-read sequencing. Additionally, we resequenced 17 C. fructicola and C. aenigma isolates varying in GLS pathogenicity using short-read sequencing. Genome comparisons revealed a conserved bipartite genome architecture involving minichromosomes (accessory chromosomes) shared by C. fructicola and other closely related species within the C. gloeosporioides species complex. Moreover, two repeat-rich genomic regions (1.61 Mb in total) were specifically conserved among GLS-pathogenic isolates in C. fructicola and C. aenigma. Single-gene deletion of 10 accessory genes within the GLS-specific regions of C. fructicola identified three that were essential for GLS pathogenicity. These genes encoded a putative non-ribosomal peptide synthetase, a flavin-binding monooxygenase and a small protein with unknown function. These results highlight the crucial role accessory genes play in the evolution of Colletotrichum pathogenicity and imply the significance of an unidentified secondary metabolite in GLS pathogenesis.

The CsPbs2-interacting protein oxalate decarboxylase CsOxdC3 modulates morphosporogenesis, virulence, and fungicide resistance in Colletotrichum siamense.

The HOG MAPK pathway mediates diverse cellular and physiological processes, including osmoregulation and fungicide sensitivity, in phytopathogenic fungi. However, the molecular mechanisms underlying HOG MAPK pathway-associated stress homeostasis and pathophysiological developmental events are poorly understood. Here, we demonstrated that the oxalate decarboxylase CsOxdC3 in Colletotrichum siamense interacts with the protein kinase kinase CsPbs2, a component of the HOG MAPK pathway. The expression of the CsOxdC3 gene was significantly suppressed in response to phenylpyrrole and tebuconazole fungicide treatments, while that of CsPbs2 was upregulated by phenylpyrrole and not affected by tebuconazole. We showed that targeted gene deletion of CsOxdC3 suppressed mycelial growth, reduced conidial length, and triggered a marginal reduction in the sporulation characteristics of the ΔCsOxdC3 strains. Interestingly, the ΔCsOxdC3 strain was significantly sensitive to fungicides, including phenylpyrrole and tebuconazole, while the CsPbs2-defective strain was sensitive to tebuconazole but resistant to phenylpyrrole. Additionally, infection assessment revealed a significant reduction in the virulence of the ΔCsOxdC3 strains when inoculated on the leaves of rubber tree (Hevea brasiliensis). From these observations, we inferred that CsOxdC3 crucially modulates HOG MAPK pathway-dependent processes, including morphogenesis, stress homeostasis, fungicide resistance, and virulence, in C. siamense by facilitating direct physical interactions with CsPbs2. This study provides insights into the molecular regulators of the HOG MAPK pathway and underscores the potential of deploying OxdCs as potent targets for developing fungicides.

Complete genome sequence of a novel alternavirus isolated from the phytopathogenic fungus Colletotrichum fioriniae.

A novel double-strand RNA (dsRNA) mycovirus, named "Colletotrichum fioriniae alternavirus1" (CfAV1), was isolated from the strain CX7 of Colletotrichum fioriniae, the causal agent of walnut anthracnose. The complete genome of CfAV1 is composed of three dsRNA segments: dsRNA1 (3528 bp), dsRNA2 (2485 bp), and dsRNA3 (2481 bp). The RNA-dependent RNA polymerase (RdRp) is encoded by dsRNA1, while both dsRNA2 and dsRNA3 encode hypothetical proteins. Based on multiple sequence alignments and phylogenetic analysis, CfAV1 is identified as a new member of the family Alternaviridae. This is the first report of an alternavirus that infects the phytopathogenic fungus C. fioriniae.

Transcriptomic and Physiological Analysis of the Effects of Exogenous Phloretin and Pterostilbene on Resistance Responses of Stylosanthes against Anthracnose.

Anthracnose caused by Colletotrichum gloeosporioides is a destructive disease of Stylosanthes (stylo). Combination treatment of phloretin and pterostilbene (PP) has been previously shown to effectively inhibit the conidial germination and mycelial growth of C. gloeosporioides in vitro. In this study, the effects of PP treatment on the growth of C. gloeosporioides in vivo and the biocontrol mechanisms were investigated. We found that exogenous PP treatment could limit the growth of C. gloeosporioides and alleviate the damage of anthracnose in stylo. Comparative transcriptome analysis revealed that 565 genes were up-regulated and 239 genes were down-regulated upon PP treatment during the infection by C. gloeosporioides. The differentially expressed genes were mainly related to oxidative stress and chloroplast organization. Further physiological analysis revealed that application of PP after C. gloeosporioides inoculation significantly reduced the accumulation of O2•- level and increased the accumulation of antioxidants (glutathione, ascorbic acid and flavonoids) as well as the enzyme activity of total antioxidant capacity, superoxide dismutase, catalase, glutathione reductase, peroxidase and ascorbate peroxidase. PP also reduced the decline of chlorophyll a + b and increased the content of carotenoid in response to C. gloeosporioides infection. These results suggest that PP treatment alleviates anthracnose by improving antioxidant capacity and reducing the damage of chloroplasts, providing insights into the biocontrol mechanisms of PP on the stylo against anthracnose.

The galactose metabolism genes UGE1 and UGM1 are novel virulence factors of the maize anthracnose fungus Colletotrichum graminicola.

Fungal cell walls represent the frontline contact with the host and play a prime role in pathogenesis. While the roles of the cell wall polymers like chitin and branched ß-glucan are well understood in vegetative and pathogenic development, that of the most prominent galactose-containing polymers galactosaminogalactan and fungal-type galactomannan is unknown in plant pathogenic fungi. Mining the genome of the maize pathogen Colletotrichum graminicola identified the single-copy key galactose metabolism genes UGE1 and UGM1, encoding a UDP-glucose-4-epimerase and UDP-galactopyranose mutase, respectively. UGE1 is thought to be required for biosynthesis of both polymers, whereas UGM1 is specifically required for fungal-type galactomannan formation. Promoter:eGFP fusion strains revealed that both genes are expressed in vegetative and in pathogenic hyphae at all stages of pathogenesis. Targeted deletion of UGE1 and UGM1, and fluorescence-labeling of galactosaminogalactan and fungal-type galactomannan confirmed that Δuge1 mutants were unable to synthesize either of these polymers, and Δugm1 mutants did not exhibit fungal-type galactomannan. Appressoria of Δuge1, but not of Δugm1 mutants, were defective in adhesion, highlighting a function of galactosaminogalactan in the establishment of these infection cells on hydrophobic surfaces. Both Δuge1 and Δugm1 mutants showed cell wall defects in older vegetative hyphae and severely reduced appressorial penetration competence. On intact leaves of Zea mays, both mutants showed strongly reduced disease symptom severity, indicating that UGE1 and UGM1 represent novel virulence factors of C. graminicola.

What's in my Pot? Six Colletotrichum Species Causing Anthracnose in Brazilian Pecan Orchards.

Pecan (Carya illinoinensis) is one important exotic forest crop cultivated in South America, specifically in Brazil, Uruguay, and Argentina. However, diseases such as anthracnose, favored by high humidity conditions and high summer temperatures, make its cultivation difficult, causing important loss to pecan farmers. This study used morphological and molecular approaches to identify the Colletotrichum species causing anthracnose in pecan plantations in Southern Brazil. The isolates obtained from pecan fruits with anthracnose symptoms were grouped through quantitative morphological characteristics into three distinct morphotypes. Molecular analysis of nuclear genes allowed the identification of six species of Colletotrichum causing anthracnose in pecan: C. nymphaeae, C. fioriniae, C. gloeosporioides, C. siamense, C. kahawae, and C. karsti. Three of these species are reported for the first time as causal agents of anthracnose in pecan. Therefore, these results provide an important basis for the adoption and/or development of anthracnose management strategies in pecan orchards cultivated in southern Brazil and neighboring countries.

Evolutionary history of the cytochrome P450s from Colletotrichum species and prediction of their putative functional roles during host-pathogen interactions.

The genomes of species belonging to the genus Colletotrichum harbor a substantial number of cytochrome P450 monooxygenases (CYPs) encoded by a broad diversity of gene families. However, the biological role of their CYP complement (CYPome) has not been elucidated. Here, we investigated the putative evolutionary scenarios that occurred during the evolution of the CYPome belonging to the Colletotrichum Graminicola species complex (s.c.) and their biological implications. The study revealed that most of the CYPome gene families belonging to the Graminicola s.c. experienced gene contractions. The reductive evolution resulted in species restricted CYPs are predominant in each CYPome of members from the Graminicola s.c., whereas only 18 families are absolutely conserved among these species. However, members of CYP families displayed a notably different phylogenetic relationship at the tertiary structure level, suggesting a putative convergent evolution scenario. Most of the CYP enzymes of the Graminicola s.c. share redundant functions in secondary metabolite biosynthesis and xenobiotic metabolism. Hence, this current work suggests that the presence of a broad CYPome in the genus Colletotrichum plays a critical role in the optimization of the colonization capability and virulence.

Response of Bacillus velezensis 83 to interaction with Colletotrichum gloeosporioides resembles a Greek phalanx-style formation: A stress resistant phenotype with antibiosis capacity.

Plant growth-promoting rhizobacteria, such as Bacillus spp., establish beneficial associations with plants and may inhibit the growth of phytopathogenic fungi. However, these bacteria are subject to multiple biotic stimuli from their competitors, causing stress and modifying their development. This work is a study of an in vitro interaction between two model microorganisms of socioeconomic relevance, using population dynamics and transcriptomic approaches. Co-cultures of Bacillus velezensis 83 with the phytopathogenic fungus Colletotrichum gloeosporioides 09 were performed to evaluate the metabolic response of the bacteria under conditions of non-nutritional limitation. The bacterial response was associated with the induction of a stress-resistant phenotype, characterized by a lower specific growth rate, but with antimicrobial production capacity. About 12% of co-cultured B. velezensis 83 coding sequences were differentially expressed, including the up-regulation of the general stress response (sigB regulon), and the down-regulation of alternative carbon sources catabolism (glucose preference). Defense strategies in B. velezensis are a determining factor in order to preserve the long-term viability of its population. Mostly, the presence of the fungus does not affect the expression of antibiosis genes, except for those corresponding to surfactin/bacillomycin D production. Indeed, the up-regulation of antibiosis genes expression is associated with bacterial growth, regardless of the presence of the fungus. This behavior in B. velezensis 83 resembles the strategy used by the classical Greek phalanx formation: by sacrificing growth rate and metabolic versatility, resources can be redistributed to defense (stress resistant phenotype) while maintaining the attack (antibiosis capacity). The presented results are the first characterization of the molecular phenotype at the transcriptome level of a biological control agent under biotic stress caused by a phytopathogen without nutrient limitation.

A novel partitivirus conferring hypovirulence by affecting vesicle transport in the fungus Colletotrichum.

Colletotrichum spp. are economically important phytopathogenic fungi that cause anthracnose in a variety of plant species worldwide. Hypovirulence-associated mycoviruses provide new options for the biological control of plant fungal diseases. Here, we found a novel partitivirus from Colletotrichum alienum and named it Colletotrichum alienum partitivirus 1 (CaPV1). CaPV1 contained two dsRNA segments encoding an RNA-dependent RNA polymerase and a capsid protein and was classified under the genus Gammapartitivirus of the family Partitiviridae. CaPV1 significantly decreased host virulence, mycelial growth, appressorial development, and appressorium turgor but increased conidial production with abnormal morphology. In addition, CaPV1 could be successfully transfected into other Colletotrichum species, including C. fructicola, C. spaethianum, and C. gloeosporioides, and caused hypovirulence, indicating the broad application potential of this virus. CaPV1 caused significant transcriptional rewiring of the host fungus C. alienum. Notably, some genes related to vesicle transport in the CaPV1-infected strain were downregulated, consistent with the impaired endocytosis pathway in this fungus. When the Rab gene CaRab7, which is associated with endocytosis in vesicle transport, was knocked out, the virulence of the mutants was reduced. Overall, our findings demonstrated that CaPV1 has the potential to control anthracnose caused by Colletotrichum, and the mechanism by which Colletotrichum induces hypovirulence is caused by affecting vesicle transport.IMPORTANCEColletotrichum is a kind of economically important phytopathogenic fungi that cause anthracnose disease in a variety of plant species worldwide. We found a novel mycovirus of the Gammapartitivirus genus and Partitiviridae family from the phytopathogenic fungus Colletotrichum alienum and named it CaPV1. This study revealed that CaPV1 infection significantly decreased host virulence and fitness by affecting mycelial growth, appressorial development, and appressorium turgor. In addition, CaPV1 could also infect other Colletotrichum species, including C. fructicola, C. spaethianum, and C. gloeosporioides, by viral particle transfection and resulting in hypovirulence of these Colletotrichum species. Transcriptomic analysis showed that CaPV1 caused significant transcriptional rewiring of the host fungus C. alienum, especially the genes involved in vesicle transport. Moreover, endocytosis and gene knockout assays demonstrated that the mechanism underlying CaPV1-induced hypovirulence is, at least in part, caused by affecting the vesicle transport of the host fungus. This study provided insights into the mechanisms underlying the pathogenesis of Colletotrichum species and mycovirus-fungus interactions, linking the role of mycovirus and fungus vesicle transport systems in shaping fungal pathogenicity.

The fatty acid 2-hydroxylase CsSCS7 is a key hyphal growth factor and potential control target in Colletotrichum siamense.

SCS7 is a fatty acid 2-hydroxylase required for the synthesis of inositol phosphorylceramide but is not essential for normal growth in Saccharomyces cerevisiae. Here, we demonstrate that the Colletotrichum siamense SCS7 homolog CsSCS7 plays a key role in hyphal growth. The CsSCS7 deletion mutant showed strong hyphal growth inhibition, small conidia, and marginally reduced sporulation and also resulted in a sharp reduction in the full virulence and increasing the fungicide sensitivity. The three protein domains (a cytochrome b5 domain, a transmembrane domain, and a hydroxylase domain) are important to CsSCS7 protein function in hyphal growth. The fatty acid assay results revealed that the CsSCS7 gene is important for balancing the contents of multiple mid-long- and short-chain fatty acids. Additionally, the retarded growth and virulence of C. siamense ΔCsSCS7 can be recovered partly by the reintroduction of homologous sequences from Magnaporthe oryzae and Fusarium graminearum but not SCS7 of S. cerevisiae. In addition, the spraying of C. siamense with naked CsSCS7-double-stranded RNA (dsRNAs), which leads to RNAi, increases the inhibition of hyphal growth and slightly decreases disease lesions. Then, we used nano material Mg-Al-layered double hydroxide as carriers to deliver dsRNA, which significantly enhanced the control effect of dsRNA, and the lesion area was obviously reduced. These data indicated that CsSCS7 is an important factor for hyphal growth and affects virulence and may be a potential control target in C. siamense and even in filamentous plant pathogenic fungi.IMPORTANCECsSCS7, which is homologous to yeast fatty acid 2-hydroxylase SCS7, was confirmed to play a key role in the hyphal growth of Colletotrichum siamense and affect its virulence. The CsSCS7 gene is involved in the synthesis and metabolism of fatty acids. Homologs from the filamentous fungi Magnaporthe oryzae and Fusarium graminearum can recover the retarded growth and virulence of C. siamense ΔCsSCS7. The spraying of double-stranded RNAs targeting CsSCS7 can inhibit hyphal growth and reduce the disease lesion area to some extent. After using nano material Mg-Al layered double hydroxide as carrier, the inhibition rates were significantly increased. We demonstrated that CsSCS7 is an important factor for hyphal growth and affects virulence and may be a potential control target in C. siamense and even in filamentous plant pathogenic fungi.

Assessment of Candidate Reference Genes for Gene Expression Studies Using RT-qPCR in Colletotrichum fructicola from Litchi.

Litchi (Litchi chinensis Sonn.) is a tropical fruit originating from southern China that is currently cultivated in subtropical and tropical regions worldwide. Litchi anthracnose, caused by Colletotrichum fructicola, a dominant species of Colletotrichum spp., is an important disease of litchi that damages the fruits in fields and in post-harvest storage. Real-time quantitative PCR (RT-qPCR) is a common technique with which to detect the expression of and function of target genes quickly and precisely, and stable reference genes are crucial. However, there is no comprehensive information on suitable reference genes of C. fructicola present. Here, we designed eight candidate genes (GAPDH, α-tubulin, 18S, ß-tubulin, EF1a, TATA, RPS5, and EF3) using RefFinder software (programs: geNorm, ΔCt, BestKeeper, and NormFinder) to investigate their reliability in the detection of C. fructicola under five different treatments (fungal development stage, temperature, UV, culture medium, and fungicide). The results showed the optimal reference genes under different conditions: EF1a and α-tubulin for developmental stage; α-tubulin and ß-tubulin for temperature; α-tubulin and RPS5 for UV treatment; RPS5 and α-tubulin for culture medium; α-tubulin, GAPDH, and TATA for fungicide treatments. The corresponding expression patterns of HSP70 (Heat shock protein 70) were significantly different when the most and the least stable reference genes were selected when treated under different conditions. Our study provides the first detailed list of optimal reference genes for the analysis of gene expression in C. fructicola via RT-qPCR, which should be useful for future functional studies of target genes in C. fructicola.

Identification of mycoparasitism-related genes in Trichoderma harzianum T4 that are active against Colletotrichum musae.

Trichoderma harzianum is a well-known biological control agent (BCA) that shows great potential in controlling many pathogenic fungi. To screen for genes associated with mycoparasitism, we sequenced and analyzed the transcriptome of T. harzianum T4 grown in dual culture with Colletotrichum musae. We analyzed differentially expressed genes (DEGs) of Trichoderma harzianum T4 in three different culture periods: before contact (BC), during contact (C) and after contact (AC). A total of 1453 genes were significantly differentially expressed compared to when T. harzianum T4 was cultured alone. During the three periods of double culture of T. harzianum T4 with C. musae, 74, 516, and 548 genes were up-regulated, respectively, and 11, 315, and 216 genes were down-regulated, respectively. The DEGs were screened using GO and KEGG enrichment analyses combined with differential expression multiples. Six gene categories related to mycoparasitism were screened: (a) pathogen recognition and signal transduction, (b) hydrolases, (c) ribosomal proteins and secreted proteins, (d) multidrug-resistant proteins and transporters, (e) heat shock proteins and detoxification, and (f) oxidative stress and antibiotics-related genes. The expression levels of 24 up-regulated genes during T. harzianum T4's antagonistic interaction with C. musae were detected via real-time fluorescence quantitative PCR (RT-qPCR). This study provided information on the transcriptional expression of T. harzianum T4 against C. musae. These results may help us to further understand the mechanism of mycoparasitism, which can provide a potential molecular target for improving the biological control capacity of T. harzianum T4.

Responsiveness of Candidate Genes on CoPv01CDRK/PhgPv01CDRK Loci in Common Bean Challenged by Anthracnose and Angular Leaf Spot Pathogens.

Anthracnose (ANT) and angular leaf spot (ALS) are significant diseases in common bean, leading to considerable yield losses under specific environmental conditions. The California Dark Red Kidney (CDRK) bean cultivar is known for its resistance to multiple races of both pathogens. Previous studies have identified the CoPv01CDRK/PhgPv01CDRK resistance loci on chromosome Pv01. Here, we evaluated the expression levels of ten candidate genes near the CoPv01CDRK/PhgPv01CDRK loci and plant defense genes using quantitative real-time PCR in CDRK cultivar inoculated with races 73 of Colletotrichum lindemuthianum and 63-39 of Pseudocercospora griseola. Gene expression analysis revealed that the Phvul.001G246300 gene exhibited the most elevated levels, showing remarkable 7.8-fold and 8.5-fold increases for ANT and ALS, respectively. The Phvul.001G246300 gene encodes an abscisic acid (ABA) receptor with pyrabactin resistance, PYR1-like (PYL) protein, which plays a central role in the crosstalk between ABA and jasmonic acid responses. Interestingly, our results also showed that the other defense genes were initially activated. These findings provide critical insights into the molecular mechanisms underlying plant defense against these diseases and could contribute to the development of more effective disease management strategies in the future.

Identification of gene modules and hub genes associated with Colletotrichum siamense infection in mango using weighted gene co-expression network analysis.

Colletotrichum siamense is a hemibiotrophic ascomycetous fungus responsible for mango anthracnose. The key genes involved in C. siamense infection remained largely unknown. In this study, we conducted weighted gene co-expression network analysis (WGCNA) of RNA-seq data to mine key genes involved in Colletotrichum siamense-mango interactions. Gene modules of Turquoise and Salmon, containing 1039 and 139 respectively, were associated with C. siamense infection, which were conducted for further analysis. GO enrichment analysis revealed that protein synthesis, organonitrogen compound biosynthetic and metabolic process, and endoplasmic reticulum-related genes were associated with C. siamense infection. A total of 568 proteins had homologs in the PHI database, 370 of which were related to virulence. The hub genes in each module were identified, which were annotated as O-methyltransferase (Salmon) and Clock-controlled protein 6 (Turquoise). A total of 24 proteins exhibited characteristics of SCRPs. By using transient expression in Nicotiana benthamiana, the SCRPs of XM_036637681.1 could inhibit programmed cell death (PCD) that induced by BAX (BCL-2-associated X protein), suggesting that it may play important roles in C. siamense infection. A mango-C. siamense co-expression network was constructed, and the mango gene of XM_044632979.1 (auxin-induced protein 15A-like) was positively associated with 5 SCRPs. These findings help to deepen the current understanding of necrotrophic stage in C. siamense infection.

Polymorphisms at amino acid positions 85 and 86 in succinate dehydrogenase subunit C of Colletotrichum siamense: Implications for fitness and intrinsic sensitivity to SDHI fungicides.

Among succinate dehydrogenase inhibiter (SDHI) fungicides, penthiopyrad and benzovindiflupyr particularly inhibit Colletotrichum. Studying SDH amino acid polymorphism in Colletotrichum, along with its fungicide binding sites, is key to understanding their mechanisms of action. This study explores the SDH amino acid polymorphisms in Colletotrichum siamense strains from rubber trees in China and their interaction with SDHI fungicides, specifically penthiopyrad and benzovindiflupyr. Sequencing revealed most polymorphisms were in the SDHC subunit, particularly at positions 85 and 86, which are key to penthiopyrad resistance. Among 33 isolates, 33.3 % exhibited a substitution at position 85, and 9 % at position 86. A strain with W85L and T86N substitutions in SDHC showed reduced SDH activity, ATP content, mycelial growth, and virulence, and decreased sensitivity to penthiopyrad but not benzovindiflupyr. Molecular docking with Alphafold2 modeling suggested distinct binding modes of the two fungicides to C. siamense SDH. These findings underscore the importance of SDHC polymorphisms in C. siamense's fitness and sensitivity to SDHIs, enhancing our understanding of pathogen-SDHI interactions and aiding the development of novel SDHI fungicides.

Integrated transcriptome and proteome analyses unravel a series of early defence responses in Sarcandra glabra against Colletotrichum gloeosporioides.

Anthracnose caused by Colletotrichum gloeosporioides critically threatens the growth and commercial cultivation of Sarcandra glabra . However, the defence responses and underlying mechanisms remain unclear. Herein, we aimed to investigate the molecular reprogramming in S. glabra leaves infected with C. gloeosporioides . Leaf tissues at 0, 24 and 48h post-inoculation (hpi) were analysed by combining RNA sequencing and Tandem Mass Tag-based liquid chromatography with tandem mass spectrometry. In total, 18 441 and 25 691 differentially expressed genes were identified at 24 and 48hpi compared to 0hpi (uninoculated control), respectively. In addition, 1240 and 1570 differentially abundant proteins were discovered at 24 and 48hpi compared to 0hpi, respectively. Correlation analysis revealed that transcription and translation levels were highly consistent regarding repeatability and expression. Analyses using databases KEGG and iPATH revealed tricitric acid cycle, glycolysis/gluconeogenesis and phenylpropanoid biosynthesis were induced, whereas photosynthesis and tryptophan were suppressed. Enzymatic activity assay results were consistent with the upregulation of defence-related enzymes including superoxide dismutases, catalases, peroxidases and chitinases. The transcriptome expression results were additionally validated by quantitative real-time polymerase chain reaction analyses. This study provides insights into the molecular reprogramming in S. glabra leaves during infection, which lay a foundation for investigating the mechanisms of host-Colletotrichum interactions and breeding disease-resistant plants.

Taxonomy-guided selection of Paraburkholderia busanensis sp. nov.: a versatile biocontrol agent with mycophagy against Colletotrichum scovillei causing pepper anthracnose.

IMPORTANCE: Traditional control methods for postharvest diseases rely on fungicides, which cause human health and environmental concerns. This study introduces a taxonomy-guided strategy for selecting biocontrol agents. By focusing on Paraburkholderia group, which harbors diverse plant-beneficial strains, the inadvertent selection of harmful strains was circumvented, thereby obviating the need for laborious in vitro screening assays. A highly promising candidate, strain P39, has been identified, exhibiting remarkable biocontrol activity against Colletotrichum scovillei. Through comprehensive genomic, physiological, and biochemical analyses, P39 was characterized as a novel species within the Paraburkholderia genus and designated Paraburkholderia busanensis. Moreover, these findings deepen our understanding of bacterial-fungal interactions, as they elucidate a potential pathway for the utilization of fungal chitin, thereby enhancing our understanding of bacterial mycophagy. P. busanensis is a promising source of antifungal volatiles and putative novel secondary metabolites.