Association of water stress and Fusarium solani exacerbated Dalbergia sissoo dieback disease.

Globally, there is a growing concern about tree mortality due to harsh climates and changes in pest and disease patterns. However, experimental studies on the interactions between biotic and abiotic stresses in plants are relatively scarce. In this study, we investigated the interaction between Fusarium solani and water-stressed Dalbergia sissoo saplings. We postulated that under drought conditions, sissoo plants would become more susceptible to dieback infestation. Five fungi, including Fusarium oxysporum, Curvularia lunata, Cladophialophora carrionii, Alternaria alternaria, and Fusarium solani, were isolated from an old shisham tree showing advanced symptoms of dieback infestation. These fungi were identified based on their ITS sequence homology and spore characteristics. Dieback development was more pronounced in plants experiencing water stress, regardless of their predisposition or whether it occurred simultaneously. Lesions were more noticeable and longer in predisposed saplings (3.8cm), followed by simultaneous (2.4cm) and much smaller lesions in seedlings that were inoculated and well-watered (0.24cm). Progressive browning of the upper leaves, which lowers sapling height in predisposed, simultaneous, and well-watered inoculated saplings to 8.09 inches, 5.93 inches, and 17.42 inches, are typical dieback symptoms. Water stress causes the loss of chlorophyll a, b, and carotenoids, which reduces stomatal conductance, transpiration rate, and photosynthetic activity, leading to poor development and mortality. Similarly, predisposed, simultaneous, and well-watered inoculated seedlings expressed increased activity of CAT (22.57, 18.148, and 9.714 U/mg) and POD (3.0, 4.848, 1.246 U/mg), to reduce the damage caused by elevated levels of H2O2 expression. It is concluded that water stress is the main cause of dieback in shisham saplings that subsequently disposed of infected seedlings to secondary agents such as fungi and insects in the advanced stages of the dieback with prolonged drought stress. The lack of dieback in native populations is attributed to the absence of several ecological stresses, including water stress, extended droughts, waterlogging, and salinity. This study emphasizes the need for additional research into the effects of abiotic factors linked with fungal diseases on the long-term production and management of D. sissoo in Pakistan.

Genome-wide association study and molecular marker development for susceptibility to Gibberella ear rot in maize.

KEY MESSAGES: Sixty-nine quantitative trait nucleotides conferring maize resistance to Gibberella ear rot were detected, including eighteen novel loci. Four candidate genes were predicted, and four kompetitive allele-specific PCR markers were developed. Maize Gibberella ear rot (GER), caused by Fusarium graminearum, is one of the most devastating diseases in maize-growing regions worldwide. Enhancing maize cultivar resistance to this disease requires a comprehensive understanding of the genetic basis of resistance to GER. In this study, 334 maize inbred lines were phenotyped for GER resistance in five environments and genotyped using the Affymetrix CGMB56K SNP Array, and a genome-wide association study of resistance to GER was performed using a 3V multi-locus random-SNP-effect mixed linear model. A total of 69 quantitative trait nucleotides (QTNs) conferring resistance to GER were detected, and all of them explained individually less than 10% of the phenotypic variation, suggesting that resistance to GER is controlled by multiple minor-effect genetic loci. A total of 348 genes located around the 200-kb genomic region of these 69 QTNs were identified, and four of them (Zm00001d029648, Zm00001d031449, Zm00001d006397, and Zm00001d053145) were considered candidate genes conferring susceptibility to GER based on gene expression patterns. Moreover, four kompetitive allele-specific PCR markers were developed based on the non-synonymous variation of these four candidate genes and validated in two genetic populations. This study provides useful genetic resources for improving resistance to GER in maize.

Using Transcriptomics to Determine the Mechanism for the Resistance to Fusarium Head Blight of a Wheat-Th. elongatum Translocation Line.

Fusarium head blight (FHB), caused by the Fusarium graminearum species complex, is a destructive disease in wheat worldwide. The lack of FHB-resistant germplasm is a barrier in wheat breeding for resistance to FHB. Thinopyrum elongatum is an important relative that has been successfully used for the genetic improvement of wheat. In this study, a translocation line, YNM158, with the YM158 genetic background carrying a fragment of diploid Th. elongatum 7EL chromosome created using 60Co-γ radiation, showed high resistance to FHB under both field and greenhouse conditions. Transcriptome analysis confirmed that the horizontal transfer gene, encoding glutathione S-transferase (GST), is an important contributor to FHB resistance in the pathogen infection stage, whereas the 7EL chromosome fragment carries other genes regulated by F. graminearum during the colonization stage. Introgression of the 7EL fragment affected the expression of wheat genes that were enriched in resistance pathways, including the phosphatidylinositol signaling system, protein processing in the endoplasmic reticulum, plant-pathogen interaction, and the mitogen-activated protein kinase (MAPK) signaling pathway at different stages after F. graminearium infection. This study provides a novel germplasm for wheat resistance to FHB and new insights into the molecular mechanisms of wheat resistance to FHB.

Identification of hub genes and potential networks by centrality network analysis of PCR amplified Fusarium oxysporum f. sp. lycopersici EF1α gene.

BACKGROUND: Fusarium wilt is a devastating soil-borne fungal disease of tomato across the world. Conventional method of disease prevention including usage of common pesticides and methods like soil solarisation are usually ineffective in the treatment of this disease. Therefore, there is an urgent need to identify virulence related genes in the pathogen which can be targeted for fungicide development. RESULTS: Pathogenicity testing and phylogenetic classification of the pathogen used in this study confirmed it as Fusarium oxysporum f. sp. lycopersici (Fol) strain. A recent discovery indicates that EF1α, a protein with conserved structural similarity across several fungal genera, has a role in the pathogenicity of Magnaporthe oryzae, the rice blast fungus. Therefore, in this study we have done structural and functional classification of EF1α to understand its role in pathogenicity of Fol. The protein model of Fol EF1α was created using the template crystal structure of the yeast elongation factor complex EEF1A:EEF1BA which showed maximum similarity with the target protein. Using the STRING online database, the interactive information among the hub genes of EF1α was identified and the protein-protein interaction network was recognized using the Cytoscape software. On combining the results of functional analysis, MCODE, CytoNCA and CytoHubba 4 hub genes including Fol EF1α were selected for further investigation. The three interactors of Fol EF1α showed maximum similarity with homologous proteins found in Neurospora crassa complexed with the known fungicide, cycloheximide. Through the sequence similarity and PDB database analysis, homologs of Fol EF1α were found: EEF1A:EEF1BA in complex with GDPNP in yeast and EF1α in complex with GDP in Sulfolobus solfataricus. The STITCH database analysis suggested that EF1α and its other interacting partners interact with guanosine diphosphate (GDPNP) and guanosine triphosphate (GTP). CONCLUSIONS: Our study offers a framework for recognition of several hub genes network in Fusarium wilt that can be used as novel targets for fungicide development. The involvement of EF1α in nucleocytoplasmic transport pathway suggests that it plays role in GTP binding and thus apart from its use as a biomarker, it may be further exploited as an effective target for fungicide development. Since, the three other proteins that were found to be tightly associated Fol EF1α have shown maximum similarity with homologous proteins of Neurospora crassa that form complex with fungicide- Cycloheximide. Therefore, we suggest that cycloheximide can also be used against Fusarium wilt disease in tomato. The active site cavity of Fol EF1α can also be determined for computational screening of fungicides using the homologous proteins observed in yeast and Sulfolobus solfataricus. On this basis, we also suggest that the other closely associated genes that have been identified through STITCH analysis, they can also be targeted for fungicide development.

Oxaloacetate anaplerosis differently contributes to pathogenicity in plant pathogenic fungi Fusarium graminearum and F. oxysporum.

Anaplerosis refers to enzymatic reactions or pathways replenishing metabolic intermediates in the tricarboxylic acid (TCA) cycle. Pyruvate carboxylase (PYC) plays an important anaplerotic role by catalyzing pyruvate carboxylation, forming oxaloacetate. Although PYC orthologs are well conserved in prokaryotes and eukaryotes, their pathobiological functions in filamentous pathogenic fungi have yet to be fully understood. Here, we delve into the molecular functions of the ortholog gene PYC1 in Fusarium graminearum and F. oxysporum, prominent fungal plant pathogens with distinct pathosystems, demonstrating variations in carbon metabolism for pathogenesis. Surprisingly, the PYC1 deletion mutant of F. oxysporum exhibited pleiotropic defects in hyphal growth, conidiation, and virulence, unlike F. graminearum, where PYC1 deletion did not significantly impact virulence. To further explore the species-specific effects of PYC1 deletion on pathogenicity, we conducted comprehensive metabolic profiling. Despite shared metabolic changes, distinct reprogramming in central carbon and nitrogen metabolism was identified. Specifically, alpha-ketoglutarate, a key link between the TCA cycle and amino acid metabolism, showed significant down-regulation exclusively in the PYC1 deletion mutant of F. oxysporum. The metabolic response associated with pathogenicity was notably characterized by S-methyl-5-thioadenosine and S-adenosyl-L-methionine. This research sheds light on how PYC1-mediated anaplerosis affects fungal metabolism and reveals species-specific variations, exemplified in F. graminearum and F. oxysporum.

Trichoderma based formulations control the wilt disease of chickpea (Cicer arietinum L.) caused by Fusarium oxysporum f. sp. ciceris, better when inoculated as consortia: findings from pot experiments under field conditions.

Background: Commercial/chemical pesticides are available to control Fusarium wilt of chickpea, but these antifungals have numerous environmental and human health hazards. Amongst various organic alternatives, use of antagonistic fungi like Trichoderma, is the most promising option. Although, Trichoderma spp. are known to control Fusarium wilt in chickpea but there are no reports that indicate the biocontrol efficacy of indigenous Trichoderma spp. against the local pathogen, in relation to environmental conditions. Methods: In the present study, biological control activity of Trichoderma species formulations viz., Trichoderma asperellum, Trichoderma harzianum (strain 1), and Trichoderma harzianum (strain 2), either singly or in the form of consortia, was investigated against Fusarium oxysporum f. sp. ciceris, the cause of Fusarium wilt in chickpea, in multiyear pot trials under open field conditions. The antagonistic effect of Trichoderma spp. was first evaluated in in vitro dual culture experiments. Then the effects of Trichoderma as well as F. oxysporum, were investigated on the morphological parameters, disease incidence (DI), and disease severity (DS) of chickpea plants grown in pots. Results: In dual culture experiments, all the Trichoderma species effectively reduced the mycelial growth of F. oxysporum. T. asperellum, T. harzianum (strain 1), and T. harzianum(strain 2) declined the mycelial growth of F. oxysporumby 37.6%, 40%, and 42%. In open field pot trials, the infestation of F. oxysporum in chickpea plants significantly reduced the morphological growth of chickpea. However, the application of T. asperellum, T. harzianum (strain 1), and T. harzianum (strain 2), either singly or in the form of consortia, significantly overcome the deleterious effects of the pathogen, thereby resulted in lower DI (22.2% and 11.1%) and DS (86% and 92%), and ultimately improved the shoot length, shoot fresh weight and shoot dry weight by 69% and 72%, 67% and 73%, 68% and 75%, during the years 1 and 2, respectively, in comparison with infested control. The present study concludes the usefulness and efficacy of Trichoderma species in controlling wilt disease of chickpea plants under variable weather conditions.

Modulation of Growth and Mycotoxigenic Potential of Pineapple Fruitlet Core Rot Pathogens during In Vitro Interactions.

Pineapple Fruitlet Core Rot (FCR) is a fungal disease characterized by a multi-pathogen pathosystem. Recently, Fusarium proliferatum, Fusarium oxysporum, and Talaromyces stollii joined the set of FCR pathogens until then exclusively attributed to Fusarium ananatum. The particularity of FCR relies on the presence of healthy and diseased fruitlets within the same infructescence. The mycobiomes associated with these two types of tissues suggested that disease occurrence might be triggered by or linked to an ecological chemical communication-promoting pathogen(s) development within the fungal community. Interactions between the four recently identified pathogens were deciphered by in vitro pairwise co-culture bioassays. Both fungal growth and mycotoxin production patterns were monitored for 10 days. Results evidenced that Talaromyces stollii was the main fungal antagonist of Fusarium species, reducing by 22% the growth of Fusarium proliferatum. A collapse of beauvericin content was observed when FCR pathogens were cross-challenged while fumonisin concentrations were increased by up to 7-fold. Antagonism between Fusarium species and Talaromyces stollii was supported by the diffusion of a red pigmentation and droplets of red exudate at the mycelium surface. This study revealed that secondary metabolites could shape the fungal pathogenic community of a pineapple fruitlet and contribute to virulence promoting FCR establishment.

FolIws1-driven nuclear translocation of deacetylated FolTFIIS ensures conidiation of Fusarium oxysporum.

Plant diseases caused by fungal pathogens pose a great threat to crop production. Conidiation of fungi is critical for disease epidemics and serves as a promising drug target. Here, we show that deacetylation of the FolTFIIS transcription elongation factor is indispensable for Fusarium oxysporum f. sp. lycopersici (Fol) conidiation. Upon microconidiation, Fol decreases K76 acetylation of FolTFIIS by altering the level of controlling enzymes, allowing for its nuclear translocation by FolIws1. Increased nuclear FolTFIIS enhances the transcription of sporulation-related genes and, consequently, enables microconidia production. Deacetylation of FolTFIIS is also critical for the production of macroconidia and chlamydospores, and its homolog has similar functions in Botrytis cinerea. We identify two FolIws1-targeting chemicals that block the conidiation of Fol and have effective activity against a wide range of pathogenic fungi without harm to the hosts. These findings reveal a conserved mechanism of conidiation regulation and provide candidate agrochemicals for disease management.

Genome-wide search and gene expression studies reveal candidate effectors with a role in pathogenicity and virulence in Fusarium graminearum.

Fusarium graminearum causes Fusarium head blight (FHB) disease in wheat worldwide. Although F. graminearum is reported to secrete several effectors, their role in virulence and pathogenicity is unknown. The study aimed at identifying candidate genes with a role in pathogenicity and virulence using two different host systems, Arabidopsis thaliana and wheat, challenged with F. graminearum TN01. Detached leaf assay and histological studies revealed the virulent nature of TN01. A genome-wide in silico search revealed several candidate genes, of which 23 genes were selected based on reproducibility. Gene expression studies by reverse transcriptase-polymerase chain reaction (RT-PCR) in leaf tissues of Arabidopsis and the two wheat genotypes, the susceptible (Sonalika) and the resistant (Nobeoka Bozu/Nobeoka), compared with mock-treated controls in a time-course study using fungal- and plant-specific genes as internal controls revealed that these genes were differentially regulated. Further, expression of these candidates in F. graminearum-inoculated Sonalika and Nobeoka spikes compared with mock-treated controls revealed their role in pathogenicity and virulence. Gene ontology studies revealed that some of these secretory proteins possessed a role in apoptosis and ceratoplatanin and KP4 killer toxin syntheses. A three-dimensional protein configuration was performed by homology modeling using trRosetta. Further, real-time quantitative PCR (RT-qPCR) studies in F. graminearum-inoculated Arabidopsis and wheat at early time points of inoculation revealed an increased expression of the majority of these genes in Sonalika, suggesting their possible role in pathogenicity, whereas low mRNA abundance was observed for 11 of these genes in the resistant genotype, Nobeoka, compared with Sonalika, indicating their role in virulence of F. graminearum.

Virulence of banana wilt-causing fungal pathogen Fusarium oxysporum tropical race 4 is mediated by nitric oxide biosynthesis and accessory genes.

Fusarium wilt of banana, caused by Fusarium oxysporum f. sp. cubense (Foc), is one of the most damaging plant diseases known. Foc race 1 (R1) decimated the Gros Michel-based banana (Musa acuminata) trade, and now Foc tropical race 4 (TR4) threatens global production of its replacement, the Cavendish banana. Here population genomics revealed that all Cavendish banana-infecting Foc race 4 strains share an evolutionary origin distinct from that of R1 strains. Although TR4 lacks accessory chromosomes, it contains accessory genes at the ends of some core chromosomes that are enriched for virulence and mitochondria-related functions. Meta-transcriptomics revealed the unique induction of the entire mitochondrion-localized nitric oxide (NO) biosynthesis pathway upon TR4 infection. Empirically, we confirmed the unique induction of a NO burst in TR4, suggesting that nitrosative pressure may contribute to virulence. Targeted mutagenesis demonstrated the functional importance of fungal NO production and the accessory gene SIX4 as virulence factors.

Enhancing tomato plant immune responses to Fusarium wilt disease by red seaweed Jania sp.

The novelty of this study lies in demonstrating a new approach to control wilt diseases using Jania ethyl acetate extract. In the current investigation, the potential impacts of Jania sp. ethyl acetate extract (JE) on Tomato Fusarium oxysporum wilt (FOW) have been studied. The in vitro antifungal potential of JE against F. oxysporum (FO) was examined. GC-MS investigation of the JE revealed that, the compounds possessing fungicidal action were Phenol,2-methoxy-4-(2-propenyl)-,acetate, Eugenol, Caryophyllene oxide, Isoespintanol, Cadinene, Caryophylla-4(12),8(13)-dien-5à-ol and Copaen. Jania sp. ethyl acetate extract exhibited strong antifungal potential against FO, achieving a 20 mmzone of inhibition. In the experiment, two different methods were applied: soil irrigation (SI) and foliar application (FS) of JE. The results showed that both treatments reduced disease index present DIP by 20.83% and 33.33% respectively. The findings indicated that during FOW, proline, phenolics, and the antioxidant enzymes activity increased, while growth and photosynthetic pigments decreased. The morphological features, photosynthetic pigments, total phenol content, and antioxidant enzyme activity of infected plants improved when JE was applied through soil or foliar methods. It is interesting to note that the application of JE had a substantially less negative effect on the isozymes peroxidase and polyphenol oxidase in tomato plants, compared to FOW. These reactions differed depending on whether JE was applied foliarly or via the soil. Finally, the use of Jania sp. could be utilized commercially as an ecologically acceptable method to protect tomato plants against FOW.

Fungal Diversity and Gibberellin Hormones Associated with Long Whips of Smut-Infected Sugarcanes.

Sugarcane smut, caused by the fungus Sporisorium scitamineum (Sydow), significantly affects sugarcane crops worldwide. Infected plants develop whip-like structures known as sori. Significant variations in these whip lengths are commonly observed, but the physiological and molecular differences causing these morphological differences remain poorly documented. To address this, we employed conventional microbe isolation, metagenomic, and metabolomic techniques to investigate smut-infected sugarcane stems and whips of varying lengths. Metagenomics analysis revealed a diverse fungal community in the sugarcane whips, with Sporisorium and Fusarium genera notably present (>1%) in long whips. Isolation techniques confirmed these findings. Ultra-performance liquid chromatography analysis (UHPLC-MS/MS) showed high levels of gibberellin hormones (GA3, GA1, GA4, GA8, and GA7) in long whips, with GA4 and GA7 found exclusively in long whips and stems. Among the prominent genera present within long whips, Fusarium was solely positively correlated with these gibberellin (GA) hormones, with the exception of GA8, which was positively correlated with Sporisorium. KEGG enrichment analysis linked these hormones to pathways like diterpenoid biosynthesis and plant hormone signal transduction. These findings suggest that Fusarium may influence GA production leading to whip elongation. Our study reveals fungal dynamics and gibberellin responses in sugarcane smut whips. Future research will explore the related molecular gibberellin synthesis mechanisms.

Genome-Wide Identification of a Maize Chitinase Gene Family and the Induction of Its Expression by Fusarium verticillioides (Sacc.) Nirenberg (1976) Infection.

Maize chitinases are involved in chitin hydrolysis. Chitinases are distributed across various organisms including animals, plants, and fungi and are grouped into different glycosyl hydrolase families and classes, depending on protein structure. However, many chitinase functions and their interactions with other plant proteins remain unknown. The economic importance of maize (Zea mays L.) makes it relevant for studying the function of plant chitinases and their biological roles. This work aims to identify chitinase genes in the maize genome to study their gene structure, family/class classification, cis-related elements, and gene expression under biotic stress, such as Fusarium verticillioides infection. Thirty-nine chitinase genes were identified and found to be distributed in three glycosyl hydrolase (GH) families (18, 19 and 20). Likewise, the conserved domains and motifs were identified in each GH family member. The identified cis-regulatory elements are involved in plant development, hormone response, defense, and abiotic stress response. Chitinase protein-interaction network analysis predicted that they interact mainly with cell wall proteins. qRT-PCR analysis confirmed in silico data showing that ten different maize chitinase genes are induced in the presence of F. verticillioides, and that they could have several roles in pathogen infection depending on chitinase structure and cell wall localization.

Metabolomics and histopathological analysis of two tomato cultivars after co-infection with soil-borne pathogens (Southern root-knot nematode and Fusarium wilt fungus).

Southern root-knot nematode (Meloidogyne incognita) and Fusarium wilt fungus (Fusarium oxysporum) are one of the most predominant pathogens responsible for substantial agricultural yield reduction of tomato. The current study planned to assess the effects of M. incognita (Mi) and F. oxysporum (Fo) and their co-infection on two tomato cultivars, Zhongza 09 (ZZ09) and Gailing Maofen 802 (GLM802). The present study examined the effects of co-infection on leaf morphology, chlorophyll content, leaf area, and histopathology. The present study used metabolomics to evaluate plant-pathogen interactions. The outcomes of the current study revealed that chlorophyll content and leaf area decreased more in GLM802 during co-infection. In co-infection (Fo + Mi), the chlorophyll content reduction in ZZ09 was 11%, while in GLM802 the reduction reached up to 31% as compared to control. Moreover, the reduction in leaf are in ZZ09 was 31%, however, in the GLM802 reduction was observed 54% as compared to control plants. Similarly, GLM802 stems exhibited larger brown patches on their vascular bundles than ZZ09 stems. The rate of browning of GLM802 stems was 247% more than ZZ09, during co-infection. Moreover, GLM802 roots exhibited a higher abundance of hyphae and larger galls than ZZ09 roots. In metabolic studies, glutathione, succinic acid, and 2-isopropylmalic acid decreased, whereas spermine and fumaric acid increased in GLM802 co-infected stems. It indicates that GLM802 is weakly resistant; therefore, F. oxysporum and other pathogens readily damage tissue. In the co-infected stem of ZZ09, L-asparagine and shikimic acid increased, but pipecolic acid, L-saccharine, and 2-isopropylmalic acid declined. L-asparagine was crucial in preserving the stability of nitrogen metabolism, chlorophyll synthesis, and leaf growth in ZZ09. Shikimic acid's substantial accumulation could explain the limited extent of browning observed in the vascular bundles of ZZ09. Thus, the present study provides insight into M. incognita and F. oxysporum co-infection in two tomato cultivars, which may aid breeding efforts to generate commercially viable resistant cultivars. However, further research on the relationship between M. incognita and F. oxysporum in different host plants is required in the future.

Protocols to enable fluorescence microscopy of microbial interactions on living maize silks (style tissue).

There is interest in studying microbes that colonize maize silks (style tissue, critical for reproduction) including the fungal pathogen Fusarium graminearum (Fg) and its interactions with the microbiome and biocontrol agents. In planta imaging of these interactions on living silks using confocal fluorescence microscopy would provide key insights. However, newly discovered microbes have unknown effects on human health, and there are regulatory requirements to prevent the release of fluorescently tagged microbes into the environment. Therefore, the microbe infection, colonization, and interaction stages on silks prior to microscopy must be contained. At the same time, silk viability must be maintained and experiments conducted that are biologically relevant (e.g. silks should remain attached to the cob), yet the silk tissue must be accessible to the researcher (i.e. not within husk leaves) and allow for multiple replicates. Here we present methods that meet these five contrasting criteria. We tested these methods using Fg and four silk-derived bacterial endophytes. The endophytes were previously known to have anti-Fg activity in vitro, but in planta observations were lacking. In Method 1, a portion of the tip of a cob was dissected, and silks remained attached to the cob in a Petri dish. The cob was placed on a water agar disc to maintain hydration. DsRed-tagged bacteria and GFP-tagged Fg were inoculated onto the silks and incubated, allowing the two microbes to grow towards one another before staining with propidium iodide for confocal microscopy. A variation of the protocol was presented in Method 2, where detached silk segments were placed directly on water agar where they were inoculated with bacteria and Fg to promote dense colonization, and to allow for many replicates and interventions such as silk wounding. The bacterial endophytes were successfully observed colonizing Fg hyphae, silk trichomes, and entering silks via cut ends and wounds. These protocols can be used to study other silk-associated microbes including several globally important fungal pathogens that enter maize grain through silks.

A ribonuclease T2 protein FocRnt2 contributes to the virulence of Fusarium oxysporum f. sp. cubense tropical race 4.

Banana Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4), is a major disease of banana plants worldwide. Effector proteins play critical roles in banana-Foc TR4 interaction. Our previous studies highlighted a ribonuclease protein belonging to the T2 family (named as FocRnt2) in the Foc TR4 secretome, which was predicted to be an effector. However, its biological function in Foc TR4 infection is still unclear. Herein, we observed significant expression of FocRnt2 during the early stage of fungal infection in planta. A yeast signal sequence trap assay showed that FocRnt2 contained a functional signal peptide for secretion. FocRnt2 possessed ribonuclease activity that could degrade the banana total RNA in vitro. Subcellular localization showed that FocRnt2 was localized in the nucleus and cytoplasm of Nicotiana benthamiana leaves. Transient expression of FocRnt2 suppressed the expression of salicylic acid- and jasmonic acid-signalling marker genes, reactive oxygen species accumulation, and BAX-mediated cell death in N. benthamiana. FocRnt2 deletion limited fungal penetration, reduced fusaric acid biosynthesis in Foc TR4, and attenuated fungal virulence against banana plants, but had little effect on Foc TR4 growth and sensitivity to various stresses. Furthermore, FocRnt2 deletion mutants induced higher expression of the defence-related genes in banana plants. These results suggest that FocRnt2 plays an important role in full virulence of Foc TR4, further improving our understanding of effector-mediated Foc TR4 pathogenesis.

Fusarium andiyazi, a Pathogenic Species Associated with Rice Bakanae Disease in Malaysia.

Rice is the main staple food crops for the Malaysian population. Rice is also susceptible to bakanae diseases caused by some Fusarium species and reducing yield, and quality of rice also profit. In this study, several rice fields were surveyed to collect Fusarium isolates associated with bakanae disease. The morphological features of Fusarium andiyazi isolates found on infected rice plants were identified in this investigation. For biological species identification, MAT-1 (Mating type idiomorphs) bearing isolates were crossed with MAT-2 isolates. Crossing was succeeded between cross of two different mating type bearing field isolates. Consequently, there is a possibility of exchange of genetic material within the F. andiyazi population in Malaysia. The identity of the isolates was further determined up to the species level by comparing DNA sequences and phylogenetic analysis of two genes. The phylogenetic analyses of the joined dataset of translation elongation factor 1-alpha (TEF1-α) and RNA polymerase subunit II (RPB2) revealed that all the isolates were F. andiyazi. In pathogenicity tests, F. andiyazi were found to be pathogenic on the susceptible rice cultivars MR211 and MR220. Inoculated rice seedling produced typical bakanae symptom like elongation, thin and yellow leaves. F. andiyazi was further confirmed as pathogenic species by Ultra High-Performance Liquid Chromatography (UPLC) detection of Gibberellic acid (GA3) and Fusaric acid. In this study, F. andiyazi strains have been identified as the responsible pathogen for causing rice bakanae disease in Malaysia and it is the first report of F. andiyazi, as a pathogenic species on rice in Malaysia.

Chitinase Gene FoChi20 in Fusarium oxysporum Reduces Its Pathogenicity and Improves Disease Resistance in Cotton.

Chitinase genes, as a class of cell wall hydrolases, are essential for the development and pathogenesis of Fusarium oxysporum f.sp. vasinfectum (F. ox) in cotton, but related research focused on chitinase genes are limited. This study explored two island cotton root secretions from the highly resistant cultivar Xinhai 41 and sensitive cultivar Xinhai 14 to investigate their interaction with F. ox by a weighted correlation network analysis (WGCNA). As a result, two modules that related to the fungal pathogenicity emerged. Additionally, a total of twenty-five chitinase genes were identified. Finally, host-induced gene silencing (HIGS) of FoChi20 was conducted, and the cotton plants showed noticeably milder disease with a significantly lower disease index than the control. This study illuminated that chitinase genes play crucial roles in the pathogenicity of cotton wilt fungi, and the FoChi20 gene could participate in the pathogenesis of F. ox and host-pathogen interactions, which establishes a theoretical framework for disease control in Sea Island cotton.

Biostimulant and antagonistic potential of endophytic fungi against fusarium wilt pathogen of tomato Fusarium oxysporum f. sp. lycopersici.

Endophytic fungal-based biopesticides are sustainable and ecologically-friendly biocontrol agents of several pests and diseases. However, their potential in managing tomato fusarium wilt disease (FWD) remains unexploited. This study therefore evaluated effectiveness of nine fungal isolates against tomato fusarium wilt pathogen, Fusarium oxysporum f. sp. lycopersici (FOL) in vitro using dual culture and co-culture assays. The efficacy of three potent endophytes that inhibited the pathogen in vitro was assessed against FWD incidence, severity, and ability to enhance growth and yield of tomatoes in planta. The ability of endophytically-colonized tomato (Solanum lycopersicum L.) plants to systemically defend themselves upon exposure to FOL were also assessed through defence genes expression using qPCR. In vitro assays showed that endophytes inhibited and suppressed FOL mycelial growth better than entomopathogenic fungi (EPF). Endophytes Trichoderma asperellum M2RT4, Hypocrea lixii F3ST1, Trichoderma harzianum KF2R41, and Trichoderma atroviride ICIPE 710 had the highest (68.84-99.61%) suppression and FOL radial growth inhibition rates compared to EPF which exhibited lowest (27.05-40.63%) inhibition rates. Endophytes T. asperellum M2RT4, H. lixii F3ST1 and T. harzianum KF2R41 colonized all tomato plant parts. During the in planta experiment, endophytically-colonized and FOL-infected tomato plants showed significant reduction of FWD incidence and severity compared to non-inoculated plants. In addition, these endophytes contributed to improved growth promotion parameters and yield. Moreover, there was significantly higher expression of tomato defence genes in T. asperellum M2RT4 colonized than in un-inoculated tomato plants. These findings demonstrated that H. lixii F3ST1 and T. asperellum M2RT4 are effective biocontrol agents against FWD and could sustainably mitigate tomato yield losses associated with fusarium wilt.

Zinc finger transcription factor ZFP1 is associated with growth, conidiation, osmoregulation, and virulence in the Polygonatum kingianum pathogen Fusarium oxysporum.

Rhizome rot is a destructive soil-borne disease of Polygonatum kingianum and adversely affects the yield and sustenance of the plant. Understanding how the causal fungus Fusarium oxysporum infects P. kingianum may suggest effective control measures against rhizome rot. In germinating conidia of infectious F. oxysporum, expression of the zinc finger transcription factor gene Zfp1, consisting of two C2H2 motifs, was up-regulated. To characterize the critical role of ZFP1, we generated independent deletion mutants (zfp1) and complemented one mutant with a transgenic copy of ZFP1 (zfp1 tZFP1). Mycelial growth and conidial production of zfp1 were slower than those of wild type (ZFP1) and zfp1 tZFP1. Additionally, a reduced inhibition of growth suggested zfp1 was less sensitive to conditions promoting cell wall and osmotic stresses than ZFP1 and zfp1 tZFP1. Furthermore pathogenicity tests suggested a critical role for growth of zfp1 in infected leaves and rhizomes of P. kingianum. Thus ZFP1 is important for mycelial growth, conidiation, osmoregulation, and pathogenicity in P. kingianum.