Genetic diversity of the banana Fusarium wilt pathogen in Cuba and across Latin America and the Caribbean.

Fusarium wilt of bananas (FWB) is a severe plant disease that leads to substantial losses in banana production worldwide. It remains a major concern for Cuban banana cultivation. The disease is caused by members of the soil-borne Fusarium oxysporum species complex. However, the genetic diversity among Fusarium species infecting bananas in Cuba has remained largely unexplored. In our comprehensive survey, we examined symptomatic banana plants across all production zones in the country, collecting 170 Fusarium isolates. Leveraging genotyping-by-sequencing and whole-genome comparisons, we investigated the genetic diversity within these isolates and compared it with a global Fusarium panel. Notably, typical FWB symptoms were observed in Bluggoe cooking bananas and Pisang Awak subgroups across 14 provinces. Our phylogenetic analysis revealed that F. purpurascens, F. phialophorum, and F. tardichlamydosporum are responsible for FWB in Cuba, with F. tardichlamydosporum dominating the population. Furthermore, we identified between five and seven distinct genetic clusters, with F. tardichlamydosporum isolates forming at least two subgroups. This finding underscores the high genetic diversity of Fusarium spp. contributing to FWB in the Americas. Our study sheds light on the population genetic structure and diversity of the FWB pathogen in Cuba and the broader Latin American and Caribbean regions.

Genome sequencing and comparative genomics reveal insights into pathogenicity and evolution of Fusarium zanthoxyli, the causal agent of stem canker in prickly ash.

BACKGROUND: Fusarium zanthoxyli is a destructive pathogen causing stem canker in prickly ash, an ecologically and economically important forest tree. However, the genome lack of F. zanthoxyli has hindered research on its interaction with prickly ash and the development of precise control strategies for stem canker. RESULTS: In this study, we sequenced and annotated a relatively high-quality genome of F. zanthoxyli with a size of 43.39 Mb, encoding 11,316 putative genes. Pathogenicity-related factors are predicted, comprising 495 CAZymes, 217 effectors, 156 CYP450s, and 202 enzymes associated with secondary metabolism. Besides, a comparative genomics analysis revealed Fusarium and Colletotrichum diverged from a shared ancestor approximately 141.1 ~ 88.4 million years ago (MYA). Additionally, a phylogenomic investigation of 12 different phytopathogens within Fusarium indicated that F. zanthoxyli originated approximately 34.6 ~ 26.9 MYA, and events of gene expansion and contraction within them were also unveiled. Finally, utilizing conserved domain prediction, the results revealed that among the 59 unique genes, the most enriched domains were PnbA and ULP1. Among the 783 expanded genes, the most enriched domains were PKc_like kinases and those belonging to the APH_ChoK_Like family. CONCLUSION: This study sheds light on the genetic basis of F. zanthoxyli's pathogenicity and evolution which provides valuable information for future research on its molecular interactions with prickly ash and the development of effective strategies to combat stem canker.

The ING protein Fng2 associated with RPD3 HDAC complex for the regulation of fungal development and pathogenesis in wheat head blight fungus.

ING proteins display a high level of evolutionary conservation across various species, and play a crucial role in modulating histone acetylation levels, thus regulating various important biological processes in yeast and humans. Filamentous fungi possess distinct biological characteristics that differentiate them from yeasts and humans, and the specific roles of ING proteins in filamentous fungi remain largely unexplored. In this study, an ING protein, Fng2, orthologous to the yeast Pho23, has been identified in the wheat head blight fungus Fusarium graminearum. The deletion of the FNG2 gene resulted in defects in vegetative growth, conidiation, sexual reproduction, plant infection, and deoxynivalenol (DON) biosynthesis. Acting as a global regulator, Fng2 exerts negative control over histone H4 acetylation and governs the expression of over 4000 genes. Moreover, almost half of the differentially expressed genes in the fng3 mutant were found to be co-regulated by Fng2, emphasizing the functional association between these two ING proteins. Notably, the fng2 fng3 double mutant exhibits significantly increased H4 acetylation and severe defects in both fungal development and pathogenesis. Furthermore, Fng2 localizes within the nucleus and associates with the FgRpd3 histone deacetylase (HDAC) to modulate gene expression. Overall, Fng2's interaction with FgRpd3, along with its functional association with Fng3, underscores its crucial involvement in governing gene expression, thereby significantly influencing fungal growth, asexual and sexual development, pathogenicity, and secondary metabolism.

FgPfn participates in vegetative growth, sexual reproduction, pathogenicity, and fungicides sensitivity via affecting both microtubules and actin in the filamentous fungus Fusarium graminearum.

Fusarium head blight (FHB), caused by Fusarium graminearum species complexes (FGSG), is an epidemic disease in wheat and poses a serious threat to wheat production and security worldwide. Profilins are a class of actin-binding proteins that participate in actin depolymerization. However, the roles of profilins in plant fungal pathogens remain largely unexplored. Here, we identified FgPfn, a homolog to profilins in F. graminearum, and the deletion of FgPfn resulted in severe defects in mycelial growth, conidia production, and pathogenicity, accompanied by marked disruptions in toxisomes formation and deoxynivalenol (DON) transport, while sexual development was aborted. Additionally, FgPfn interacted with Fgα1 and Fgß2, the significant components of microtubules. The organization of microtubules in the ΔFgPfn was strongly inhibited under the treatment of 0.4 µg/mL carbendazim, a well-known group of tubulin interferers, resulting in increased sensitivity to carbendazim. Moreover, FgPfn interacted with both myosin-5 (FgMyo5) and actin (FgAct), the targets of the fungicide phenamacril, and these interactions were reduced after phenamacril treatment. The deletion of FgPfn disrupted the normal organization of FgMyo5 and FgAct cytoskeleton, weakened the interaction between FgMyo5 and FgAct, and resulting in increased sensitivity to phenamacril. The core region of the interaction between FgPfn and FgAct was investigated, revealing that the integrity of both proteins was necessary for their interaction. Furthermore, mutations in R72, R77, R86, G91, I101, A112, G113, and D124 caused the non-interaction between FgPfn and FgAct. The R86K, I101E, and D124E mutants in FgPfn resulted in severe defects in actin organization, development, and pathogenicity. Taken together, this study revealed the role of FgPfn-dependent cytoskeleton in development, DON production and transport, fungicides sensitivity in F. graminearum.

Unveiling the A-to-I mRNA editing machinery and its regulation and evolution in fungi.

A-to-I mRNA editing in animals is mediated by ADARs, but the mechanism underlying sexual stage-specific A-to-I mRNA editing in fungi remains unknown. Here, we show that the eukaryotic tRNA-specific heterodimeric deaminase FgTad2-FgTad3 is responsible for A-to-I mRNA editing in Fusarium graminearum. This editing capacity relies on the interaction between FgTad3 and a sexual stage-specific protein called Ame1. Although Ame1 orthologs are widely distributed in fungi, the interaction originates in Sordariomycetes. We have identified key residues responsible for the FgTad3-Ame1 interaction. The expression and activity of FgTad2-FgTad3 are regulated through alternative promoters, alternative translation initiation, and post-translational modifications. Our study demonstrates that the FgTad2-FgTad3-Ame1 complex can efficiently edit mRNA in yeasts, bacteria, and human cells, with important implications for the development of base editors in therapy and agriculture. Overall, this study uncovers mechanisms, regulation, and evolution of RNA editing in fungi, highlighting the role of protein-protein interactions in modulating deaminase function.

Development of transgenic Paulownia trees expressing antimicrobial thionin genes for enhanced resistance to fungal infections using chitosan nanoparticles.

There is an increasing focus on genetically altering Paulownia trees to enhance their resistance against fungal infections, given their rapid growth and quality wood production. The aim of this research was to establish a technique for incorporating two antimicrobial thionin genes, namely thionin-60 (thio-60) and thionin-63 (thio-63), into Paulownia tomentosa and Paulownia hybrid 9501 through the utilization of chitosan nanoparticles. The outcomes revealed the successful gene transfer into Paulownia trees utilizing chitosan nanoparticles. The effectiveness of thionin proteins against plant pathogens Fusarium and Aspergillus was examined, with a specific focus on Fusarium equiseti due to limited available data. In non-transgenic Paulownia species, the leaf weight inhibition percentage varied from 25 to 36 %, whereas in transgenic species, it ranged from 22 to 7 %. In general, Paulownia species expressing thio-60 displayed increased resistance to F. equiseti, while those expressing thio-63 exhibited heightened resistance to A. niger infection. The thionin proteins displayed a strong affinity for the phospholipid bilayer of the fungal cell membrane, demonstrating their capability to disrupt its structure. The transgenic plants created through this technique showed increased resistance to fungal infections. Thionin-60 demonstrated superior antifungal properties in comparison to thio-63, being more effective at disturbing the fungal cell membrane. These findings indicate that thio-60 holds potential as a novel antifungal agent and presents a promising approach for enhancing the antimicrobial traits of genetically modified Paulownia trees.

Quantitative Analysis of Fungal Contamination of Different Herbal Medicines in China.

Herbal medicines are widely used for clinical purposes worldwide. These herbs are susceptible to phytopathogenic fungal invasion during the culturing, harvesting, storage, and processing stages. The threat of fungal and mycotoxin contamination requires the evaluation of the health risks associated with these herbal medicines. In this study, we collected 138 samples of 23 commonly used herbs from 20 regions in China, from which we isolated a total of 200 phytopathogenic fungi. Through morphological observation and ITS sequencing, 173 fungal isolates were identified and classified into 24 genera, of which the predominant genera were Fusarium (27.74%) and Alternaria (20.81%), followed by Epicoccum (11.56%), Nigrospora (7.51%), and Trichocladium (6.84%). Quantitative analysis of the abundance of both Fusarium and Alternaria in herbal medicines via RT-qPCR revealed that the most abundant fungi were found on the herb Taraxacum mongolicum, reaching 300,000 copies/µL for Fusarium and 700 copies/µL for Alternaria. The in vitro mycotoxin productivities of the isolated Fusarium and Alternaria strains were evaluated by using liquid chromatography-tandem mass spectrometry (LC-MS/MS), and it was found that the Fusarium species mainly produced the acetyl forms of deoxynivalenol, while Alternaria species mainly produced altertoxins. These findings revealed widely distributed fungal contamination in herbal medicines and thus raise concerns for the sake of the quality and safety of herbal medicines.

Exploring class III cellobiose dehydrogenase: sequence analysis and optimized recombinant expression.

BACKGROUND: Cellobiose dehydrogenase (CDH) is an extracellular fungal oxidoreductase with multiple functions in plant biomass degradation. Its primary function as an auxiliary enzyme of lytic polysaccharide monooxygenase (LPMO) facilitates the efficient depolymerization of cellulose, hemicelluloses and other carbohydrate-based polymers. The synergistic action of CDH and LPMO that supports biomass-degrading hydrolases holds significant promise to harness renewable resources for the production of biofuels, chemicals, and modified materials in an environmentally sustainable manner. While previous phylogenetic analyses have identified four distinct classes of CDHs, only class I and II have been biochemically characterized so far. RESULTS: Following a comprehensive database search aimed at identifying CDH sequences belonging to the so far uncharacterized class III for subsequent expression and biochemical characterization, we have curated an extensive compilation of putative CDH amino acid sequences. A sequence similarity network analysis was used to cluster them into the four distinct CDH classes. A total of 1237 sequences encoding putative class III CDHs were extracted from the network and used for phylogenetic analyses. The obtained phylogenetic tree was used to guide the selection of 11 cdhIII genes for recombinant expression in Komagataella phaffii. A small-scale expression screening procedure identified a promising cdhIII gene originating from the plant pathogen Fusarium solani (FsCDH), which was selected for expression optimization by signal peptide shuffling and subsequent production in a 5-L bioreactor. The purified FsCDH exhibits a UV-Vis spectrum and enzymatic activity similar to other characterized CDH classes. CONCLUSION: The successful production and functional characterization of FsCDH proved that class III CDHs are catalytical active enzymes resembling the key properties of class I and class II CDHs. A detailed biochemical characterization based on the established expression and purification strategy can provide new insights into the evolutionary process shaping CDHs and leading to their differentiation into the four distinct classes. The findings have the potential to broaden our understanding of the biocatalytic application of CDH and LPMO for the oxidative depolymerization of polysaccharides.

Co-overexpression of chitinase and ß-1,3-glucanase significantly enhanced the resistance of Iranian wheat cultivars to Fusarium.

Fusarium head blight (FHB) is a devastating fungal disease affecting different cereals, particularly wheat, and poses a serious threat to global wheat production. Chitinases and ß-glucanases are two important proteins involved in lysing fungal cell walls by targeting essential macromolecular components, including chitin and ß-glucan micro fibrils. In our experiment, a transgenic wheat (Triticum aestivum) was generated by introducing chitinase and glucanase genes using Biolistic technique and Recombinant pBI121 plasmid (pBI-ChiGlu (-)). This plasmid contained chitinase and glucanase genes as well as nptII gene as a selectable marker. The expression of chitinase and glucanase was individually controlled by CaMV35S promoter and Nos terminator. Immature embryo explants from five Iranian cultivars (Arta, Moghan, Sisun, Gascogen and A-Line) were excised from seeds and cultured on callus induction medium to generate embryonic calluses. Embryogenic calluses with light cream color and brittle texture were selected and bombarded using gold nanoparticles coated with the recombinant pBI-ChiGlu plasmid. Bombarded calluses initially were transferred to selective callus induction medium, and later, they were transfferd to selective regeneration medium. The selective agent was kanamycin at a concentration of 25 mg/l in both media. Among five studied cultivars, A-Line showed the highest transformation percentage (4.8%), followed by the Sisun, Gascogen and Arta in descending order. PCR and Southern blot analysis confirmed the integration of genes into the genome of wheat cultivars. Furthermore, in an in-vitro assay, the growth of Fusarium graminearum was significantly inhibited by using 200 µg of leaf protein extract from transgenic plants. According to our results, the transgenic plants (T1) showed the resistance against Fusarium when were compared to the non-transgenic plants. All transgenic plants showed normal fertility and no abnormal response was observed in their growth and development.

Crucial Involvement of Heme Biosynthesis in Vegetative Growth, Development, Stress Response, and Fungicide Sensitivity of Fusarium graminearum.

Heme biosynthesis is a highly conserved pathway from bacteria to higher animals. Heme, which serves as a prosthetic group for various enzymes involved in multiple biochemical processes, is essential in almost all species, making heme homeostasis vital for life. However, studies on the biological functions of heme in filamentous fungi are scarce. In this study, we investigated the role of heme in Fusarium graminearum. A mutant lacking the rate-limiting enzymes in heme synthesis, coproporphyrinogen III oxidase (Cpo) or ferrochelatase (Fc), was constructed using a homologous recombination strategy. The results showed that the absence of these enzymes was lethal to F. graminearum, but the growth defect could be rescued by the addition of hemin, so we carried out further studies with the help of hemin. The results demonstrated that heme was required for the activity of FgCyp51, and its absence increased the sensitivity to tebuconazole and led to the upregulation of FgCYP51 in F. graminearum. Additionally, heme plays an indispensable role in the life cycle of F. graminearum, which is essential for vegetative growth, conidiation, external stress response (especially oxidative stress), lipid accumulation, fatty acid ß-oxidation, autophagy, and virulence.

Development of Rapid Detection Methods for Fusarium oysporum f. sp. melonis in Melon Seeds.

Melon (Cucumis melo L.) is a global commercial crop that is sensitive to seed-borne wilt infections caused by Fusarium oxysporum f. sp. melonis (Fom). To address the challenge of detecting Fom contamination, we designed a probe-based real-time PCR method, TDCP2, in combination with rapid or column-based DNA extraction protocols to develop reliable molecular detection methods. Utilizing TDCP2, the detection rate reached 100% for both artificially Fom-inoculated (0.25-25%) and pod-inoculated melon seeds in conjunction with DNA samples from either the rapid or column-based extraction protocol. We performed analyses of precision, recall, and F1 scores, achieving a maximum F1 score of 1 with TDCP2, which highlights the robustness of the method. Additionally, intraday and interday assays were performed, which revealed the high reproducibility and stability of column-based DNA extraction protocols combined with TDCP2. These metrics confirm the reliability of our developed protocols, setting a foundation for future enhancements in seed pathology diagnostics and potentially broadening their applicability across various Fom infection levels. In the future, we hope that these methods will reduce food loss by improving the control and management of melon diseases.

Characterization of Fusarium species causing head blight of highland barley (qingke) in Tibet, China.

Most of the research on the characterization of Fusarium species focused on wheat, barley, rice, and maize in China. However, there has been limited research in highland barley (qingke). Recently, Fusarium head blight (FHB) of qingke was recently observed in Tibet, China, especially around the Brahmaputra River. To gain a better understanding of the pathogens involver, 201 Fusarium isolates were obtained from qingke samples in 2020. Among these isolates, the most abundant species was F. avenaceum (45.3 %), followed by F. equiseti (27.8 %), F. verticillioides (13.9 %), F. acuminatum (9.0 %), F. flocciferum (3.5 %), and F. proliferatum (0.5 %). The distribution of Fusarium species varied along the Brahmaputra River, with F. avenaceum being predominant in the midstream and downstream regions, while F. equiseti was more common in the upstream region. Chemical analyses of all the isolates revealed the production of different mycotoxins by various Fusarium species. It was found that enniatins were produced by F. acuminatum, F. avenaceum, and F. flocciferum, beauvericin (BEA) and fumonisins were produced F. proliferatum and F. verticillioides, and zearalenone (ZEN) and nivalenol (NIV) were produced by F. equiseti. Pathogenicity test showed that F. avenaceum was more aggressive in causing FHB compared to F. acuminatum, F. equiseti, and F. flocciferum. The disease severity, measured by the area under the disease progress curve (AUDPC), was significantly positively (P < 0.01) correlated with the concentration of total toxins produced by each species. Furthermore, all the Fusarium strains which were used for pathogenicity test were susceptible to carbendazim, and the 50 % effective concentration (EC50) ranged from 0.406 µg/mL to 0.673 µg/mL with an average EC50 of 0.551 ± 0.012 µg/mL.

A comparison between the role of enniatins and deoxynivalenol in Fusarium virulence on different tissues of common wheat.

BACKGROUND: Fusarium graminearum and Fusarium avenaceum are two of the most important causal agents of Fusarium head blight (FHB) of wheat. They can produce mycotoxins that accumulate in infected wheat heads, including deoxynivalenol (DON) and enniatins (ENNs), produced by F. graminearum and F. avenaceum, respectively. While the role of DON as a virulence factor in F. graminearum toward wheat is well known, ENNs in F. avenaceum has been poorly explored. Results obtained to-date indicate that ENNs may confer an advantage to F. avenaceum only on particular hosts. RESULTS: In this study, with the use of ENN-producing and ENN non-producing F. avenaceum strains, the role of ENNs on F. avenaceum virulence was investigated on the root, stem base and head of common wheat, and compared with the role of DON, using DON-producing and DON non-producing F. graminearum strains. The DON-producing F. graminearum strain showed a significantly higher ability to cause symptoms and colonise each of the tested tissues than the non-producing strain. On the other hand, the ability to produce ENNs increased initial symptoms of the disease and fungal biomass accumulation, measured by qPCR, only in wheat heads, and not in roots or stem bases. LC-MS/MS analysis was used to confirm the presence of ENNs and DON in the different strains, and results, both in vitro and in wheat heads, were consistent with the genetics of each strain. CONCLUSION: While the key role of DON on F. graminearum virulence towards three different wheat tissues was noticeable, ENNs seemed to have a role only in influencing F. avenaceum virulence on common wheat heads probably due to an initial delay in the appearance of symptoms.

Fusarium keratitis in a Brazilian tropical semi-arid area: Clinical-epidemiological features, molecular identification and antifungal susceptibility.

BACKGROUND: Fungal keratitis is a severe eye infection that can result in blindness and visual impairment, particularly in developing countries. Fusarium spp. are the primary causative agents of this condition. Diagnosis of Fusarium keratitis (FK) is challenging, and delayed treatment can lead to serious complications. However, there is limited epidemiological data on FK, especially in tropical areas. OBJECTIVES: This study aimed to describe the clinical, laboratorial and epidemiological characteristics of FK in a tropical semi-arid region of Brazil. PATIENTS/METHODS: Adult patients with laboratory-confirmed FK diagnosed between October 2019 and March 2022 were evaluated. Fusarium isolates were characterized at molecular level and evaluated regarding antifungal susceptibility. RESULTS: A total of 226 clinical samples from patients suspected of keratitis were evaluated; fungal growth was detected in 50 samples (22.12%); out of which 42 were suggestive of Fusarium spp. (84%). Molecular analysis of a randomly selected set of 27 isolates identified F. solani species complex (n = 14); F. fujikuroi sensu lato (n = 6) and F. dimerum sensu lato (n = 7); a total of 10 haplotypes were identified among the strains. All but one Fusarium strains were inhibited by amphotericin B, natamycin and fluconazole. Most patients were male (71.42%; 30 out of 42), aged from 27 to 73 years old. Trauma was the most important risk factor for FK (40.47%; 17 out of 42). Patients were treated with antifungals, corticoids and antibiotics; keratoplasty and eye enucleation were also performed. CONCLUSIONS: The study provided insights into the characteristics of FK in tropical regions and emphasized the importance of enhanced surveillance and management strategies.

Revealing the diversity of Jojoba-associated fungi using amplicon metagenome approach and assessing the in vitro biocontrol activity of its cultivable community.

Jojoba shrubs are wild plants cultivated in arid and semiarid lands and characterized by tolerance to drought, salinity, and high temperatures. Fungi associated with such plants may be attributed to the tolerance of host plants against biotic stress in addition to the promotion of plant growth. Previous studies showed the importance of jojoba as jojoba oil in the agricultural field; however, no prior study discussed the role of jojoba-associated fungi (JAF) in reflecting plant health and the possibility of using JAF in biocontrol. Here, the culture-independent and culture-dependent approaches were performed to study the diversity of the jojoba-associated fungi. Then, the cultivable fungi were evaluated for in-vitro antagonistic activity and in vitro plant growth promotion assays. The metagenome analysis revealed the existence of four fungal phyla: Ascomycota, Aphelidiomycota, Basidiomycota, and Mortierellomycota. The phylum Ascomycota was the most common and had the highest relative abundance in soil, root, branch, and fruit samples (59.7%, 50.7%, 49.8%, and 52.4%, respectively). Alternaria was the most abundant genus in aboveground tissues: branch (43.7%) and fruit (32.1%), while the genus Discosia had the highest abundance in the underground samples: soil (24%) and root (30.7%). For the culture-dependent method, a total of 14 fungi were isolated, identified, and screened for their chitinolytic and antagonist activity against three phytopathogenic fungi (Fusarium oxysporum, Alternaria alternata and Rhizoctonia solani) as well as their in vitro plant growth promotion (PGP) activity. Based on ITS sequence analysis, the selected potent isolates were identified as Aspergillus stellatusEJ-JFF3, Aspergillus flavus EJ-JFF4, Stilbocrea sp. EJ-JLF1, Fusarium solani EJ-JRF3, and Amesia atrobrunneaEJ-JSF4. The endophyte strain A. flavus EJ-JFF4 exhibited the highest chitinolytic activity (9 Enzyme Index) and antagonistic potential against Fusarium oxysporum, Alternaria alternata, and Rhizoctonia solani phytopathogens with inhibitory percentages of 72, 70, and 80 respectively. Also, A. flavus EJ-JFF4 had significant multiple PGP properties, including siderophore production (69.3%), phosphate solubilization (95.4 µg ml-1). The greatest production of Indol-3-Acetic Acid was belonged to A. atrobrunnea EJ-JSF4 (114.5 µg ml-1). The analysis of FUNGuild revealed the abundance of symbiotrophs over other trophic modes, and the guild of endophytes was commonly assigned in all samples. For the first time, this study uncovered fungal diversity associated with jojoba plants using a culture-independent approach and in-vitro assessed the roles of cultivable fungal strains in promoting plant growth and biocontrol. The present study indicated the significance of jojoba shrubs as a potential source of diverse fungi with high biocontrol and PGP activities.

Secreted in Xylem (SIX) genes in Fusarium oxysporum f.sp. cubense (Foc) unravels the potential biomarkers for early detection of Fusarium wilt disease.

Secreted in Xylem (SIX) are small effector proteins released by Fusarium oxysporum f.sp. cubense (Foc) into the plant's xylem sap disrupting the host's defence responses causing Fusarium wilt disease resulting in a significant decline in banana crop yields and economic losses. Notably, different races of Foc possess unique sets of SIX genes responsible for their virulence, however, these genes remain underutilized, despite their potential as biomarkers for early disease detection. Herein, we identified seven SIX genes i.e. SIX1, SIX2, SIX4, SIX6, SIX8a, SIX9a and SIX13 present in Foc Tropical Race 4 (FocTR4), while only SIX9b in Foc Race 1 (Foc1). Analysis of SIX gene expression in infected banana roots revealed differential patterns during infection providing valuable insights into host-pathogen interactions, virulence level, and early detection time points. Additionally, a comprehensive analysis of virulent Foc1_C2HIR and FocTR4_C1HIR isolates yielded informative genomic insights. Hence, these discoveries contribute to our comprehension of potential disease control targets in these plants, as well as enhancing plant diagnostics and breeding programs.

Investigation of the Inhibitory Effects of Illicium verum Essential Oil Nanoemulsion on Fusarium proliferatum via Combined Transcriptomics and Metabolomics Analysis.

Fusarium proliferatum is the main pathogen that causes Panax notoginseng root rot. The shortcomings of strong volatility and poor water solubility of Illicium verum essential oil (EO) limit its utilization. In this study, we prepared traditional emulsion (BDT) and nanoemulsion (Bneo) of I. verum EO by ultrasonic method with Tween-80 and absolute ethanol as solvents. The chemical components of EO, BDT, and Bneo were identified by gas chromatography-mass spectrometry (GC-MS) and the antifungal activity and mechanism were compared. The results show that Bneo has good stability and its particle size is 34.86 nm. The contents of (-) -anethole and estragole in Bneo were significantly higher than those in BDT. The antifungal activity against F. proliferatum was 5.8-fold higher than BDT. In the presence of I. verum EO, the occurrence of P. notoginseng root rot was significantly reduced. By combining transcriptome and metabolomics analysis, I. verum EO was found to be involved in the mutual transformation of pentose and glucuronic acid, galactose metabolism, streptomycin biosynthesis, carbon metabolism, and other metabolic pathways of F. proliferatum, and it interfered with the normal growth of F. proliferatum to exert antifungal effects. This study provide a theoretical basis for expanding the practical application of Bneo.

Development of a PCR-based assay for specific and sensitive detection of Fusarium buharicum from infected okra plant.

Fusarium wilt, caused by the fungus Fusarium buharicum, is an emerging disease of okra in Japan. The disease was first reported in Japan in 2015, causing significant damage to okra seedlings. Due to the potential threat in okra cultivation, the development of an accurate detection method for F. buharicum is needed for the surveillance and management of the disease. In this study, we designed a primer set and developed conventional and nested PCR assays for the specific detection of F. buharicum in infected okra plants and contaminated soil, respectively. We compared the diversity of the translation elongation factor 1 alpha (EF-1α) gene of F. buharicum with 103 other fungal species/isolates to design a species-specific primer. This primer pair successfully amplified approximately 400 bp of PCR product that was only detected in the F. buharicum isolate, not in the other fungal isolates. The developed nested PCR method was highly sensitive and could detect the fungus from a 0.01 fg DNA sample. The primer successfully detected the pathogen in artificially infected plants and soil by conventional and nested PCR, respectively. This is the first report of the development of the F. buharicum-specific primer set and detection assays, which can be used for the specific and sensitive detection of F. buharicum in field samples and for taking early control measures.

Comparative transcriptome analysis provides insights into the resistance regulation mechanism and inhibitory effect of fungicide phenamacril in Fusarium asiaticum.

Fusarium asiaticum is a destructive phytopathogenic fungus that causes Fusarium head blight of wheat (FHB), leading to serious yield and economic losses to cereal crops worldwide. Our previous studies indicated that target-site mutations (K216R/E, S217P/L, or E420K/G/D) of Type I myosin FaMyo5 conferred high resistance to phenamacril. Here, we first constructed one sensitive strain H1S and three point mutation resistant strains HA, HC and H1R. Then we conducted comparative transcriptome analysis of these F. asiaticum strains after 1 and 10 µg·mL-1 phenamacril treatment. Results indicated that 2135 genes were differentially expressed (DEGs) among the sensitive and resistant strains. The DEGs encoding ammonium transporter MEP1/MEP2, nitrate reductase, copper amine oxidase 1, 4-aminobutyrate aminotransferase, amino-acid permease inda1, succinate-semialdehyde dehydrogenase, 2, 3-dihydroxybenzoic acid decarboxylase, etc., were significantly up-regulated in all the phenamacril-resistant strains. Compared to the control group, a total of 1778 and 2097 DEGs were identified in these strains after 1 and 10 µg·mL-1 phenamacril treatment, respectively. These DEGs involved in 4-aminobutyrate aminotransferase, chitin synthase 1, multiprotein-bridging factor 1, transcriptional regulatory protein pro-1, amino-acid permease inda1, ATP-dependent RNA helicase DED1, acetyl-coenzyme A synthetase, sarcoplasmic/endoplasmic reticulum calcium ATPase 2, etc., showed significantly down-regulated expression in phenamacril-sensitive strain but not in resistant strains after phenamacril treatment. In addition, cyanide hydratase, mating-type protein MAT-1, putative purine nucleoside permease, plasma membrane protein yro2, etc., showed significantly co-down-regulated expression in all the strains after phenamacril treatment. Taken together, This study provides deep insights into the resistance regulation mechanism and the inhibitory effect of fungicide phenamacril and these new annotated proteins or enzymes are worth for the discovery of new fungicide targets.

Influence of Bacillus subtilis strain Z-14 on microbial ecology of cucumber rhizospheric vermiculite infested with fusarium oxysporum f. sp. cucumerinum.

Fusarium oxysporum (FO) is a typical soil-borne pathogenic fungus, and the cucumber wilt disease caused by F. oxysporum f. sp. cucumerinum (FOC) seriously affects crop yield and quality. Vermiculite is increasingly being used as a culture substrate; nevertheless, studies exploring the effectiveness and mechanisms of biocontrol bacteria in this substrate are limited. In this study, vermiculite was used as a culture substrate to investigate the control effect of Bacillus subtilis strain Z-14 on cucumber wilt and the rhizospheric microecology, focusing on colonization ability, soil microbial diversity, and rhizosphere metabolome. Pot experiments showed that Z-14 effectively colonized the cucumber roots, achieving a controlled efficacy of 61.32% for wilt disease. It significantly increased the abundance of Bacillus and the expression of NRPS and PKS genes, while reducing the abundance of FO in the rhizosphere. Microbial diversity sequencing showed that Z-14 reduced the richness and diversity of the rhizosphere bacterial community, increased the richness and diversity of the fungal community, and alleviated the effect of FO on the community structure of the cucumber rhizosphere. The metabolomics analysis revealed that Z-14 affected ABC transporters, amino acid synthesis, and the biosynthesis of plant secondary metabolites. Additionally, Z-14 increased the contents of phenylacetic acid, capsidol, and quinolinic acid, all of which were related to the antagonistic activity in the rhizosphere. Z-14 exhibited a significant control effect on cucumber wilt and influenced the microflora and metabolites in rhizospheric vermiculite, providing a theoretical basis for further understanding the control effect and mechanism of cucumber wilt in different culture substrates.