Fusarium Tricinctum Research Articles

Bioprospecting Endophytic Fungi of Forest Plants for Bioactive Metabolites with Antibacterial, Antifungal, and Antioxidant Potentials.

The growing emergence of multi-drug resistant microbial strains has kept the scientific world searching for novel bioactive compounds with specific chemical characteristics. Accordingly, researchers have started exploring the understudied metabolites from endophytes as a new source of bioactive compounds. In this context, the current study was designed to evaluate the bioactive properties of endophytic fungi from the Mokrzański forest in Wrocław, Poland that have not yet been fully researched. Forty-three endophytic fungi were isolated from twelve distinct plants. Following their cultivation, fungal extracts were separately prepared from biomass and cell-free filtrates, and their antibacterial, antifungal (against human and plant pathogens), and antioxidant properties were examined. Five promising fungi after screening were identified to possess all of these activities. These strains and their respective plant hosts were Trichoderma harzianum BUK-T (Fagus sylvatica), Aspergillus ochraceus ROB-L1 (Robinia pseudoacacia), Chaetomium cochliodes KLON-L1, Fusarium tricinctum KLON-L2 (Acer platanoides), and Penicillium chrysogenum SOS-B2 (Pinus sylvestris). Moreover, gamma irradiation at several doses (Gy) was separately applied to the fungal cultures to study their effects on the recorded activities. Finally, compounds after preparative thin-layer chromatography fractionation of the five fungal strains were identified by GC-MS. These findings suggest that the isolated endophytic fungi could serve as novel sources of bioactive metabolites with antibacterial, antifungal, and antioxidant properties, potentially paving the way for future research and the development of new bioactive compounds.

Molecular identification and morphological characterisation of multiple fungal pathogens of triticale in Northern Kazakhstan

Background and Aim. Тtriticale is classified as an amphidiploids and is the first grain crop created by humans, possessing high yield potential along with favorable biochemical and technological characteristics. For a long time, it was believed that triticale, during selection, inherited disease resistance from wheat and resistance to abiotic factors from rye. However, in recent years, there have been several reports that triticale is to fungal diseases, which reduces the quality of the harvested crop. The aim of our study is to examine and characterize the fungal pathogens of triticale in Northern Kazakhstan, and conduct molecular-genetic identification of the main fungal pathogens.MaterialsandMethods. The study was conducted of triticale from two varieties, Dauren and Rossika. Primary fungal isolation was carried out on agarized nutrient media, with preliminary identification using microscopy. Molecular genetic analysis was performed to determine the species of fungi. Results. During the study, we isolated five major fungal pathogens from different parts of the plant. Data on the percentage of infection by the main fungal pathogens were provided. Three of them are pathogens of alternariosis – Alternaria alternate (more common in grains 39%, in leaves and scales of seeds 19-21%), fusariosis – Fusarium tricinctum (occurrence: in roots 57%, in grains 17%, in leaves 10%) and helminthosporiosis – Bipolaris sorokiniana (occurrence in roots and leaves 0.83%) of grain crops, which can lead to a decrease and loss of yield due to their production of mycotoxins. A cultural and morphological description of the main fungal pathogens of grain crops was provided. Moleculargenetic identification was carried out using the ribosomal marker ITS (internal transcribed spacer).Conclusion. According to result of our research, we characterized and molecular-genetically identified the most common fungi found on different parts of the triticale plant.

Targeted delivery of 5-fluorouracil and shikonin by blended and coated chitosan/pectin nanoparticles for treatment of colon cancer

Herein, 5-fluorouracil and shikonin (extracted from Fusarium tricinctum) were loaded in chitosan/pectin nanoparticle (CS/PEC-NPs), prepared by blending (B-CS/PEC-NPs) and coating (C-CS/PEC-NPs) methods. The nanoparticles characterized by Fourier Transform Infrared (FTIR), X-ray diffraction (XRD), Energy-dispersive X-ray (EDX), Scanning Electron Microscope (SEM) and Differential Light Scattering (DLS). Then, some properties of the nanoparticles such as drug release rate and the nanoparticles cytotoxicity were studied. The FTIR, XRD, EDX, SEM and DLS results showed that the nanoparticles synthesized properly with an almost spherical morphology, an average size of 82–93 nm for B-CS/PEC-NPs, an average diameter of below 100 nm (mostly 66–89 nm) for C-CS/PEC-NPs, and hydrodynamic diameter of 310–817 nm. The drug release results indicated the lower release rate of drugs for B-CS/PEC-NPs relative to C-CS/PEC-NPs at different pHs, high release rate of drugs for the nanoparticles in the simulated large intestinal fluids containing pectinase, and Korsmeyer-Peppas model for release of the drugs. The results showed more cytotoxicity of B-CS/PEC-NPs containing drugs, especially B-CS/PEC-NPs containing both drugs (B-CS/PEC/5-FU/SHK-NPs) after treating with pectinase (IC50 of 18.6 μg/mL). In conclusion, despite the limitation of C-CS/PEC-NPs for simultaneous loading of hydrophilic and hydrophobic drugs, B-CS/PEC-NPs showed suitable potency for loading and targeted delivery of the drugs.

Isolation and genome characterization of Paenibacillus polymyxa 188, a potential biocontrol agent against fungi.

In this work, we aimed to isolate marine bacteria that produce metabolites with antifungal properties. Paenibacillus polymyxa 188 was isolated from a marine sediment sample, and it showed excellent antifungal activity against many fungi pathogenic to plants (Fusarium tricinctum, Pestalotiopsis clavispora, Fusarium oxysporum, F. oxysporum f. sp. Cubense (Foc), Curvularia plantarum, and Talaromyces pinophilus) and to humans (Aspergillus terreus, Penicillium oxalicum, and Microsphaeropsis arundinis). The antifungal compounds produced by P. polymyxa 188 were extracted and analyzed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The complete genome sequence and biosynthetic gene clusters of P. polymyxa 188 were characterized and compared with those of other strains. A total of 238 carbohydrate-active enzymes (CAZymes) were identified in P. polymyxa 188. Two antibiotic gene clusters, fusaricidin and tridecaptin, exist in P. polymyxa 188, which is different from other strains that typically have multiple antibiotic gene clusters. Paenibacilluspolymyxa 188 was identified with numerous biosynthetic gene clusters, and its antifungal ability against pathogenic fungi was verified.

Significance of Gnomoniopsis smithogilvyi as kernel rot of sweet chestnut in Turkey

AbstractChestnut nut rots caused by fungi, especially by Gnomoniopsis smithogilvyi, showed a significant increase not only in Turkey but also in the other chestnut‐producing countries. In this study, the prevalence of chestnut kernel rot was determined by using 800 fruits collected from eight significant chestnut‐growing provinces of Turkey located in the Black Sea, Marmara, and Aegean Regions. Six types of kernel rot symptoms were determined from the necrotic nuts, brown rot being the most common accounting for 62% of the necrotic nuts. Along with G. smithogilvyi, seven additional fungal species Colletotrichum fioriniae, C. godetiae, Cryphonectria parasitica, Diplodina agrifoliae, Fusarium concentricum, Fusarium tricinctum, and Trichothecium roseum were found to be associated with kernel rot. In addition, some other ubiquitous fungi such as Penicillium spp. were detected. All the pathogenic fungi were identified through a combination of morphological traits and phylogenetic analysis involving partial internal transcribed spacer (ITS) sequences, as well as tef1 and tub2 loci sequences. The most frequently isolated fungus was G. smithogilvyi, and it was found at 98, 31, 10, and 1 of the necrotic kernels out of 291 showing brown rot, brown to black rot, cream colour decay, and white rot symptoms, respectively. It was not found from the kernels showing green rot and greenly‐blue rot symptoms. G. smithogilvyi was prevalent in six of eight chestnut‐growing locations in Turkey. Based on the findings, G. smithogilvyi was shown to be the main fungal agent of brown kernel rot of chestnuts, and some of the other fungi obtained from the kernels were shown to contribute kernel rots of chestnuts.

First Report of Fusarium ipomoeae Causing leaf rot on Hosta plantaginea in China.

Hosta plantaginea is an important horticultural plant with ornamental value and is widely cultivated in China. Since April 2022, leaf rot has been observed in the H. plantaginea plants in Wanzhou District, Chongqing City, China (31º14'58"N, 108º53'25"E), the initial symptom is a yellow and brown lesion on the edge of the leaf, in the late stage, brown blighted tissue caused leaves to curl and abscise. Ten typical diseased leaves were collected, the margins of infected tissues were cut into small pieces (5×5 mm) and were sterilized in 75% Ethanol for 30 s, 3% sodium hypochlorite for 3 min, rinsed three times with sterile water, then dried on sterile filter paper and placed to potato dextrose agar (PDA) medium at 25℃for 4 days. Thirteen isolates with morphological characteristics similar to those of Fusarium spp. (Nelson et al. 1983) were recovered. These isolates had white, pink and yellowish mycelia, two isolates produced irregular colonies, and remaining isolates showed round. Two of each type were selected for intensive study (yz2, yz11, yz9 and yz17). The colony of yz2 reached 62 mm in diameter on PDA medium after seven days, macroconidia were elongated sickle-shaped, 3-5 septa, and 12.92 to 21.49 × 3.42 to 5.90 μm in size, microconidia were oval and measured 5.69 to 12.95 × 3.41 to 9.80 μm in size, conidiophores were whorled and branched, yz9 attained 74 mm in diameter after nine days, macroconidia were curved sickle-shaped and apex cell acuminate, 26.9 to 57.2 × 2.4 to 7.1 μm, 3-5 septa. The microconidia were fusiform, 17.8 to 28.8 × 11.2 to 14.5 μm. Conidiophores variable in length. Genomic DNA was extracted from 7-day-old aerial mycelia of four strains (yz2, yz9, yz11 and yz17). The internal transcribed spacer (ITS) region (White et al. 1990), translation elongation factor (EF-1α) (Cao et al. 2014) and partial RNA polymerase second largest subunit (RPB2) (Wang et al. 2019) gene regions were amplified and multilocus phylogenetic analysis was conducted, their sequences were deposited in NCBI Genbank with the following accession numbers: the strains of yz2 and yz11 with OQ829372 and OR236201 for ITS, OQ848594 and OR282462 for EF-1α, OR492296 and OR492297 for RPB2; yz9 and yz17 with OQ829383 and OR236222 for ITS, OQ848595 and OR282463 for EF-1α, OR492295 and OR492298 for RPB2. The ModelFinder was used to select the best-fit model in PhyloSuite v1.2.2, the Bayesian Inference method (BI) analysis was used to estimate the system relationship, yz9 and yz17 were identified as Fusarium ipomoeae, yz2 and yz11 were identified as Fusarium tricinctum. To verify Koch's postulates, 8 healthy plants of H. plantaginea (two-year-old) grown were rinsed with sterile water, after 5 leaves per plant were stabbed with a sterilized needle, 4 plants were inoculated with conidial suspension (1×106 conidia mL-1), other plants injected with sterile water as control, then placed in a greenhouse maintained with 95% relative humidity at 25 ± 1°C. The symptoms on the leaves were similar to field after inoculation for 7 days, whereas all control leaves remained healthy. The same pathogen was re-isolated and re-identified based on multilocus phylogenetic analysis, fulfilling Koch's postulates. To our knowledge, this is the first report of F. ipomoeae causing leaf rot on H. plantaginea in China. In addition, F. ipomoeae was reported to cause leaf spot in Peanut (Xu et al. 2021), and F. tricinctum can cause fruit rot on navel orange in China (Yang et al. 2023). H. plantaginea as a horticultural plant is popular with some people, but it has long been threatened by Fusarium.spp. The finding can provide a theoretical basis for control leaf rot on H. plantaginea.

First report of Fusarium tricinctum Causing Leaf Blight on Amorphophallus konjac in Hunan Province of China.

Konjac (Amorphophallus konjac) is a perennial herbaceous plant of the Araceae family, cultivated mainly in south-western China and used extensively for weight loss (Chua et al. 2010). In June 2022, leaf blight was detected on a 2,00 ha A. konjac plantation in Chenxi County, Huaihua City, Hunan Province. It infected almost 20% of the area under cultivation and tends to occur each year during warm, humid weather from May to July, causing significant economic losses to A. konjac production. There were small brown spots on the leaves which gradually spread to form irregular brown lesions. In severe cases the entire plant turned yellow and died. Nine samples were collected randomly from different plants in three plantation forests to isolate the pathogens. They were washed with sterile water and the lesions were excised. They were subsequently disinfected with 3% hydrogen peroxide for 30 s, 75% ethanol for 90 s and rinsed three times with sterile water. The cut sections were then placed on water agar plates and grown in the dark in a constant temperature incubator at 28℃ for 3-5 days, when mycelia grew they were transferred to potato dextrose agar medium and grown in the dark at 28℃ for 3-5 days. Eleven purified fungal isolates were obtained, ten of which looked like Fusarium (90.9% isolation rate), and three representative isolates (MY5, MY7 and MY9) were chosen for further study. The fungal colonies initially appeared white and gradually turnned dark red. Macroconidia were crescent-shaped, elongated, slightly curved and had 2 to 4 septations, with a predominance of 3 septations. They measured 15.540 to 42.083 × 2.760 to 4.558 μm (n=100). Microconidia were oval or pyriform, with a maximum of one septum and measured 6.135 to 24.990 × 2.158 to 4.412 μm (n=100). Two genetic regions, the translation elongation factor-1 (TEF1-α) and RNA polymerase II largest subunit (RPB1) genes, were amplified and sequenced to verify the identity of the fungus (Qiu et al. 2023). The universal primers TEF1-F/R, G2R/Fa were used for amplification and sequencing, and the sequences were submitted to GenBank (TEF1-α: OR545395, OR545397, OR545399; RPB1: OR545394, OR545396, OR545398). A joint phylogenetic tree of the two genes was constructed and analysis showed that the three isolates were significantly clustered with Fusarium tricinctum. Based on the results of morphological identification and phylogenetic tree analysis, the three isolates were identified as F. tricinctum. Pathogenicity tests were carried out on 12 uniformly growing leaf expansion stages of konjac plants，each inoculated with five young leaves. Mycelial blocks of 6 × 6 mm grown on PDA media for 5 days were placed on the surface of the leaves, while sterile PDA blocks were placed on the control plant. After 10 days of rearing the treated plants in a constant temperature chamber at 28°C and 90% relative humidity, the lesions appeared and the pathogens re-isolated from the diseased tissues had the same morphological characteristics as representative isolates. F. tricinctum has been shown to be the major pathogenic fungus causing leaf blight in wheat (Triticum aestivum L.) (Castañares et al. 2011) and orchardgrass (Dactylis glomerata L.) (Wu et al. 2020). To our knowledge, this is the first time in the world that F. tricinctum has been reported to cause leaf blight in A. konjac. This research could provide a foundation for future control of leaf blight disease.

Morphological and Molecular Identification of Fusarium tricinctum Causing Fruit Rot of Pumpkin (Cucurbita pepo) in Iraq

Thanoon, A.H. 2024. Morphological and Molecular Identification of Fusarium tricinctum Causing Fruit Rot of Pumpkin (Cucurbita pepo) in Iraq. Arab Journal of Plant Protection, 42(2): 168-173. https://doi.org/10.22268/AJPP-001241 In November 2019, samples of pumpkin fruits with post harvest rotting symptoms were collected from farmers' stores. The causative fungus was isolated from infected pumpkin fruits. Based on morphological traits, Koch's hypotheses and molecular diagnostic tests were employed to confirm the infection and identify the causal agent. Molecular diagnosis using the polymerase chain reaction (PCR) confirmed the identity of the causal fungus. The results of electrophoresis using a 2% agarose gel showed the presence of a 550 bp amplified band. The ITS sequences were found to be homologous. to that of Fusarium tricinctum in GenBank database at a similarity level of 99%. GenBank assigned the number MZ166321.1 to the Iraqi isolate. This is the first record of F. tricinctum that causes post-harvest pumpkin rot in Iraq. Keywords: Pumpkin fruit rot, postharvest diseases, Fusarium tricinctum.

First Report of Fusarium tricinctum Causing Root Rot on Chinese dwarf cherry (Cerasus humilis) in China.

Chinese dwarf cherry (Cerasus humilis) is a perennial small shrub indigenous to northern China, highly regarded for its calcium-rich fruit known as 'calcium fruit'. These fruits have a remarkable capacity to aid in human calcium absorption. Additionally, they contain beneficial flavonoids that hold promise for applications in the healthcare industry (Wang et al., 2018). In July 2020, a concerning development occurred on the farms in Jingtai County (37.48o N, 103.82o E), Baiyin City, Gansu Province in China. Approximately 20 to 30% of C. humilis at the full culture stage exhibited symptoms of root rot, including brownish leaves, rotten roots, and plant mortality, leading to a decrease of over 30% in calcium fruit yield. To identify the pathogen, the surface of symptomatic roots was sterilized with 3% NaOCl for 2 minutes, followed by 70% ethanol for 30 seconds, and rinsed three times with sterile water. Tissue sections (5×5mm) from the margin of the necrotic lesion were cut and cultured on potato dextrose agar (PDA) medium, and incubated for 7 days at 25℃. Five pure culture isolates were obtained from individual spores. Initially, the isolates exhibited abundant white aerial mycelia that turned light pink on the third day. Macroconidia were falciform, two to five septate, straight or slightly curved, and measuring 20.1 to 32.5×2.2 to 3.8 μm (n=50). Napiform microconidia were oval-ellipsoid, non-septate, and measuring 6.2 to 9.3×4.2 to 5.8 μm (n=50). Based on these morphological characteristics, the fungus was tentatively identified as Fusarium species (Leslie and Summerell, 2006). To confirm the identification, the internal transcribed spacer region (ITS) and the translation elongation factor (EF1α) of the isolate CH-2 were partially amplified and sequenced using the primers ITS1/ITS4 and EF2T/EF3 (Li et al., 2013, Yang et al., 2022). Upon comparison with the sequences in GenBank, 857 bp ITS sequence showed 100% homology to Fusarium tricinctum isolate QY3-1 (GenBank accessions no. MZ572963.1), and 665 bp EF1α sequence showed 99.7% homology to F. tricinctum strain TQC-C2 (GenBank accessions no. KF939493.1). The resulting sequences were deposited into GenBank with accession nos. OQ581576 and OQ848462 respectively. A maximum likelihood (ML) phylogenetic analysis based on combined partial ITS and EF1α data set was conducted via ML bootstrapping using MEGA 11. According to morphology and phylogenetic analysis, the isolate was identified as F. tricinctum (Wang et al., 2022). For a pathogenicity test, a pot experiment was conducted in a greenhouse with a temperature range of 20-27℃ and 60% relative humidity. Roots of C. humilis were immersed in a spore suspension (1×107 conidia/ml) of isolate F. tricinctum CH-2 for approximately 5 minutes. Subsequently the treated roots were planted in pots filled with sterilized field soil, while roots dipped in sterilized water were used as the control. The experiment considered of ten pots for the inoculation treatment and six pots for the control treatment. All pots were maintained in the greenhouse. After 15 days, it was observed that 80% of the inoculated plants displayed symptoms consistent with the field observations, indicating successful infection. In contrast, plants in the control treatment did not exhibit any symptoms. The same fungal pathogen as F. tricinctum CH-2 was reisolated from the diseased root tissue and confirmed through morphological and molecular assays, thereby satisfying Koch's postulates. This is the first documented report of F. tricinctum causing root rot in C. humilis in China. This disease has the potential to become one of the most significant diseases affecting C. humilis in China.

Endophytic Fungal Diversity in Cirsium kawakamii from Taiwan.

The endophytic fungal diversity of Cirsium kawakamii, a herb indigenous to Taiwan, was analyzed in this study. In addition, some fungal isolates were evaluated for the risk they pose as plant pathogens. In total, 1836 endophytic fungi were isolated from C. kawakamii from Hehuanjian, Puli Township, and Tatachia. They were classified into 2 phyla, 8 classes, 40 families, and 68 genera. Colletotrichum, Fusarium, Phomopsis, and Xylaria, (Ascomycota, Sordariomycetes) were the dominant genera. The genus accumulation curve (based on the bootstrap estimator) was non-asymptotic, with estimated richness significantly exceeding the richness captured by our sampling to date. Considering the collection time, the data indicated significant differences in the proportions of the C. kawakamii endophyte genus from Hehuanjan, Puli Township (across two seasons), and Tatachia. The Shannon and Gini-Simpson indices revealed variations in diversity, with C. kawakamii endophytes (Puli Township in winter) significantly reducing alpha diversity compared with other seasons and locations. Meanwhile, the Gini-Simpson index suggested that there were no significant differences in richness among the four sampling sites. The PCA results unveiled distinct community structures across different locations and seasons, explaining 46.73% of the total variation in fungal community composition significantly affected diversity and richness. In addition, a considerable number of Fusarium isolates exhibited harmful properties towards wheat, potatoes, and apples. It is postulated that these fungi belong to the Fusarium tricinctum species complex (FTSC).

The 2′,4′-Dichloro-chalcone Inhibits the In Vitro Growth and Pathogenicity of Fusarium tricinctum and Trichothecium roseum by Activating Cyanide-Resistant Respiration

Chalcones are a class of flavonoids possessing antimicrobial properties and have potential for use as coatings of plant products for the control of postharvest diseases. The effects of 2′,4′-dichloro-chalcone on the in vitro growth and in vivo pathogenicity of Fusarium tricinctum and Trichothecium roseum were investigated. First, 1 µM of 2′,4′-dichloro-chalcone strongly inhibited the mycelial growth and conidial production of F. tricinctum (32.3%) and T. roseum (65.2%) in vitro. Meanwhile, the cell membrane permeability was increased by 25% and 22.5% and the accumulation of reactive oxygen species was increased by 41.7 and 65.4%, respectively, of F. tricinctum and T. roseum. This treatment also significantly inhibited the total respiration rate and activated the cyanide-resistant respiratory pathway in both pathogens. The expression level of AOX was enhanced in F. tricinctum and T. roseum by 52.76 and 39.13%, respectively. This treatment also significantly inhibited the expansion of potato dry rot from F. tricinctum (48.6%) and apple rot spot from T. roseum (36.2%). Therefore, 2′,4′-dichloro-chalcone has potential use as an alternative safety method in the control of postharvest diseases by F. tricinctum and T. roseum in agricultural practices.

First report of Fusarium tricinctum causing root rot on sugar beet (Beta vulgaris L.) in Morocco

First report of Fusarium tricinctum causing root rot on sugar beet (Beta vulgaris L.) in Morocco

First report of Fusarium tricinctum Species complex causing root rot of tobacco in China.

Tobacco (Nicotiana tabacum L.) is an economically important crop in China. In June 2021, a root rot disease was observed on tobacco (cv. Yunyan99) in Lushi, Mianchi, and Luoning counties of western Henan, China. Diseased tobacco plants exhibited wilting with leaf chlorosis and root rot accompanied by purplish to brown vascular discoloration. The symptoms were observed in four surveyed fields, 57 ha in total, and disease incidence ranged from 21 to 56%. Five symptomatic plants with leaf chlorosis and root rot were randomly collected from each field for pathogen isolation. Tissue pieces from diseased roots were surface sterilized in 75% ethanol for 30 s then rinsed with sterile distilled water three times, air dried, and placed onto potato dextrose agar (PDA) medium. Five isolates, SL1, SL2, SL3, LN and KC, were purified by single-spore culturing. On PDA, colonies grew at a rate of 2-5 mm/day and produced abundant cottony, white to pink aerial mycelia and rose pigment on the reverse side of the culture plate. From 7-day-old cultures grown on carnation leaf agar (CLA), macroconidia were straight to subarcuate, with blunt and slightly hooked apical and basal cells, had three to four septa, measured 23.4 to 44.6×3.5 to 4.2 μm (n=30). Cylindrical, napiform or oval microconidia were one to two-celled, measuring 6.3 to 22.9×2.2 to 4.9 μm (n=30). Spherical chlamydospores were intercalary or terminal, in chains. Such characteristics resembled those of the Fusarium tricinctum species comples (FTSC; Batra and Lichtwardt 1962; Leslie and Summerell 2006). To confirm the morphological diagnosis, the genomic DNA of the isolates were extracted, the translation elongation factor 1-alpha (EF-1α), RNA polymerase I largest subunit (RPB1) and second largest subunit (RPB2) genes were amplified with primers EF1/EF2, F5/G2R and 5f2/7cr respectively (O'Donnell et al. 2010), and sequenced. Maximum likelihood analysis was carried out using MEGA 7. Sequences were 97.55% to 100% identical to corresponding DNA sequences of FTSC based on GenBank and Fusarium MLST BLASTn analysis, and deposited in GenBank (ON637268.1-ON637272.1, ON637275.1-ON637279.1, ON637282.1-ON637286.1). Based on the morphological characteristics and phylogenetic analysis, the isolates were identified as F. acuminatum (SL1, SL2, SL3 and LN; Senatore et al. 2021) and F. reticulatum (KC; Moreira et al. 2019). Koch's postulates were conducted to verify the pathogenicity of individual isolates. The four-leaf stage healthy tobacco seedlings (Yunyan99, n=30) were inoculated by pouring 20 mL conidial suspension (1×107 conidia/mL) around the rhizosphere. Control seedlings were inoculated with sterilized water (n=30). All the treatments were maintained under greenhouse conditions with a 12-h light/dark photoperiod at 25±0.5℃ and 70% relative humidity for 30 days. The assay was conducted three times. Foliage chlorosis and root rot were observed on the inoculated tobacco seedlings, whereas the control seedlings remained asymptomatic after 30 days. The pathogens were reisolated from the necrotic tissue from all inoculated seedlings and were identified by sequencing partial EF-1α and RPB2 genes. Fusarium tricinctum species complex are known as an important causal of cereals Fusarium Head Blight (FHB; Laraba, et al. 2022). In China, F. acuminatum can also infect herb plants and fruits, such as Angelica sinensis, Schisandra chinensis (Ma et al. 2022; Shen et al. 2021). To our knowledge, this is the first report of root rot on tobacco caused by FTSC members in China as well as the world. This finding expands the host range known for FTSC and will be helpful for developing effective control strategies of tobacco root rot.

First report of seedling blight caused by Fusarium tricinctum on Cynanchum stauntonii in China.

Cynanchum stauntonii is a perennial herb plant of the Asclepiadaceae family. The dried roots and rhizomes have been used as medicine in China for 1500 years and are considered a remedy for cough and phlegm. In recent years, the wild C. stauntonii resources have not been sufficient for market demand, therefore, a large artificial cultivation area was established in Xinzhou, Tuanfeng and Macheng in Hubei province. In March and April 2022, serious outbreaks of seedling blight were observed in C. stauntonii in Xinzhou county (N30°48'12″, E114°49'24″), and the disease occurred on 10 to 15% of plants in five C. stauntonii nursery beds. Early symptoms included withered tips, chlorosis, stunting, yellow leaves and leaf drop, and later, seedlings die in patches. To determine the causal agent of disease, pieces (5 mm × 5 mm) of diseased tissue at the junction of disease and healthy tissue were surface disinfected by soaking in 75% ethanol for 3 min, rinsed three times with sterilized water, and pieces were placed on PDA at 25°C. Fungal isolates obtained were yellow-brown at the center and pink to white toward the periphery, and dark red pigments were observed in the agar. Isolates were cultured in synthetic low nutrient agar (SNA) and carnation leaf agar to observe the spore morphology. The macroconidia were sickle-shaped with 3-4 septate, with sizes of 30.26±2.36×3.77±0.53 μm on SNA and 33.52±2.20×3.81±0.48 μm on carnation leaf agar (n=30). Morphological characteristics of the isolates were consistent with those of Fusarium sp in the Fusarium Laboratory Manual (Leslie et al. 2006). Furthermore, the genomic DNA from a representative isolate BQ-2 was extracted, the ITS, TEF-1α, RPB1 and RPB2 genes were amplified with primers ITS1/ITS4, EF1/EF2, Fa/G2R and 5F2/7cr, respectively (Zhang et al. 2022). BLAST analysis showed that the ITS (ON935780.1), TEF-1α (OP985126.1), RPB1 (OP985125.1) and RPB2 (OP985124.1) amplicon sequence were 99.44%, 98.94%, 99.88% and 100% identical to the sequences of F. tricinctum strain (KU350724.1, AB674264.1, LC701712.1, MW474678.1), respectively. A phylogenetic tree constructed based on a concatenated sequence (ITS, TEF-1α, RPB1, RPB2) using the neighbor-joining and maximum likelihood method in MEGA7 revealed that BQ-2 grouped with concatenated sequences from four representative F. tricinctum isolates in GenBank. Based on the morphological characteristics and molecular identification, the strain BQ-2 was identified as F. tricinctum. For pathogenicity tests, 5 mm pieces of a BQ-2 colony on PDA were placed on excised leaves of healthy C. stauntonii wounded with a needle (n=5) and kept at 25±2℃. Leaves treated PDA were used as a control (Li et al.2020). After three days inoculation, the mycelia proliferated and began to infect leaf tissues. Ten days later, large parts of the detached leaves were extensively infested with the pathogen and brown. For live plant inoculation, stem bases of five healthy seedlings were punctured with sterile needle and then inoculated with BQ-2 mycelia from PDA. Controls were treated with only PDA. The seedlings began wilting after three days and at five days showed typical disease symptoms, similar to those observed in the field. The controls were asymptomatic. The pathogen was reisolated from the diseased tissues, and the colonies and microscopic characteristics were similar to those of BQ-2. To the best of our knowledge, this is the first report of F. tricinctum causing seedling blight on C. stauntonii in China. This report will provide resources and reference for controlling the increased incidence and economic losses of seedling blight on C. stauntonii.

Impact of Erwinia gerundensis as a Biocontrol Agent on the Sanitary and Technological Quality of Barley Malt

This study investigates the use of a barley-associated Erwinia gerundensis strain as a biocontrol agent for the protection of malt against fungal development and mycotoxin production during the malting of barley. First, the antifungal activity of E. gerundensis was studied via simultaneous co-cultures in Yeast Malt broth and on barley kernels artificially infected with Fusarium tricinctum. Then, the effect of E. gerundensis on the fungal load and mycotoxin content of malt obtained from naturally contaminated barley was studied by applying different doses of the bacterial strain at the steeping step of the malting process. Mycotoxin concentrations were determined using the technique of liquid chromatography coupled to the mass spectrometry detector in tandem (LC-MS/MS). Also, the effect of E. gerundensis on the technological quality of the final malt was evaluated using European Brewery Convention standard methods. The bacterial strain reduced the production of enniatin (ENN) by 97.7–99.8% and 38–68% in the liquid medium and on barley kernels, respectively, along with 0–82% Fusarium tricinctum growth reduction on barley kernels. Application of E. gerundensis at the beginning of the steeping step reduced the fungal load of the final malt by 67–72% and its ENN and deoxynivalenol (DON) contents by 32–50% and 15–33%, respectively, depending on the treatment dose. Barley malts treated with E. gerundensis showed no degradation in their technological quality, thus making E. gerundensis a potentially interesting biocontrol agent for the malting and brewing industries. Further investigation should be focused on improving the bioprotective capability of the proposed biocontrol agent.

First report of stem rot of Scrophularia ningpoensis caused by Fusarium commune in China.

First report of stem rot of Scrophularia ningpoensis caused by Fusarium commune in China.

Antibacterial Metabolites from Kiwi Endophytic Fungus Fusarium tricinctum, a Potential Biocontrol Strain for Kiwi Canker Disease

Pseudomonas syringae pv. actinidiae (Psa) is a Gram-negative bacterium causing the kiwifruit canker disease, resulting in serious economic losses to the kiwifruit industry. This study investigated the use of an endophytic fungus, Fusarium tricinctum, obtained from the kiwi plant (Actinidia chinesis) as a potential biocontrol strain against the Psa. F. tricinctum showed an inhibition rate of 59.5% in vitro against Psa. Bioassay-guided isolation was conducted on the cultural broth of F. tricinctum and seven new imidazole alkaloids, (±)-fusaritricine J ((±)-1) and fusaritricines K-P (2-7), and four enniatins (8-11) were identified. Their absolute configurations were established through extensive spectroscopic methods, quantum chemical calculations, and X-ray single crystal diffraction. Compounds 1, 4, 5, and 8-11 showed comparable anti-bacterial activities against Psa as positive control, with MIC values of 25-50 μg/mL. Further cell membrane permeability assay suggested that the most active compound 4 could destroy the bacterial cell wall structure. Hence, F. tricinctum metabolites could be applied as potential anti-Psa agents, and F. tricinctum could be considered a biocontrol strain for the control of the kiwifruit canker disease.

Activity of the botanical compound thymol against kiwifruit rot caused by Fusarium tricinctum and the underlying mechanisms.

Kiwifruit rot is an important disease caused by different fungal pathogens, which can lead to huge economic loss in the kiwifruit industry. The aims of this study were to discover an effective botanical compound that significantly inhibits the pathogens causing kiwifruit rot, evaluate its control efficacy against this disease, and reveal the underlying mechanisms. A strain of Fusarium tricinctum (GF-1), isolated from diseased kiwifruit, could cause fruit rot in both Actinidia chinensis var. chinensis and Actinidia chinensis var. deliciosa. Different botanical chemicals were used for antifungal activity test against GF-1 and thymol was the most effective one with a 50% effective concentration (EC50 ) of 30.98 mg L-1 . The minimal inhibitory concentration (MIC) of thymol against GF-1 was 90 mg L-1 . Control efficacy of thymol against kiwifruit rot was evaluated and the results indicated that thymol could effectively decrease the occurrence and spread of kiwifruit rot. The mechanisms underlying the antifungal activity of thymol against F. tricinctum were investigated, and it showed that thymol could significantly damage the ultrastructure, destroy the plasma membrane integrity, and instantaneously increase energy metabolisms of F. tricinctum. Further investigations indicated that thymol could extend shelf life of kiwifruit by increasing their storability. Thymol can effectively inhibit F. tricinctum that is one of the causal agents of kiwifruit rot. Multiple modes of action are involved in the antifungal activity. The results of this study indicate that thymol can be a promising botanical fungicide to control kiwifruit rot and provide useful references for thymol application in agriculture system. © 2023 Society of Chemical Industry.

Antifungal Activity and Effect of Plant-Associated Bacteria on Phenolic Synthesis of Quercus robur L.

Europe's forests, particularly in Ukraine, are highly vulnerable to climate change. The maintenance and improvement of forest health are high-priority issues, and various stakeholders have shown an interest in understanding and utilizing ecological interactions between trees and their associated microorganisms. Endophyte microbes can influence the health of trees either by directly interacting with the damaging agents or modulating host responses to infection. In the framework of this work, ten morphotypes of endophytic bacteria from the tissues of unripe acorns of Quercus robur L. were isolated. Based on the results of the sequenced 16S rRNA genes, four species of endophytic bacteria were identified: Bacillus amyloliquefaciens, Bacillus subtilis, Delftia acidovorans, and Lelliottia amnigena. Determining the activity of pectolytic enzymes showed that the isolates B. subtilis and B. amyloliquefaciens could not cause maceration of plant tissues. Screening for these isolates revealed their fungistatic effect against phytopathogenic micromycetes, namely Fusarium tricinctum, Botrytis cinerea, and Sclerotinia sclerotiorum. Inoculation of B. subtilis, B. amyloliquefaciens, and their complex in oak leaves, in contrast to phytopathogenic bacteria, contributed to the complete restoration of the epidermis at the sites of damage. The phytopathogenic bacteria Pectobacterium and Pseudomonas caused a 2.0 and 2.2 times increase in polyphenol concentration in the plants, respectively, while the ratio of antioxidant activity to total phenolic content decreased. Inoculation of Bacillus amyloliquefaciens and Bacillus subtilis isolates into oak leaf tissue were accompanied by a decrease in the total pool of phenolic compounds. The ratio of antioxidant activity to total phenolic content increased. This indicates a qualitative improvement in the overall balance of the oak leaf antioxidant system induced by potential PGPB. Thus, endophytic bacteria of the genus Bacillus isolated from the internal tissues of unripe oak acorns have the ability of growth biocontrol and spread of phytopathogens, indicating their promise for use as biopesticides.

First report of Fusarium tricinctum causing fruit rot on navel orange (Citrus sinensis (L.) Osbeck) in China.

Citrus sinensis (L.) Osbeck is popular with consumers for its delicious taste. In December 2020, a rot symptom causing about 15% losses of a total of 450 fruits was observed on 'Newhall' navel oranges after 70 d storage (20℃, 85%-90% RH) at Jiangxi Key Laboratory for Postharvest Technology and Nondestructive Testing of Fruits & Vegetables (28.68° N, 115.85° E). The fruits were harvested from an orchard in Ganzhou City, Jiangxi Province, China (25.53° N, 114.79° E). Incipiently, the pedicles of infected fruits were brown, the peels became softened and showed yellowish-brown lesions which, gradually expanded and had white hyphae (Fig. S1A). To isolate the pathogen, the surface of diseased fruits was disinfected with NaClO (2%) for 2 min and ethanol (75%) for 0.5 min, then washed with sterile water three times. Tissues (5 × 5 mm) around the lesion were incubated on potato dextrose agar (PDA) at 28 ± 1℃ (L: D=12: 12) for 5 days. Five cultures with similar morphology were obtained and colonies initially produced white aerial hyphae and became khaki then turned pink on PDA (Fig. S1F, G, H). Abundant microconidia, macroconidia and rare chlamydospores were observed after 10 days on PDA and no glucose PDA media (Zhang et al. 2020). Macroconidia were falciform and curved to lunate, 2-4 septa, 29.38 × 3.75 µm in size (n=50) (Fig. S1K, Fig. S3). Microconidia were oval, napiform or pyriform, 0-1 septa, 12.00 × 3.43 µm in size (n=50) (Fig. S1L1 to L4, Fig. S3). Chlamydospores were found in hyphae, ellipsoidal or orbicular (Fig. S1I-1 to I-2, J-1 to J-2). The morphological features of five isolates were similar to Fusarium (Leslie and Summerell 2006). Genomic DNA of five isolates was extracted with DNA Extraction Kit (Yeasen, Shanghai, China), ITS1/ITS4, EF1Ha/EF2Tb and fRPB2-5F/fRPB2-7cR primers were used to amplify the internal transcribed spacer region (ITS), and the transcriptional elongation factor-1 alpha (TEF-1α), and RNA polymerase II (RPB2) gene sequences (White et al. 1990; Carbone and Kohn 1999; Liu et al. 1999). The ITS, TEF-1α and RPB2 sequences of five isolates were deposited in GenBank and showed 99-100% identity with corresponding sequences from F. tricinctum (Table S1). A phylogenetic tree was constructed with ITS-TEF-1α-RPB2 concatenated sequences in MEGA7.0 (Li et al. 2021) and all five isolates were placed in F. tricinctum clade with 100% bootstrap support (Fig. S2). To confirm pathogenicity, ten healthy C. sinensis fruits were surface-sterilized with 75% ethanol and inoculated with 10 µL spore suspension (1.0 × 106 spore/mL) including five wounded (with sterilized needle) and five unwounded (Fig. S1B to E). Control fruits were inoculated with 10 µL sterile water. All fruits were incubated at 28 ± 1℃, 90% RH for 7 days. The experiment was conducted three times. The lesion diameter of inoculated wounded fruits was 21.01 ± 2.52 mm and showed similar symptoms to original rotten fruits. However, the control and unwounded fruits remained healthy. To fulfill Koch's postulates, F. tricinctum was re-isolated from the inoculated fruits and deposited in Collaborative Innovation Center of Postharvest Key Technology and Quality Safety of Fruits and Vegetables in Jiangxi Province. To our knowledge, F. tricinctum has been reported on apple tree and kiwi plant in China (Zhang et al. 2021; Ma et al. 2022), but this is the first report of F. tricinctum causing fruit rot on navel orange in China. This finding provides important information for preventing postharvest disease of citrus.