Visible light-activated ZnO nanoparticles for microbial control of wheat crop.

Every year 15-50% of cereals all around the world are lost due to fungal contamination and deterioration. In addition, 25% of crops, which are used for human and animal consumption, are contaminated with mycotoxins. It is obvious, that more effective and sustainable technologies for better microbial control of crops are required. For this purpose we evaluated antibacterial and antifungal activity of ZnO nanoparticles (NPs) (10-3-5 × 10-3M) activated with visible light (405 nm, 18-30 J/cm2). Obtained data indicated that this treatment can inactivate human pathogen E. coli B by 6 log CFU without any possibility to regrowth after treatment. Wheat pathogen Fusarium oxysporum was inactivated by 51.7%. Results indicated that reactive oxygen species took place in the mechanisms of inactivation. Moreover, visible light activated ZnO NPs reduced the population of mesophiles on the surface of grains by 2.5 log CFU/g, inoculated E. coli- by 2.0 log CFU/g and naturally distributed fungi-by 2.1 log CFU/g. This treatment had no impact on visual quality of grains, did not inhibit grain germination rate and slightly promoted grain seedling growth. Concluding, the use of visible light driven photocatalysis in ZnO nanoparticles has huge potential to control plant pathogens, reduce food-borne diseases and subsequently enhance the sustainability in agriculture, meeting the increasing demands of a growing world population.

Effect of pulsed light on postharvest disease control-related metabolomic variation in melon (Cucumis melo) artificially inoculated with Fusarium pallidoroseum.

Pulsed light, as a postharvest technology, is an alternative to traditional fungicides, and can be used on a wide variety of fruit and vegetables for sanitization or pathogen control. In addition to these applications, other effects also are detected in vegetal cells, including changes in metabolism and secondary metabolite production, which directly affect disease control response mechanisms. This study aimed to evaluate pulsed ultraviolet light in controlling postharvest rot, caused by Fusarium pallidoroseum in 'Spanish' melon, in natura, and its implications in disease control as a function of metabolomic variation to fungicidal or fungistatic effects. The dose of pulsed light (PL) that inhibited F. pallidoroseum growth in melons (Cucumis melo var. Spanish) was 9 KJ m-2. Ultra-performance liquid chromatography (UPLC) coupled to a quadrupole-time-of-flight (QTOF) mass analyzer identified 12 compounds based on tandem mass spectrometry (MS/MS) fragmentation patterns. Chemometric analysis by Principal Components Analysis (PCA) and Orthogonal Partial Least Squared Discriminant Analysis (OPLS-DA) and corresponding S-Plot were used to evaluate the changes in fruit metabolism. PL technology provided protection against postharvest disease in melons, directly inhibiting the growth of F. pallidoroseum through the upregulation of specific fruit biomarkers such as pipecolic acid (11), saponarin (7), and orientin (3), which acted as major markers for the defense system against pathogens. PL can thus be proposed as a postharvest technology to prevent chemical fungicides and may be applied to reduce the decay of melon quality during its export and storage.

Dual effect of blue light on Fusariumsolani clinical corneal isolates in vitro.

The purpose was to investigate the effect of daylight-intensity blue light on F. solani isolated from the cornea of patients with fungal keratitis. Spore suspensions of 5 F. solani strains (one standard strain and 4 clinical corneal isolates) were prepared in 6-well plates. Blue light groups were irradiated by a light-emitting diode (LED) device with a peak wavelength of 454 nm at 0.5 mW/cm2 for 0 to 48 h, while the controls were maintained in darkness. Hyphal morphology in the 6-well plates was recorded at 0, 12, 24, 36, 48 h. One hundred microliters of spore suspensions of each strain at these five time points was transferred to SGA plates and cultured for 36 h at 29 °C; the number of colonies formed was counted as a measure of conidia quality and viability. Blue light has dual effects on F. solani. The hyphal length of F. solani exposed to blue light was significantly shorter than that of the control (P < 0.01), indicating that fungal growth was inhibited. Meanwhile, instead of reducing the viability of spores, blue light significantly enhanced the conidia quality and viability after at least 24 h irradiation. Daylight-intensity blue light exposure will inhibit the hyphal growth of F. solani but promote conidiation, which would be more harmful to fungal keratitis. Eliminating the influence of blue light for these patients should be taken into account.

Fusarium concolor X4 Pretreatment Suppresses Light-Induced Yellowing of High-Yield Pulp.

High-yield pulps (HYPs), such as CTMP (chemi-thermo-mechanical pulp), are attractive due to their low cost and high wood utilization. However, their drawback of rapid brightness reversion (yellowing) limits wide use of the HYPs. In this study, a fungus, Fusarium concolor X4, was applied to treat poplar CTMP for exploring the effects of biotreatment on brightness and light-induced yellowing of the pulp. The results indicated that the biotreatment with Fusarium concolor X4 could improve the brightness of poplar CTMP and inhibit light-induced yellowing of the pulp. The yellowing inhibition mechanism was explored by the analysis of enzyme production regularity during biotreatment, changes in chemical components, and the UV-Vis absorption spectra and FTIR-ATR spectra of pulps before and after biotreatment.

Inactivation of molds on the surface of wheat sprouts by chlorophyllin-chitosan coating in the presence of visible LED-based light.

The present study clearly demonstrated the significant antifungal activity of chlorophyllin-chitosan complex (Chl-CHS) after activation with visible light. This phenomenon afterwards was successfully applied for better microbial control of highly popular food- germinated wheat sprouts. Obtained results indicated that photoactivated Chl-CHS complex (0.001% Chl-0.1% CHS and 0.005% Chl-0.5% CHS, 405 nm, 76 J/cm2) considerably inhibited (83%) the growth of dominating sprout pathogenic microfungus Fusarium graminearum in vitro. Moreover, obvious delay of fungus growth by 4 days after treatment was observed. The efficiency of antifungal treatment strongly depended on used Chl-CHS complex concentration. The coating of wheat grains with Chl-CHS (0.005% Chl-0.5% CHS) and illumination with visible light (405 nm; 76 J/cm2) inactivated the molds on the surface of grains by 79%. It is important to note, that no grain surface microstructure damage observed by SEM imaging have been found. No inhibiting effects on seed germination process, viability, average weight of grains, length of seedlings and content of chlorophyll a and chlorophyll b in the seedlings or eventually visual quality after Chl-CHS coating of grains and illumination with visible light have been observed. In conclusion, chlorophyllin-chitosan coating in the concert with visible light has great potential as cost-effective, environmentally friendly and sustainable strategy for better microbial control of highly contaminated sprouts.

Biofilm characterization of Fusarium solani keratitis isolate: increased resistance to antifungals and UV light.

Fusarium solani has drawn phytopathogenic, biotechnological, and medical interest. In humans, it is associated with localized infections, such as onychomycosis and keratomycosis, as well as invasive infections in immunocompromised patients. One pathogenicity factor of filamentous fungi is biofilm formation. There is still only scarce information about the in vitro mechanism of the formation and composition of F. solani biofilm. In this work, we describe the biofilm formed by a clinical keratomycosis isolate in terms of its development, composition and susceptibility to different antifungals and ultraviolet light (UV) at different biofilm formation stages. We found five biofilm formation stages using scanning electron microscopy: adherence, germination, hyphal development, maturation, and cell detachment. Using epifluorescence microscopy with specific fluorochromes, it was elucidated that the extracellular matrix consists of carbohydrates, proteins, and extracellular DNA. Specific inhibitors for these molecules showed significant biofilm reductions. The antifungal susceptibility against natamycin, voriconazole, caspofungin, and amphotericin B was evaluated by metabolic activity and crystal violet assay, with the F. solani biofilm preformation to 24 h increased in resistance to natamycin, voriconazole, and caspofungin, while the biofilm preformation to 48 h increased in resistance to amphotericin B. The preformed biofilm at 24 h protected and reduced UV light mortality. F. solani isolate could produce a highly structured extra biofilm; its cellular matrix consists of carbohydrate polymers, proteins, and eDNA. Biofilm confers antifungal resistance and decreases its susceptibility to UV light. The fungal biofilm functions as a survival strategy against antifungals and environmental factors.

Comparative transcriptomic analysis unveils interactions between the regulatory CarS protein and light response in Fusarium.

BACKGROUND: The orange pigmentation of the agar cultures of many Fusarium species is due to the production of carotenoids, terpenoid pigments whose synthesis is stimulated by light. The genes of the carotenoid pathway and their regulation have been investigated in detail in Fusarium fujikuroi. In this and other Fusarium species, such as F. oxysporum, deep-pigmented mutants affected in the gene carS, which encodes a protein of the RING-finger family, overproduce carotenoids irrespective of light. The induction of carotenogenesis by light and its deregulation in carS mutants are achieved on the transcription of the structural genes of the pathway. We have carried out global RNA-seq transcriptomics analyses to investigate the relationship between the regulatory role of CarS and the control by light in these fungi. RESULTS: The absence of a functional carS gene or the illumination exert wide effects on the transcriptome of F. fujikuroi, with predominance of genes activated over repressed and a greater functional diversity in the case of genes induced by light. The number of the latter decreases drastically in a carS mutant (1.1% vs. 4.8% in the wild-type), indicating that the deregulation produced by the carS mutation affects the light response of many genes. Moreover, approximately 27% of the genes activated at least 2-fold by light or by the carS mutation are coincident, raising to 40% for an 8-fold activation threshold. As expected, the genes with the highest changes under both regulatory conditions include those involved in carotenoid metabolism. In addition, light and CarS strongly influence the expression of some genes associated with stress responses, including three genes with catalase domains, consistent with roles in the control of oxidative stress. The effects of the CarS mutation or light in the transcriptome of F. oxysporum were partially coincident with those of F. fujikuroi, indicating the conservation of the objectives of their regulatory mechanisms. CONCLUSIONS: The CarS RING finger protein down-regulates many genes whose expression is up-regulated by light in wild strains of the two investigated Fusarium species, indicating a regulatory interplay between the mechanism of action of the CarS protein and the control by light.

Design and effectiveness of pulsed electric fields towards seed disinfection.

BACKGROUND: Seeds harbor different microorganisms on their surfaces that degrade seed quality, thus causing an economic loss. Even though different approaches are available for the disinfection of seed surfaces, there is a need to develop environmentally friendly and sustainable technologies. A bench-scale pulsed electric field (PEF) unit was designed to inactivate microflora of eight seeds after which the resultant vigor of the treated seeds was determined. RESULTS: Significant reductions were obtained in endogenous natural and inoculated pathogenic (Alternaria brassica and Xanthomonas campestris pv. campestris, Drechslera graminea and Fusarium graminearum) microflora of seeds. The survival ratios of total aerobic mesophilic bacteria and of total mold and yeast decreased significantly for winter wheat and barley, parsley, onion, lettuce, tomato, and garden rocket with the PEF treatments of 240 and 960 J. A significant increase in germination ratio was observed for winter wheat and barley, lettuce, and tomato with 960 J. Germination energy increased for parsley with 240 J and for winter wheat and barley, lettuce, tomato, and garden rocket with 960 J. A better root development and seedling were found for winter barley. CONCLUSION: PEFs are a viable option to both disinfect seed surfaces and improve seed vigor. © 2019 Society of Chemical Industry.

Feasibility of 3D UV-C treatment to reduce fungal growth and mycotoxin loads on maize and wheat kernels.

Fungal disease of grain crops is a concern for the agricultural industry, resulting in economic losses. Aside from severe yield losses, mycotoxigenic fungi such as Penicillium and Fusarium can produce harmful mycotoxins, including deoxynivalenol (DON), zearalenone (ZEN), and ochratoxin A (OTA). This proof-of-concept study explored the feasibility and effects of ultraviolet (UV) C light at 253.7 nm to reduce fungal and mycotoxin loads on model surfaces as well as on maize and wheat kernels using benchtop 2D and 3D illumination strategies. Reduction of Penicillium verrucosum (98.6%) and Fusarium graminearum (88.8%) on agar was achieved using a UV-C dose of 100 mJ cm-2. Naturally occurring fungal growth resembling P. verrucosum on maize was reduced by 79% after exposure to 5000 mJ cm-2. Similarly, fungal growth resembling F. graminearum on maize was reduced by 60% with 1000 mJ cm-2. On wheat, significant reduction of fungal growth was not observed. Maximal reduction of DON (97.3%), ZEN (75.4%), and OTA (91.2%) on filter paper was obtained using 15,000 mJ cm-2. The overall reduction of DON (30%; 14%), ZEN (52%; 42%), and OTA (17%; 6%) on maize and wheat, respectively, was lower than on filter paper. Moisture and crude protein content as well as percent germination of maize kernels were not affected by UV-C treatment up to 5000 mJ cm-2. This study has shown that 3D UV-C treatment is a feasible option for reducing Fusarium and Penicillium growth on maize kernels and, at higher doses, decreasing ZEN by ~ 50%.

Effect of management of organic wastes on inactivation of Brassica nigra and Fusarium oxysporum f.sp. lactucae using soil biosolarization.

BACKGROUND: Soil biosolarization is a promising alternative to conventional fumigation. Volatile fatty acids (VFAs) produced in the soil through fermentation of amended organic matter can affect pest inactivation during biosolarization. The objective was to determine how soil amended with organic wastes that were partially stabilized through either composting or anaerobic digestion affected the inactivation of Brassica nigra (BN; a weed) and Fusarium oxysporum f. sp. lactucae (FOL; a phytopathogenic fungus). RESULTS: The mortality of BN seeds in the biosolarized soil was 12% higher than in the solarized soil, although this difference was not significant. However, a significant correlation between BN mortality and VFA accumulation was observed. The number of FOL colony-forming units (CFU) in solarized samples at 5 cm was 34 CFU g-1 of soil, whereas in the biosolarized samples levels were below the limit of quantification. At 15 cm, these levels were 100 CFU g-1 for solarized samples and < 50 CFU g-1 of soil for the biosolarized samples. Amendment addition positively affected the organic matter and potassium content after the solarization process. CONCLUSION: The organic waste stabilization method can impact downstream biosolarization performance and final pest inactivation levels. This study suggests that organic waste management practices can be leveraged to improve pest control and soil quality. © 2018 Society of Chemical Industry.

Fusarium graminearum in Stored Wheat: Use of CO2 Production to Quantify Dry Matter Losses and Relate This to Relative Risks of Zearalenone Contamination under Interacting Environmental Conditions.

Zearalenone (ZEN) contamination from Fusarium graminearum colonization is particularly important in food and feed wheat, especially during post-harvest storage with legislative limits for both food and feed grain. Indicators of the relative risk from exceeding these limits would be useful. We examined the effect of different water activities (aw; 0.95-0.90) and temperature (10-25 °C) in naturally contaminated and irradiated wheat grain, both inoculated with F. graminearum and stored for 15 days on (a) respiration rate; (b) dry matter losses (DML); (c) ZEN production and (d) relationship between DML and ZEN contamination relative to the EU legislative limits. Gas Chromatography was used to measure the temporal respiration rates and the total accumulated CO2 production. There was an increase in temporal CO2 production rates in wetter and warmer conditions in all treatments, with the highest respiration in the 25 °C × 0.95 aw treatments + F. graminearum inoculation. This was reflected in the total accumulated CO2 in the treatments. The maximum DMLs were in the 0.95 aw/20-25 °C treatments and at 10 °C/0.95 aw. The DMLs were modelled to produce contour maps of the environmental conditions resulting in maximum/minimum losses. Contamination with ZEN/ZEN-related compounds were quantified. Maximum production was at 25 °C/0.95-0.93 aw and 20 °C/0.95 aw. ZEN contamination levels plotted against DMLs for all the treatments showed that at ca <1.0% DML, there was a low risk of ZEN contamination exceeding EU legislative limits, while at >1.0% DML, the risk was high. This type of data is important in building a database for the development of a post-harvest decision support system for relative risks of different mycotoxins.

Complete cure of Fusarium solani sp. complex onychomycosis with Qs NdYAG treatment.

The incidence of non dermatophytic mould (NDM) onychomycosis (OM) has been steadily increasing Fusarium spp is the most common cause of NDM OM in most geographical locations. Fusarium spp and other NDMs are largely resistant to commonly used anti-fungals. The successful use of laser and light based devices has been demonstrated in dermatophytic OM, but there is no previous report of their successful use in any NDM OM. We describe a patient with OM caused by Fusarium solani spp, who was clinically (with a normal appearing nail) and mycologically (with negative microscopy and culture on repeated samples) cured of her infection following treatment with 2 sessions of Qs NdYAG (532nm and 1064nm) given 1 month apart.

New insight into the disinfection mechanism of Fusarium monoliforme and Aspergillus niger by TiO2 photocatalyst under low intensity UVA light.

Titanium dioxide (TiO2) photocatalytic reaction has great potential for the disinfection of harmful pathogens. However, the disinfection mechanisms of TiO2 photocatalysis are not yet well-known for fungi and protozoa. In this work, the photocatalytic disinfection mechanism of Fusarium monoliforme and Aspergillus niger under low intensity UVA light (365nm, <10W/m2) was studied at the ultrastructural level. Photocatalytic treatments showed that the photocatalytic oxidation of 10% TiO2 based paint was efficacious in the complete disinfection of F. monoliforme under low intensity UVA light. No growth of F. monoliforme was observed on agar plate in the subsequent dark. Transmission electron microscopy (TEM) of F. monoliforme exposed to TiO2 photocatalysis treatment showed a distinct damage to electron-dense outer cell wall, but not to an underlying electron-transparent layer cell wall. The TEM image revealed that the UVA-light only did not damage cell wall, cell membrane and cellular organelles. Unlike, A. niger was more sensitive to UVA-light. Serious destructions of cell membrane and cellular organelles were shown in A. niger exposed to UVA-light only and photocatalytic treatments. However, morphological change in A. niger cell wall was only observed in photocatalytic treatment. Changes to the outermost melanin like layer and cell wall of A. niger spore due to photocatalytic treatment were greatly apparent while the intracellular organelles of A. niger spore were not affected. Therefore, regrowth of A. niger on agar plate was expected from the germination of A. niger spore in the subsequent dark. These observations give a better understanding of the photocatalytic disinfection mechanism toward fungi.

Survival of Fusarium solani f. sp. cucurbitae and Fungicide Application, Soil Solarization, and Biosolarization for Control of Crown and Foot Rot of Zucchini Squash.

Fusarium crown and foot rot of zucchini squash (Cucurbita pepo L.) caused by Fusarium solani f. sp. cucurbitae is one of the major diseases affecting zucchini squash production in Almería, Spain. Experiments were conducted to determine the pathogen's ability to survive in infested bags of perlite and to test several control methods under greenhouse conditions. The pathogen survived in the bags for at least 20 months with enough inoculum at that time to produce disease symptoms in zucchini plants, although disease severity was significantly reduced after 14 months. A total of 14 zucchini cultivars were inoculated with F. solani f. sp. cucurbitae, and all were highly susceptible to the disease. Eight fungicides and two microbial products, Trichoderma harzianum and Rhizophagus irregularis, were tested to determine their efficacy for the control of this disease. Prochloraz, carbendazim, and thiophanate-methyl, which are not labeled for use in zucchini in Spain, were highly effective for the control of the disease, while the other products were ineffective. Two soil solarization and biosolarization experiments were conducted in a greenhouse for 45-day periods during the summer. Inocula in the soil samples decreased by more than 99%, indicating the efficacy of completely closing the greenhouse windows, solarization, and biosolarization in reducing inoculum. Fungicide applications, crop rotation for at least two years, and soil solarization or biosolarization are promising control methods for this disease.

Controlled biosynthesis of silver nanoparticles using nitrate reductase enzyme induction of filamentous fungus and their antibacterial evaluation.

The controlled synthesis of silver nanoparticles (AgNPs) using cell-free filtrate of Fusarium oxysporum fungus was investigated. The effect of fungal incubation period on nanoparticle formation and nitrate reductase enzyme activity was studied using UV-visible spectroscopy and Harley assay, respectively. The highest AgNP formation was observed in the cell-free filtrate of biomass harvested at the early stationary phase where the NR enzyme activity is the maximum. Mixing of the cell-free filtrates of fungal cultures obtained at 23, 28, and 33 °C with silver nitrate solution confirms the higher productivity of AgNP biosynthesis using the cell-free filtrate of fungus incubated at 28 °C. The effect of some factors such as carbon and nitrate sources and light in fungal incubation period on nitrate reductase induction and AgNP formation was also evaluated. In conclusion, increasing nitrate and carbon sources and presence of light induced NR enzyme and produced AgNPs with smaller size, higher monodispersity, and productivity. Results revealed that the presence of ammonium prevents the NR enzyme secretion and causes to the lower productivity of AgNPs.

Enhancement of pectinase production by ultraviolet irradiation and diethyl sulfate mutagenesis of a Fusarium oxysporum isolate.

Fusarium oxysporum strain BM-201 was treated with ultraviolet (UV) radiation to obtain a high pectinase-producing strain. Mutant UV-10-41 was obtained and then treated by diethyl sulfate. Next, the mutant UV-diethyl sulfate-43 derived from UV-10-41 was selected as high pectinase-producing strain. Mutant UV-diethyl sulfate-43 was incubated on slant for 10 generations, demonstrating that the pectinase-producing genes were stable. Pectinase activity reached 391.2 U/mL, which is 73.6% higher than that of the original strain.

A non-linear model for temperature-dependent sporulation and T-2 and HT-2 production of Fusarium langsethiae and Fusarium sporotrichioides.

This research has produced new quantitative data on the sporulation and T-2+HT-2 toxin production that could be further integrated to develop a comprehensive disease or toxin prediction model for Fusarium langsethiae and Fusarium sporotrichioides. Experiments were conducted to determine the effect of temperature or incubation time on sporulation and the effect of temperature on T-2+HT-2 toxin production of strains of the two species. F. sporotrichioides demonstrated a preference for higher temperatures than F. langsethiae during sporulation; the optimum temperature was 24.5 ± 0.7 °C for F. langsethiae and 32.3 ± 2.1 °C for F. sporotrichioides, according to the Beta equation fitted to the data. The dynamics of sporulation over different incubation times were fitted by a Gompertz function. The maximum spore production was estimated to be after 18 and 8 d incubation at optimum temperatures for F. langsethiae and F. sporotrichioides, respectively. F. sporotrichioides produced more T-2+HT-2 than F. langsethiae. The best fit of the effect of temperature on T-2+HT-2 production in wheat grains was obtained with a Beta equation showing an optimum temperature of 14.7 ± 0.8 °C for F. langsethiae and 12.1 ± 0.2 °C for F. sporotrichioides. The optimum temperature for mycotoxin production was lower than for sporulation.

Ag doped hollow TiO2 nanoparticles as an effective green fungicide against Fusarium solani and Venturia inaequalis phytopathogens.

Chemical-based pesticides are widely used in agriculture to protect crops from insect infestation and diseases. However, the excessive use of highly toxic pesticides causes several human health (neurological, tumor, cancer) and environmental problems. Therefore nanoparticle-based green pesticides have become of special importance in recent years. The antifungal activities of pure and Ag doped (solid and hollow) TiO2 nanoparticles are studied against two potent phytopathogens, Fusarium solani (which causes Fusarium wilt disease in potato, tomato, etc) and Venturia inaequalis (which causes apple scab disease) and it is found that hollow nanoparticles are more effective than the other two. The antifungal activities of the nanoparticles were further enhanced against these two phytopathogens under visible light exposure. The fungicidal effect of the nanoparticles depends on different parameters, such as particle concentration and the intensity of visible light. The minimum inhibitory dose of the nanoparticles for V. inaequalis and F. solani are 0.75 and 0.43 mg/plate. The presence of Ag as a dopant helps in the formation of stable Ag-S and disulfide bonds (R-S-S-R) in cellular protein, which leads to cell damage. During photocatalysis generated (•)OH radicals loosen the cell wall structure and this finally leads to cell death. The mechanisms of the fungicidal effect of nanoparticles against these two phytopathogens are supported by biuret and triphenyl tetrazolium chloride analyses and field emission electron microscopy. Apart from the fungicidal effect, at a very low dose (0.015 mg/plate) the nanoparticles are successful in arresting production of toxic napthoquinone pigment for F. solani which is related to the fungal pathogenecity. The nanoparticles are found to be effective in protecting potatoes affected by F. solani or other fungi from spoiling.

The tubulin cofactor A is involved in hyphal growth, conidiation and cold sensitivity in Fusarium asiaticum.

BACKGROUND: Tubulin cofactor A (TBCA), one of the members of tubulin cofactors, is of great importance in microtubule functions through participating in the folding of α/ß-tubulin heterodimers in Saccharomyces cerevisiae. However, little is known about the roles of TBCA in filamentous fungi. RESULTS: In this study, we characterized a TBCA orthologue FaTBCA in Fusarium asiaticum. The deletion of FaTBCA caused dramatically reduced mycelial growth and abnormal conidiation. The FaTBCA deletion mutant (ΔFaTBCA-3) showed increased sensitivity to low temperatures and even lost the ability of growth at 4°C. Microscopic observation found that hyphae of ΔFaTBCA-3 exhibited blebbing phenotypes after shifting from 25 to 4°C for 1- or 3-day incubation and approximately 72% enlarged nodes contained several nuclei after 3-day incubation at 4°C. However, hyphae of the wild type incubated at 4°C were phenotypically indistinguishable from those incubated at 25°C. These results indicate that FaTBCA is involved in cell division under cold stress (4°C) in F. asiaticum. Unexpectedly, ΔFaTBCA-3 did not exhibit increased sensitivity to the anti-microtubule drug carbendazim although quantitative real-time assays showed that the expression of FaTBCA was up-regulated after treatment with carbendazim. In addition, pathogenicity assays showed that ΔFaTBCA-3 exhibited decreased virulence on wheat head and on non-host tomato. CONCLUSION: Taken together, results of this study indicate that FaTBCA plays crucial roles in vegetative growth, conidiation, temperature sensitivity and virulence in F. asiaticum.

The white collar complex is involved in sexual development of Fusarium graminearum.

Sexual spores (ascospores) of Fusarium graminearum, a homothallic ascomycetous fungus, are believed to be the primary inocula for epidemics of the diseases caused by this species in cereal crops. Based on the light requirement for the formation of fruiting bodies (perithecia) of F. graminearum under laboratory conditions, we explored whether photoreceptors play an important role in sexual development. Here, we evaluated the roles of three genes encoding putative photoreceptors [a phytochrome gene (FgFph) and two white collar genes (FgWc-1 and FgWc-2)] during sexual development in F. graminearum. For functional analyses, we generated transgenic strains lacking one or two genes from the self-fertile Z3643 strain. Unlike the wild-type (WT) and add-back strains, the single deletion strains (ΔFgWc-1 and ΔFgWc-2) produced fertile perithecia under constant light on complete medium (CM, an unfavorable medium for sexual development) as well as on carrot agar (a perithecial induction condition). The expression of mating-type (MAT) genes increased significantly in the gene deletion strains compared to the WT under both conditions. Deletion of FgFph had no significant effect on sexual development or MAT gene expression. In contrast, all of the deletion strains examined did not show significant changes in other traits such as hyphal growth, mycotoxin production, and virulence. A split luciferase assay confirmed the in vivo protein-protein interactions among three photoreceptors along with FgLaeA, a global regulator of secondary metabolism and fungal development. Introduction of an intact copy of the A. nidulans LreA and LreB genes, which are homologs of FgWc-1 and FgWc-2, into the ΔFgWc-1 and ΔFgWc-2 strains, respectively, failed to repress perithecia formation on CM in the gene deletion strains. Taken together, these results demonstrate that FgWc-1 and FgWc-2, two central components of the blue-light sensing system, negatively regulate sexual development in F. graminearum, which differs from the regulation pattern in A. nidulans.