Conidial Germination Research Articles

Comprehensive analysis of the composition of the major phospholipids during the asexual life cycle of the filamentous fungus Aspergillus nidulans

Filamentous fungi undergo significant cellular morphological changes during their life cycle. It has recently been reported that deletions of genes that are involved in phospholipid synthesis led to abnormal hyphal morphology and differentiation in filamentous fungi. Although these results suggest the importance of phospholipid balance in their life cycle, comprehensive analyses of cellular phospholipids are limited. Here, we performed lipidomic analysis of A. nidulans during morphological changes in a liquid medium and of colonies on a solid medium. We observed that the phospholipid composition and transcription of the genes involved in phospholipid synthesis changed dynamically during the life cycle. Specifically, the levels of phosphatidylethanolamine, and highly unsaturated phospholipids increased during the establishment of polarity. Furthermore, we demonstrated that the phospholipid composition in the hyphae at colony margins is similar to that during conidial germination. Furthermore, we demonstrated that common and characteristic phospholipid changes occurred during germination in A. nidulans and A. oryzae, and that species-specific changes also occurred. These results suggest that the exquisite regulation of phospholipid composition is crucial for the growth and differentiation of filamentous fungi.

Identification of Colletotrichum aenigma as the new causal agent of leaf blight disease on Aucuba japonica Thunb., and screenings of effective fungicides for its sustainable management.

Aucuba japonica Thunb is an evergreen woody ornamental plant with significant economic and ecological values. It also produces aucubin, showing a variety of biological activities. It is widely planted in the southwest region of China, including karst landscape areas in Guizhou Province. In January 2022, a serious leaf blight disease was observed on the leaves of A. japonica in the outdoor gardens of Guizhou University, Guiyang, Guizhou, China. The causal agent was identified as Colletotrichum aenigma through amplification and sequencing of the internal transcribed spacer (ITS) region, translation of the chitin synthase (CHS) and actin (ACT) genes, and morphological characterizations. Koch's postulates were confirmed by its pathogenicity on healthy leaves, including re-isolation and identification. To our knowledge, this is the first report of C. aenigma causing leaf blight on A. japonica worldwide. To identify pathogen characteristics that could be utilized for future disease management, the effects of temperature and light on mycelial growth, conidia production, and conidial germination, and the effects of humidity on conidial germination were studied. Optimal temperatures for mycelial growth of C. aenigma BY827 were 25-30°C, while 15°C and 35°C were favorable for conidia production. Concurrently, alternating 10-h light and 14-h dark, proved to be beneficial for mycelial growth and conidial germination. Additionally, conidial germination was enhanced at 90% humidity. In vitro screenings of ten chemical pesticides to assess their efficacy in suppressing C. aenigma representative strain BY827. Among them, difenoconazole showed the best inhibition rate, with an EC50 (concentration for 50% of maximal effect) value of 0.0148 μg/ml. Subsequently, field experiment results showed that difenoconazole had the highest control efficiency on A. japonica leaf blight (the decreasing rate of disease incidence and decreasing rate of disease index were 44.60 and 47.75%, respectively). Interestingly, we discovered that C. aenigma BY827 may develop resistance to mancozeb, which is not reported yet among Colletotrichum spp. strains. In conclusion, our study provided new insights into the causal agent of A. japonica leaf blight, and the effective fungicides evaluated provided an important basis and potential resource for the sustainable control of A. japonica leaf blight caused by C. aenigma in the field.

Effect of adjuvants, fungicides and insecticides on the growth of Trichoderma koningiopsis Th003

The effect of 44 agrochemicals (fungicides, insecticides and adjuvants) on the mycelial growth and germination conidia of Trichoderma koningiopsis Th003 was determined by seeding of 5 mm discs of fungal mycelium in Sabouraud agar supplemented with each agrochemical and seeding of conidia suspended in solutions of agrochemicals in water agar. For the adjuvants, their effect on the phyllospheric establishment of T. koningiopsis Th003 was determined by following their population in cape gooseberry leaflets inoculated with the fungus suspended in them. Eight fungicides did not inhibit the fungus mycelial growth or the conidia germination. Fenhexamid - Tebuconazole, Flutriafol and Kasugamicina inhibited it between 34 and 48% without affecting germination, Thiram - Pyrimethanil, Prochloraz, Tiabendazol, Spiroxamina and Triadimenol - Tebuconazole inhibited the growth and Thiram - Pyrimethanil and Dodine did not allow conidia germination. Insecticides and adjuvants presented an inhibition of up to 70% but did not affect the conidia germination. No negative effects of the adjuvants on the phyllosphere establishment of T. koningiopsis Th003 were observed, recovering from the treatments approximately 1x103 CFU g-1.

Calcium disodium edetate controls citrus green mold by chelating Mn2+ and targeting pyruvate synthesis in the pathogen

Edetate salts are a kind of food additive regarded as candidates for chemical fungicides because of their lower toxicity and higher safety for the environment and human health. The antifungal activities of edetate salts have been determined in some phytopathogens except Penicillium digitatum, which causes the most destructive fungal disease during the citrus postharvest period. In the present study, we investigated the molecular mechanisms of calcium disodium edetate (EDTA-Na2Ca) against P. digitatum in deepth. The results suggested that EDTA-Na2Ca inhibited the P. digitatum P44 strain in vitro at a minimum inhibitory/fungicidal concentration (MIC/MFC) of 50 mmol L−1. Meanwhile, EDTA-Na2Ca also inhibited P. digitatum in vivo by enhancing the host defense system. The P44 strain treated with EDTA-Na2Ca displayed a delayed conidial germination rate, expanded hyphal tips, thickened septa and cell walls, and destroyed mitochondria. Transcriptome sequencing and qRT-PCR analyses indicated that EDTA-Na2Ca suppressed the P44 strain primarily by disturbing the carbohydrate metabolism pathway, especially the synthesis of pyruvate in glycolysis. In addition, the adverse effects, including those on mycelial growth, pyruvate synthesis and superoxide dismutase activity, of EDTA-Na2Ca could be reversed by exogenous Mn2+, implying that Mn2+ chelation is an essential mode of action. In summary, the current work verifies that EDTA-Na2Ca is a potential antifungal drug that cures citrus green mold probably by chelating Mn2+ and targeting pyruvate synthesis in P. digitatum.

Initial host invasion of Colletotrichum gloeosporioides is dampened by 1-methylcyclopropene due to blocking ethylene sensitivity of the fungus

Ethylene (ET) is not only a fruit ripening hormone, but also plays important roles in plant-pathogen interactions, as it is involved in both plant defense and fungal virulence. 1-methylcyclopropene (1-MCP) is a competitive inhibitor for ET being applied as a postharvest preservation agent to delay fruit ripening. Plant pathogenic fungi can also sense ET to promote fungal conidial germination, appressorium development, and pathogenicity. However, it is unclear whether 1-MCP could regulate postharvest disease development by interfering with the ET-sensing in pathogenic fungi. In this study, we identified the effects of ET and 1-MCP on the growth and development of Colletotrichum gloeosporioides, a fungal pathogen that causes anthracnose disease in various crops, and explored the regulatory effects of 1-MCP on the pathogenic fungus during its host invasions. The results showed that 1-MCP can inhibit the ET responses of C. gloeosporioides, including the ET-induced conidial germination, appressorium formation, and expression of the ethylene-responsive marker gene. Further analysis revealed that 1-MCP treatment dampened the host invasion of the fungus, and thus decreased disease severity on lime and strawberry fruit. Collectively, this study reveals the action mode of 1-MCP in inhibiting the fungal ET sensitivity and highlights the unexplored potential of using 1-MCP as a postharvest treatment to control postharvest diseases caused by fungal pathogens.

Effector secretion and stability in the maize anthracnose pathogen Colletotrichum graminicola requires N-linked protein glycosylation and the ER chaperone pathway.

N-linked protein glycosylation is a conserved and essential modification mediating protein processing and quality control in the endoplasmic reticulum (ER), but how this contributes to the infection cycle of phytopathogenic fungi is largely unknown. In this study, we discovered that inhibition of protein N-glycosylation severely affected vegetative growth, hyphal tip development, conidial germination, appressorium formation, and, ultimately, the ability of the maize (Zea mays) anthracnose pathogen Colletotrichum graminicola to infect its host. Quantitative proteomics analysis showed that N-glycosylation can coordinate protein O-glycosylation, glycosylphosphatidylinositol anchor modification, and endoplasmic reticulum quality control (ERQC) by directly targeting the proteins from the corresponding pathway in the ER. We performed a functional study of the N-glycosylation pathway-related protein CgALG3 and of the ERQC pathway-related protein CgCNX1, which demonstrated that N-glycosylation of ER chaperone proteins is essential for effector stability, secretion, and pathogenicity of C. graminicola. Our study provides concrete evidence for the regulation of effector protein stability and secretion by N-glycosylation.

Aspergillus ochraceus biocontrol by Hanseniaspora opuntiae in vitro and on coffee fruits

Aspergillus ochraceus is an ochratoxin-producing fungus which contaminates coffee. In this study the antifungal effect of the yeast Hanseniaspora opuntiae on three Aspergillus ochraceus strains (IOC 4417, IOC 4462, Ao 14) was evaluated in vitro and on coffee fruits. H. opuntiae (106 and 107 cells mL−1) reduced in vitro fungal growth from 82% to 87%, when co-cultivated with A. ochraceus. The yeast cell free supernatant (CFS) inhibited conidial germination from 76.5% to 92.5%, and hyphal growth from 54% to 78%. The yeast (107 and 109 cells mL−1) applied on coffee fruits delayed fruit decay by A. ochraceus (IOC 4417 and Ao 14) until the 9th day, and was significantly different (p < 0.05) from the controls. Furthermore, the ultrastructure of the yeast-fungus interaction on the coffee fruit surface showed yeast attachment to A. ochraceus hyphae, and morphological alterations in fungal structures, with hyphal abnormalities, such as tortuous hyphae with irregular, non-uniform surface compared to the control without yeast. H. opuntiae showed efficacy as biocontrol agent and, to the best of our knowledge, this is the first study on the antifungal activity of H. opuntiae against A. ochraceus on coffee fruits Nevertheless, application of H. opuntiae to the crop in the field requires further studies.

Herbicide 2,4-dichlorophenoxyacetic acid interferes with MAP kinase signaling in Fusarium graminearum and is inhibitory to fungal growth and pathogenesis

Plant hormones are important for regulating growth, development, and plant-pathogen interactions. Some of them are inhibitory to growth of fungal pathogens but the underlying mechanism is not clear. In this study, we found that hyphal growth of Fusarium graminearum was significantly reduced by high concentrations of IAA and its metabolically stable analogue 2,4-dichlorophenoxyacetic acid (2,4-D). Besides inhibitory effects on growth rate, treatments with 2,4-D also caused significant reduction in conidiation, conidium germination, and germ tube growth. Treatments with 2,4-D had no obvious effect on sexual reproduction but significantly reduced TRI gene expression, toxisome formation, and DON production. More importantly, treatments with 2,4-D were inhibitory to infection structure formation and pathogenesis at concentrations higher than 100 µM. The presence of 1000 µM 2,4-D almost completely inhibited plant infection and invasive growth. In F. graminearum, 2,4-D induced ROS accumulation and FgHog1 activation but reduced the phosphorylation level of Gpmk1 MAP kinase. Metabolomics analysis showed that the accumulation of a number of metabolites such as glycerol and arabitol was increased by 2,4-D treatment in the wild type but not in the Fghog1 mutant. Transformants expressing the dominant active FgPBS2S451D T455D allele were less sensitive to 2,4-D and had elevated levels of intracellular glycerol and arabitol induced by 2,4-D in PH-1. Taken together, our results showed that treatments with 2,4-D interfere with two important MAP kinase pathways and are inhibitory to hyphal growth, DON biosynthesis, and plant infection in F. graminearum.

3-Octanol controls gray mold on postharvest fruit by inducing autophagy of Botrytis cinerea

Gray mold caused by Botrytis cinerea is a devastating postharvest disease, resulting in severe fruit decay and enormous economic loss. Volatile secondary metabolites produced by plants or microorganisms are potential resources for developing fungicide alternatives. Here, we explored the antifungal activity of 3-octanol, a safe fungal volatile secondary metabolite, against B. cinerea. 3-Octanol exhibited a dose-dependent inhibitory effect on the development of B. cinerea both in vitro and in vivo. 3-Octanol treatment reduced the cell viability and suppressed the conidial germination of B. cinerea. Moreover, the transmission electron microscope analysis suggested that 3-octanol induces autophagy and autolysosome formation of B. cinerea. Further quantitative real-time PCR analysis indicated that 3-octanol induces the expression of autophagy-related genes and inhibits the expression of germination-related genes. These results suggest that 3-octanol controls gray mold by inducing autophagy, reducing cell viability, and suppressing the conidial germination of B. cinerea. Our findings suggest the potential role of 3-octanol in managing gray mold on postharvest fruit and provide a theoretical basis for the antifungal activity of 3-octanol against B. cinerea.

Recombinant Fusion Protein Containing Plant Nigellothionin Regulates the Growth of Food-Spoiling Fungus (Aspergillus niger).

This study aimed to obtain a recombinant chimeric protein named trx-NsW2 via theheterologous expression of the multifunctional antimicrobial peptide nigellothionin from black cumin (Nigella sativa L.) seeds in the Escherichia coli system. The protein was purified using a combination of Ni-NTA affinity chromatography and reversed-phase HPLC. Based on the HPLC calibration, the total yield of the protein was calculated to be 650 mg/L of bacterial culture. The fungistatic activity of trx-NsW2 against the food-spoiling fungus Aspergillus niger was demonstrated as itinhibited the maturation of conidiawithout affecting conidial germination or fungal growth. In contrast to mature nigellothionin NsW2, the fusion protein showeda low level of cytotoxicity towards both normal and tumor cell lines at concentrationsof up to 100-200 µM. Interestingly, at lower concentrations, it even stimulated cytokinesis. These findings are of critical importance for applying chimeric antimicrobial proteins obtained via microbiological synthesis in applied science.

Effective management of powdery mildew in cantaloupe plants using nighttime applications of UV light.

Ultraviolet light at wavelengths from 254 to 283 nm has been reported to effectively suppress powdery mildews in several crops, including some cucurbits. Its use to suppress powdery mildew (Podosphaera xanthii) specifically in cantaloupe has not been previously reported. We evaluated the foregoing technology in cantaloupe fields for suppression of powdery mildew and possible effects on plant growth, and yield. In a controlled laboratory study, greenhouse-grown cantaloupe plants were exposed to a gradient of UV-C (254 nm) doses during darkness, and the effects upon powdery mildew development and the plant were evaluated. We also evaluated the efficacy of nighttime applications of UV-C at 100 and 200 J/m2 against powdery mildew on adaxial leaf surfaces in greenhouse, high-tunnel and open-field plantings. UV-C at the foregoing doses reduced sporulation and germination of conidia P. xanthii without damaging plants. On cantaloupe seedlings in the greenhouse, disease severity was equivalently suppressed at all doses and frequencies of applications of the light. In high-tunnel and open-field experiments, the most effective control of powdery mildew was provided by UV-C applied at 200 J/m2 twice every week, where suppression provided by UV-C was generally equal to and sometimes better than the fungicide treatment. The foregoing UV-C dose and frequency of application also provided the highest yield under field conditions, indication that UV-C treatment is a promising technology for commercially-relevant suppression of powdery mildew on cantaloupe in a variety of growing systems.

MoJMJD6, a Nuclear Protein, Regulates Conidial Germination and Appressorium Formation at the Early Stage of Pathogenesis in Magnaporthe oryzae.

In plant-pathogen interactions, Magnaporthe oryzae causes blast disease on more than 50 species of 14 monocot plants, including important crops such as rice, millet, and most 15 recently wheat. M. oryzae is a model fungus for studying plant-microbe interaction, and the main source for fungal pathogenesis in the field. Here we report that MoJMJD6 is required for conidium germination and appressorium formation in M. oryzae. We obtained MoJMJD6 mutants (ΔMojmjd6) using a target gene replacement strategy. The MoJMD6 deletion mutants were delayed for conidium germination, glycogen, and lipid droplets utilization and consequently had decreased virulence. In the ΔMojmjd6 null mutants, global histone methyltransferase modifications (H3K4me3, H3K9me3, H3K27me3, and H3K36me2/3) of the genome were unaffected. Taken together, our results indicated that MoJMJD6 function as a nuclear protein which plays an important role in conidium germination and appressorium formation in the M. oryzae. Our work provides insights into MoJMJD6-mediated regulation in the early stage of pathogenesis in plant fungi.

Green synthesis of chitosan silver nanocomposites and their antifungal activity against Colletotrichum truncatum causing anthracnose in chillies

Biopolymer-based metal nanocomposites are gaining importance due to their safety, stability, and ease of synthesis. Chitosan and silver-based nanoparticles have been found to be effective as anti-fungal agents. In this study, we tested the fungicidal effect of green synthesized chitosan silver nanocomposites (CS-AgNPs) against the chilli anthracnose pathogen Colletotrichum truncatum (syn. C. capsici). The nanoparticles were synthesized at 90 ± 1 °C under alkaline condition. Chitosan was used as a reducing and stabilizing agent in the synthesis. The formation of nanoparticles (NPs) was indicated by a change in the colour of the solution to yellow. UV-Visible spectroscopy of the synthesized nanoparticles showed a surface plasmon resonance peak between 406 and 420 nm. The synthesized silver nanoparticles were small, and the average particle size distribution was 4 nm, as characterized by TEM and FESEM. The Zeta potential measurement of the synthesized nanoparticles ranged from + 41.9 mV and + 50.5 mV. The nanoparticles were further characterized by XRD and FTIR analysis. The nanocomposites showed antifungal activity against C. truncatum in an in vitro conidial germination assay even at a concentration of 0.0005%. In vivo assay using detached chilli fruit (Capsicum annuum) showed that the nanocomposites significantly inhibited anthracnose in both preventive and curative measures. Even when applied at a concentration of 0.0625%, the nanocomposites exhibited growth-promoting activity with chilli seeds. Thus, the synthesized nanocomposites have a dual benefit of growth promotion as well as an effective antifungal agent in preventing postharvest anthracnose disease in chilli.

Effects of temperature and leaf wetness duration on pathogens causing preharvest fruit rots on tomato

Preharvest fruit rots are serious tomato diseases. This study showed that Botrytis cinerea was a significant problem, and that Alternaria (A. alternata, A. solani) was the main pathogen identified from tomatoes showing the symptoms of preharvest fruit rots. Fusarium oxysporum and Rhizoctonia solani were also found causing important preharvest fruit rots in tomatoes. This is the first time F. oxysporum and R. solani have been identified in Greece as pathogens causing preharvest fruit rots on tomatoes. The optimal temperatures for A. alternata, A. solani, B. cinerea, and F. oxysporum mycelial growth and conidial germination were generally found to be between 20 and 30 °C, while the wetness duration of 24 hours seemed to be optimal for conidial germination and 84 hours for R. solani sclerotial germination. Based on the above findings, logistic regression models that adequately described the impact of pre-inoculation temperature and wetness on infection incidence in tomato fruit can be created. As the minimum, maximum, optimal temperatures and wetness duration are generally similar for the main pathogens causing preharvest fruit rots in tomato, it is possibly to develop a predicting predictive model to forecast the preharvest infection of tomato fruit. So enabling advice to be given to growers to when control methods should be applied.

Resistance risk and molecular mechanism associated with resistance to picoxystrobin in Colletotrichum truncatum and Colletotrichum gloeosporioides

Anthracnose, caused by Colletotrichum truncatum and C. gloeosporioides, is amongst the most serious diseases of soybean in China. Picoxystrobin, a quinone outside inhibitor fungicide, is commonly used for the control of anthracnose. Its resistance risk and mechanism in C. truncatum and C. gloeosporioides are unclear. In this study, the sensitivities of 128 C. truncatum and 121 C. gloeosporioides isolates to picoxystrobin were investigated, and unimodal distributions were observed with average EC50 values of 0.7740 and 1.1561 μg mL−1, respectively. Eleven picoxystrobin-resistant mutants of C. truncatum and six mutants of C. gloeosporioides were acquired, with EC50 values varying from 5.40–152.96 and 13.53–28.30 μg mL−1, respectively. Compared to the parental isolates, mutants showed similar or higher relative fitness in conidial production and germination, and pathogenicity. Collectively, the resistance risk of C. truncatum and C. gloeosporioides to picoxystrobin is moderate to high. There was positive cross-resistance between picoxystrobin and pyraclostrobin, but not between picoxystrobin and fluazinam, difenoconazole, or propiconazole. The G143S mutation in Cyt b protein was detected in seven high-resistant mutants of C. truncatum (RF>100), and G137R occurred in four moderate-resistant mutants (RF<50). Contrastingly, there were no point mutations in Cyt b of any C. gloeosporioides mutants. Molecular docking confirmed that two mutations conferred different resistance levels to picoxystrobin. Under greenhouse trials, picoxystrobin did not control mutants with the G143S mutation, those bearing G137R or no point mutation were somewhat controlled, but at a lower level compared to wild-type isolates. These results showed that integrated management strategies should be implemented to preserve fungicide effectiveness.

Homeobox transcription factor HbxA influences expression of over one thousand genes in the model fungus Aspergillus nidulans.

In fungi, conserved homeobox-domain proteins are transcriptional regulators governing development. In Aspergillus species, several homeobox-domain transcription factor genes have been identified, among them, hbxA/hbx1. For instance, in the opportunistic human pathogen Aspergillus fumigatus, hbxA is involved in conidial production and germination, as well as virulence and secondary metabolism, including production of fumigaclavines, fumiquinazolines, and chaetominine. In the agriculturally important fungus Aspergillus flavus, disruption of hbx1 results in fluffy aconidial colonies unable to produce sclerotia. hbx1 also regulates production of aflatoxins, cyclopiazonic acid and aflatrem. Furthermore, transcriptome studies revealed that hbx1 has a broad effect on the A. flavus genome, including numerous genes involved in secondary metabolism. These studies underline the importance of the HbxA/Hbx1 regulator, not only in developmental processes but also in the biosynthesis of a broad number of fungal natural products, including potential medical drugs and mycotoxins. To gain further insight into the regulatory scope of HbxA in Aspergilli, we studied its role in the model fungus Aspergillus nidulans. Our present study of the A. nidulans hbxA-dependent transcriptome revealed that more than one thousand genes are differentially expressed when this regulator was not transcribed at wild-type levels, among them numerous transcription factors, including those involved in development as well as in secondary metabolism regulation. Furthermore, our metabolomics analyses revealed that production of several secondary metabolites, some of them associated with A. nidulans hbxA-dependent gene clusters, was also altered in deletion and overexpression hbxA strains compared to the wild type, including synthesis of nidulanins A, B and D, versicolorin A, sterigmatocystin, austinol, dehydroaustinol, and three unknown novel compounds.

Melatonin targets MoIcl1 and works synergistically with fungicide isoprothiolane in rice blast control.

Melatonina natural harmless molecule-displays versatile roles in human health and crop disease control such as for rice blast. Rice blast, caused by the filamentous fungus Magnaporthe oryzae, is one devastating disease of rice. Application of fungicides is one of the major measures in the control of various crop diseases. However, fungicide resistance in the pathogen and relevant environmental pollution are becoming serious problems. By screening for possible synergistic combinations, here, we discovered an eco-friendly combination for rice blast control, melatonin, and the fungicide isoprothiolane. These compounds together exhibited significant synergistic inhibitory effects on vegetative growth, conidial germination, appressorium formation, penetration, and plant infection by M. oryzae. The combination of melatonin and isoprothiolane reduced the effective concentration of isoprothiolane by over 10-fold as well as residual levels of isoprothiolane. Transcriptomics and lipidomics revealed that melatonin and isoprothiolane synergistically interfered with lipid metabolism by regulating many common targets, including the predicted isocitrate lyase-encoding gene MoICL1. Furthermore, using different techniques, we show that melatonin and isoprothiolane interact with MoIcl1. This study demonstrates that melatonin and isoprothiolane function synergistically and can be used to reduce the dosage and residual level of isoprothiolane, potentially contributing to the environment-friendly and sustainable control of crop diseases.

Biological control of green and blue molds on postharvest lemon by lactic acid bacteria

Postharvest green and blue molds caused mainly by Penicillium digitatum and P. italicum conduce to economic losses in citrus production. Here, the potential antifungal activity of four lactobacilli strains was assessed against local isolates of these fungi to control postharvest diseases on lemons. Inhibitory activity of L. paraplantarum CRL 1905, L. fermentum CRL 973, L. casei CRL 1110 and L. reuteri CRL1101 strains was assayed by the culture overlay method. Also, inhibition of conidial germination by cell-free supernatants (CFS) obtained from 24 and 48 h-cultures of these lactobacilli strains (CFS24 and CFS48, respectively) was tested by microtiter assay. CRL 1905 showed the strongest fungicidal activity and was selected for further studies. When the antifungal compound/s produced by CRL 1905 strain were investigated, it was observed that the ability of CFS24 to inhibit conidial germination was lost after pH neutralization, inferring an acidic nature of the metabolite/s. Finally, to detect whether CFS24 had a potential application to control of Penicillium rot on lemons, a fruit decay test was performed. It was demonstrated that ten-fold concentrated CFS24 was able to decrease the disease incidence by ∼ 80 %. In conclusion, the use of extracellular components derived from CRL 1905 with antifungal activity could be an eco-friendly alternative to control postharvest decays of lemon.

Analysis of the emerging physical network in young mycelia

Filamentous fungi develop intricate hyphal networks that support mycelial foraging and transport of resources. These networks have been analyzed recently using graph theory, enabling the development of models that seek to predict functional traits. However, attention has focused mainly on mature colonies. Here, we report the extraction and analysis of the graph corresponding to Trichoderma atroviride mycelia only a few hours after conidia germination. To extract the graph for a given mycelium, a mosaic conformed of multiple bright-field, optical microscopy images is digitally processed using freely available software. The resulting graphs are characterized in terms of number of nodes and edges, average edge length, total mycelium length, hyphal growth unit, maximum edge length and mycelium diameter, for colonies between 8 h and 14 h after conidium germination. Our results show that the emerging hyphal network grows first by hyphal elongation and branching, and then it transitions to a stage where hyphal-hyphal interactions become significant. As a tangled hyphal network develops with decreasing hyphal mean length, the mycelium maintains long (∼2 mm) hyphae—a behavior that suggests a combination of aggregated and dispersed architectures to support foraging. Lastly, analysis of early network development in Podospora anserina reveals striking similarity with T. atroviride, suggesting common mechanisms during initial colony formation in filamentous fungi.

Germination and Invasion of Paraphoma radicina on Roots of a Susceptible and a Resistant Alfalfa Cultivar (Medicago sativa).

Alfalfa Paraphoma root rot (APRR) (Paraphoma radicina) is a recently described alfalfa disease widely distributed in China, first reported in 2020. So far, the resistance levels of 30 alfalfa cultivars to APRR have been characterized; however, the resistance mechanisms among these cultivars remain unknown. In the present study, the alfalfa resistance mechanisms against APRR were investigated by studying the difference of P. radicina infecting susceptible (Gibraltar) and resistant (Magnum II) alfalfa cultivars under the light microscope and scanning electronic microscope. The conidial germination and germ tube growth in the root exudates of different resistant cultivars were also compared. The results revealed that conidial germination, germ tube development, and P. radicina penetration into root tissues of resistant plants were delayed. In susceptible and resistant cultivars, P. radicina infected roots by penetrating epidermal cells and the intercellular space between epidermal cells. During the infection process, germ tubes penetrated the root surface directly or formed appressoria. However, the penetration percentage on the susceptible cultivar was significantly higher than on the resistant cultivar, irrespective of the infection route. Moreover, disintegrated conidia and germ tubes were observed on resistant cultivar roots at 48 h postinoculation. The conidial germination and germ tube growth in root exudates of susceptible cultivars were significantly higher than in resistant cultivars. The current findings implied that the alfalfa resistance mechanism might be related to root exudates. These findings could provide insights into the alfalfa resistance mechanism following P. radicina infection.