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Background Hyperspectral imaging is emerging as a promising approach for plant disease identification. The large and possibly redundant information contained in hyperspectral data cubes makes deep learning based identification of plant diseases a natural fit. Here, we deploy a novel 3D deep convolutional neural network (DCNN) that directly assimilates the hyperspectral data. Furthermore, we interrogate the learnt model to produce physiologically meaningful explanations. We focus on an economically important disease, charcoal rot, which is a soil borne fungal disease that affects the yield of soybean crops worldwide. Results Based on hyperspectral imaging of inoculated and mock-inoculated stem images, our 3D DCNN has a classification accuracy of 95.73% and an infected class F1 score of 0.87. Using the concept of a saliency map, we visualize the most sensitive pixel locations, and show that the spatial regions with visible disease symptoms are overwhelmingly chosen by the model for classification. We also find that the most sensitive wavelengths used by the model for classification are in the near infrared region (NIR), which is also the commonly used spectral range for determining the vegetative health of a plant. Conclusion The use of an explainable deep learning model not only provides high accuracy, but also provides physiological insight into model predictions, thus generating confidence in model predictions. These explained predictions lend themselves for eventual use in precision agriculture and research application using automated phenotyping platforms.

The mung bean crop ( Vigna radiata (L.) R. Wilczek) is widely recognized as a key source of pulse food worldwide. However, this crop suffers substantial yield losses due to humid environments, particularly from infestations by the fungal pathogen Macrophomina phaseolina , which causes charcoal rot disease. This infestation results in significant agronomic losses, affecting both the crop’s growth characteristics and overall yield. Previous research suggests that these losses can be mitigated through environmentally friendly soil amendments, such as biochar, as well as by applying various nanofungicides. This study aims to explore the potential of biochar and zinc oxide nanoparticles (ZnONPs) to reduce the severity of charcoal rot disease and enhance the agronomic traits and yield of mung bean plants affected by this disease. The experiment was conducted in triplicate, applying ZnONPs at three concentrations (5, 10, and 20 mg. L − 1 ) via foliar spraying, combined with two levels of biochar (20 g and 40 g per pot). Positive and negative control treatments were also included for comparison. The results demonstrated that applying 40 g of biochar per pot and 20 mg. L − 1 of foliar-applied ZnONPs increased the activities of the anti-oxidative defence enzymes. Additionally, this treatment strategy boosted the plants’ disease resistance mechanisms, leading to lower mortality rates and reduced levels of malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) by 61.7% and 49.23%. Moreover, the treatment positively impacted key growth parameters, increasing total chlorophyll content by 43%, plant height by 47%, and legume count per plant by 80.4%. The application of biochar and ZnONPs also improved seed protein content, reflecting an enhancement in nutritional quality. This study supports the use of biochar and ZnONPs as biostimulants to manage yield losses in mung bean crops affected by charcoal rot disease. The future prospects of using ZnONPs and biochar as treatments in agriculture are promising, as they offer innovative, eco-friendly solutions to enhance crop productivity, improve soil health, and reduce reliance on synthetic chemicals, paving the way for more sustainable and resilient agricultural systems.

Endophytes can induce the defence responses and modulates physiological attributes in host plants during pathogen attacks. In the present study, 127 bacterial endophytes (BEs) were isolated from different parts of healthy soybean plant. Among them, two BEs (M-2 and M-4) resulted a significant antagonistic property against Macrophomina phaseolina, causes charcoal rot disease in soybean. The antagonistic potential was evaluated through dual culture plate assay, where M-4 expressed higher antifungal activity than M-2 against M. phaseolina. The M-4 produces cell wall degrading enzymes viz. cellulase (145.71 ± 1.34 μgmL−1), chitinase (0.168 ± 0.0009 unitmL−1) and β,1–3 endoglucanase (162.14 ± 2.5 μgmL−1), which helps in cell wall disintegration of pathogens. Additionally, M-4 also can produce siderophores, indole-3-acetic acid (IAA) (17.03 ± 1.10 μgmL−1) and had a phosphate solubilization potential (19.89 ± 0.26 μgmL−1). Further, GC–MS profiling of M-4 has been carried out to demonstrate the production of lipophilic secondary metabolites which efficiently suppress the M. phaseolina defensive compounds under co-culture conditions. Bio-efficacy study of M-4 strain shown a significant reduction in disease incidence around 60 and 80% in resistant and susceptible varieties of soybean, respectively. The inoculation of M-4 potentially enhances the physiological attributes and triggers various defence responsive enzymes viz. superoxide dismutase (SOD), phenol peroxidase (PPO), peroxidase (PO) and catalase (CAT). The histopathological study also confirmed that M-4 can reduce the persistence of microsclerotia in root and shoot tissue. Conclusively, M-4 revealed as an efficient biocontrol agent that can uses multifaceted measures for charcoal rot disease management, by suppress the M. phaseolina infection and enhance the physiological attributes of soybean.

Charcoal rot disease, caused by the fungus Macrophomina phaseolina, results in major economic losses in soybean production in southern USA. M. phaseolina has been proposed to use the toxin (-)-botryodiplodin in its root infection mechanism to create a necrotic zone in root tissue through which fungal hyphae can readily enter the plant. The majority (51.4%) of M. phaseolina isolates from plants with charcoal rot disease produced a wide range of (-)-botryodiplodin concentrations in a culture medium (0.14–6.11 µg/mL), 37.8% produced traces below the limit of quantification (0.01 µg/mL), and 10.8% produced no detectable (-)-botryodiplodin. Some culture media with traces or no (-)-botryodiplodin were nevertheless strongly phytotoxic in soybean leaf disc cultures, consistent with the production of another unidentified toxin(s). Widely ranging (-)-botryodiplodin levels (traces to 3.14 µg/g) were also observed in the roots, but not in the aerial parts, of soybean plants naturally infected with charcoal rot disease. This is the first report of (-)-botryodiplodin in plant tissues naturally infected with charcoal rot disease. No phaseolinone was detected in M. phaseolina culture media or naturally infected soybean tissues. These results are consistent with (-)-botryodiplodin playing a role in the pathology of some, but not all, M. phaseolina isolates from soybeans with charcoal rot disease in southern USA.

The soil fungus Macrophomina phaseolina, the charcoal rot disease agent, poses a major threat to cotton fields. In Israel, highly infected areas are also inhabited by the maize pathogen Magnaporthiopsis maydis. This study reveals the relationships between the two pathogens and their impact on cotton sprouts. Infecting the soil 14 days before sowing (DBS) with each pathogen or with M. phaseolina before M. maydis caused a strong inhibition (up to 50–65%) of the sprouts’ development and survival, accompanied by each pathogen’s high DNA levels in the plants. However, combined or sequence infection with M. maydis first led to two distinct scenarios. This pathogen acted as a beneficial protective endophyte in one experiment, leading to significantly high emergence and growth indices of the plants and a ca. 10-fold reduction in M. phaseolina DNA in the sprouts’ roots. In contrast, M. maydis showed strong virulence potential (with 43–69% growth and survival suppression) in the other experiment, proving its true nature as an opportunist. Interestingly, soil inoculation with M. phaseolina first, 14 DBS (but not at sowing), shielded the plants from M. maydis’ devastating impact. The results suggest that the two pathogens restrict each other, and this equilibrium may lead to a moderate disease burst.

The application of algae has been considered a key element for integrated disease management in sustainable agriculture. These organisms can act as a bio-stimulant for induction of resistance against a variety of abiotic and biotic agents that cause economical loss to crop production globally. Charcoal rot disease caused by Macrophomina phaseolina (Tassi) Goid. is one of the biotic agents restricting strawberry ( Fragaria × ananassa Duch.) yield in many cultivation sites. Herein, the foliar application of brown alga ( Sargassum angustifolium ) was investigated for the reduction of the disease symptoms and improvement of vegetative and reproductive indices in strawberries under greenhouse conditions. The results showed that alga-treated infected plants showed symptom remission. Moreover, vegetative and reproductive indices of alga-treated plants were significantly improved. Biochemical analysis showed that in alga-treated infected plants the total phenol, flavonoids, and total antioxidant activity were significantly increased compared to non-treated infected plants. Furthermore, the content of defense-related enzymes, viz. phenylalanine ammonia-lyase and polyphenol oxidase, were significantly increased in the infected plants pre-treated with the alga extract. Foliar application of S. angustifolium extract can induce defense responses in strawberry plants infected by M. phaseolina leading to improved growth indices of the plants. It can be concluded that S. angustifolium extract is a promising source of bio-stimulants for induction of disease resistance against charcoal rot disease in strawberry cultivations.

An isolate of Macrophomina phaseolina from muskmelons (Cucumis melo) was reported by Dunlap and Bruton to produce red pigment(s) in melons and in culture in the presence of added glycine, alanine, leucine, or asparagine in the medium, but not with some other amino acids and nitrogen-containing compounds. We explored the generality and mechanism of this pigment production response using pathogenic M. phaseolina isolates from soybean plants expressing symptoms of charcoal rot disease. A survey of 42 M. phaseolina isolates growing on Czapek-Dox agar medium supplemented with glycine confirmed pigment production by 71% of isolates at the optimal glycine concentration (10 g/L). Studies in this laboratory have demonstrated that some pathogenic isolates of M. phaseolina produce the mycotoxin (−)-botryodiplodin, which has been reported to react with amino acids, proteins, and other amines to produce red pigments. Time course studies showed a significant positive correlation between pigment and (−)-botryodiplodin production by selected M. phaseolina isolates with maximum production at seven to eight days. Pigments produced in agar culture medium supplemented with glycine, beta-alanine, or other amines exhibited similar UV-vis adsorption spectra as did pigments produced by (±)-botryodiplodin reacting in the same agar medium. In a separate study of 39 M. phaseolina isolates, red pigment production (OD520) on 10 g/L glycine-supplemented Czapek-Dox agar medium correlated significantly with (−)-botryodiplodin production (LC/MS analysis of culture filtrates) in parallel cultures on un-supplemented medium. These results support pigment production on glycine-supplemented agar medium as a simple and inexpensive in-culture method for detecting (−)-botryodiplodin production by M. phaseolina isolates.

The effects of temperature and pH on the growth of 45 Hungarian Macrophomina phaseolina isolates from different locations and hosts were compared on the basis of their genetic diversity. One Spanish and two Serbian isolates were also included in the experiment. The most favourable temperature regimes for the development of the isolates ranged between 25 and 35°C. The optimal pH for the pathogen varied between 4.0 and 6.0, but growth was observed on potato dextrose agar even at pH values of 3.0, 7.0 and 8.0. RAPD analysis with 13 different primer pairs generated 148 unambiguous bands. RFLP analysis involving 8 different restriction endonucleases was performed on a 1550 bp fragment of the rDNA region containing internal transcribed spacers (ITS1, ITS2), the 5.8S rDNA and part of the 25S rDNA. The greatest genetic distance values were obtained for three isolates, two from Hungary and one from Spain, which had similar values, but were quite distinct from all the others. A strong positive correlation was observed between the genetic distances and the growth parameters measured at various temperatures, and between the geographical data and the growth data sets at different pH values, but the correlation was less strong in the latter case. While Hungarian M. phaseolina populations are thought to reproduce clonally, the present results indicate the coexistence of different haplotypes in this area, and besides the geographical dominance of a given haplotype it was found that a closer genetic relationship might exist between spatially distinct haplotypes.

The effects of soil solarization combined or not with millet residues or paunch contents amendments, on the survival of Macrophomina phaseolina (Tassi) Goid. and development of charcoal rot of cowpea (Vigna unguiculata), were assessed in a naturally infested soil. Solarization increased the soil temperature to 50 degree C for at least 4 h per day during June, leading to a significant reduction (44%) in soil inoculum of M. phaseolina. Paunch contents or millet residues amendment (3 t ha super(-1)) caused 16% or 35% reduction of initial inoculum density, respectively. The combination of paunch contents or millet residues amendments followed by solarization, resulted in the strongest effects on inoculum density, with reductions of 46% or 66%, respectively. The reduction in disease severity, as expressed by the area under the disease progress curve, was 78% or 96% for the combination of millet residues or paunch contents amendments and solarization, respectively. The stronger effect of the treatments on disease severity than on inoculum density may be explained by a weakening effect caused by the treatments on the remaining inoculum. Our results suggest that in the Sahelian zone the combination of solarization and organic amendment can be a credible alternative to pesticides for managing charcoal rot disease and improving cowpea yield in fields with heavy infestations with M. phaseolina.

Charcoal rot, caused by Macrophomina phaseolina (Tassi) Goid., is an economically important disease of oilseed plants in Northern Iran. Seventy isolates of M. phaseolina were obtained from different hosts, including soybean ( Glycine max L.), and sunflower ( Helianthus annuus L.) in the northern oilseed planting regions of Iran. RAPD-PCR amplification profiles, by using of six random OPA primers (kit A) showed polymorphisms among the isolates. The primer OPA-13 (5'_ CAGCACCCAC_3') amplified the genomic DNA of all isolates of M. phaseolina producing 4-12 bands of sizes between 0.25 and 2.3 kb. Unweighted pair-group method with arithmetic mean (UPGMA) analysis classified the isolates into the nine major groups with 64% similarity. Pathogenicity of isolates was evaluated at seedling stage of soybean, sunflower, and maize ( Zea mays L.) plants under in vitro conditions. None of the isolates were pathogenic on corn, while all of the isolates infected soybean and sunflower seedlings. Isolates were more virulent on soybean than sunflower. The disease index on sunflower and soybean varied between 19-24 and 27-30 respectively. These results indicated a significant pathogenic and genetic variability within the Iranian isolates of M. phaseolina. Cultivation with crop rotation was probably tended to induce less diversity of the pathogen isolates.