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Today's fungal plant disease control efforts tend towards environmentally friendly and reduced chemical applications. While traditional broad-spectrum fungicides provide efficient protection to many field crops, they pose a risk to the soil's beneficial microflora and a potential health hazard. Moreover, their intensive use often evokes the appearance of resistant pathogens. On the other hand, biocontrol agents such as Trichoderma spp. provide a green solution but often cannot shield the plants from aggressive disease outbreaks. Integrated biological and chemical disease control can combine the benefits of both methods while reducing their drawbacks. In the current study, such a bio-chemo approach was developed and evaluated for the first time against the maize late wilt pathogen, Magnaporthiopsis maydis. Combinations of four Trichoderma species and Azoxystrobin were tested, starting with an in vitro seed assay, then a growth room sprouts trial, and finally a semi-field, full-season pot experiment. In the plates assay, all four Trichoderma species, Trichoderma sp. O.Y. (T14707), T. longibrachiatum (T7407), T. asperellum (P1) and T. asperelloides (T203), grew (but with some delay) in the presence of Azoxystrobin minimal inhibition concentration (0.005 mg/L). The latter two species provided high protection to sprouts in the growth room and to potted plants throughout a full season in a semi-field open-enclosure trial. At harvest, the P1 and T203 bio-shielding exhibited the best parameters (statistically significant) in plant growth promotion, yield increase and late wilt protection (up to 29% health recovery and 94% pathogen suppression tracked by real-time PCR). When applied alone, the Azoxystrobin treatment provided minor (insignificant) protection. Adding this fungicide to Trichoderma spp. resulted in similar (statistically equal) results to their sole application. Still, the fact that Azoxystrobin is harmless to the beneficial Trichoderma species over a complete semi-field condition is a great opening stage for carrying out follow-up studies validating the integrated control in a commercial field situation challenged with acute disease stress. [ABSTRACT FROM AUTHOR]

Simple Summary: Maize late wilt disease (LWD) is considered the most severe corn disease in Israel and Egypt and a significant threat in other countries. The utilization of disease-resistant maize cultivars is currently our best LWD control method. Here we evaluated the predictive ability of a rapid test to examine the susceptibility/tolerance of corn varieties to the disease. The fastest assay is based on the sensitivity of corn seeds to metabolites secreted by the pathogen. However, it is only reliable in detecting highly resistant or extremely susceptible maize varieties. To identify LWD immunity of mildly tolerant hybrid plants, a full-season pot assay in the open air, under field conditions, is needed. New detection methods, including aerial visible dehydration symptom monitoring and real-time PCR-based tracking of M. maydis DNA inside the plants' tissues, were evaluated here and proved to provide a great advantage. No complete overlap exists between the fungal DNA amount and the severity of symptoms. Such a correlation exists in high sensitivity or resistance cases but not in intermediate situations. Still, the valuation of the pathogen's establishment in asymptomatic corn hybrids is a powerful means to predict each maize variety's LWD immunity. Magnaporthiopsis maydis late wilt disease (LWD) in corn is considered to be the most severe in Israel and Egypt and poses a significant threat in other countries. Research efforts extending over a period of five decades led to the development of chemical, biological, agrotechnical, physical (solar disinfection) and other means for controlling late wilt disease. Today, some applications can reduce damage even in severe cases. However, cultivating disease-resistant maize varieties is the primary means for reducing the disease's impact. The current work uses a rapid (six days) laboratory seedling pathogenicity test and a full-season open encloser semi-field conditioned pots assay (101 days) to classify maize varieties according to their LWD resistance. To better evaluate differences between the cultivars, a real-time based molecular assay was applied to track the pathogen's presence in the plants' tissues, and visible light aerial imaging was used in parallel. The findings show that in cases of extreme sensitivity or tolerance (for example, in the highly susceptible Megaton cultivar (cv.) or the resistant Hatai cv.), a similarity in the results exists between the different methods. Thus, a reliable estimate of the varieties' sensitivity can be obtained in a seed assay without the need for a test carried out throughout an entire growing season. At the same time, in most situations of partial or reduced LWD sensitivity/resistance, there is no match between the various tests, and only the entire growing season can provide the most reliable results. Tracking the amount of M. maydis DNA in the plants' bodies is a precise, sensitive scientific tool of great importance for studying the development of the disease and the factors affecting it. Yet, no complete overlap exists between the fungal DNA amount and symptom severity. Such a correlation exists in high sensitivity or resistance cases but not in intermediate situations. Still, the valuation of the pathogen's establishment in asymptomatic corn hybrids can indicate the degree of LWD immunity and the chance of susceptibility development. [ABSTRACT FROM AUTHOR]

6-pentyl-α-pyrone (6-PP) is a powerful Magnaporthiopsis maydis antifungal compound, recently discovered when the potent growth medium of Trichoderma asperellum was analyzed. Despite its high potential in plate assay, it was not inspected for plant treatment prevention. Late wilt disease, caused by the fungus M. maydis, threatens commercial maize production in high-risk areas. Thus, the search for control options against the pathogen is one of the top priorities in Israel, Egypt, and other countries. Disease-resistant maize genotypes can reduce the damages. Yet, aggressive variants of the fungus can overcome host resistance. The current study aimed at inspecting T. asperellum and its secreted metabolite, pure 6-PP, against the pathogen in plants over a full growth period. First, adding T. asperellum directly to seeds with sowing provides significant protection to sprouts (up to 42 days) in a growth room, with more than two-fold growth promotion and reduced pathogen root infection (detected by real-time PCR). The same procedure applied in a commercial field was less beneficial in rescuing the plants' growth and yield. Still, it reduced the cobs' symptoms by 11% and resulted in nine-fold lower levels of the pathogen's DNA in the stem tissue. Second, the T. asperellum purified 6-PP compound (30 µg/seed) was used in seed coating and tested against the T. asperellum secretory metabolites' crude (diluted to 50%). At the season's end, these treatments improved plant biomass by 90–120% and cob weight by 60%. Moreover, the treatments significantly (p < 0.05) reduced the symptoms (up to 20%) and pathogen infection (94–98%). The current study's results reveal the potential of 6-PP as a new fungicide against M. maydis. Such a treatment may protect maize plants from other soil diseases. [ABSTRACT FROM AUTHOR]

In recent years, worldwide scientific efforts towards controlling maize late wilt disease (LWD) have focused on eco-friendly approaches that minimize the environmental impact and health risks. This disease is considered to be the most severe threat to maize fields in Israel and Egypt, and a major growth restraint in India, Spain, and Portugal. Today's most commonly used method for LWD control involving resistant maize genotypes is under constant risk from aggressive pathogen lines. Thus, this study's objectives were to evaluate the effect of crop rotation and avoiding tillage on restraining the disease. Such an agrotechnical approach will support the continuity of soil mycorrhiza networks, which antagonize the disease's causal agent, Magnaporthiopsis maydis. The method gained positive results in previous studies, but many knowledge gaps still need to be addressed. To this end, a dual-season study was conducted using the LWD hyper-susceptible maize hybrid, Megaton cv. The trials were performed in a greenhouse and in the field over full dual-growth seasons (wheat or clover as the winter crop followed by maize as the summer crop). In the greenhouse under LWD stress, the results clearly demonstrate the beneficial effect of maize crop rotation with clover and wheat on plant weight (1.4-fold), height (1.1–1.2-fold) and cob yield (1.8–2.4-fold), especially in the no-till soil. The clover-maize growth sequence excels in reducing disease impact (1.7-fold) and pathogen spread in the host tissues (3-fold). Even though the wheat-maize crop cycle was less effective, it still had better results than the commercial mycorrhizal preparation treatment and the uncultivated non-infected soil. The results were slightly different in the field. The clover-maize rotation also achieved the best growth promotion and disease restraint results (2.6-fold increase in healthy plants), but the maize rotation with wheat showed only minor efficiency. Interestingly, pre-cultivating the soil with clover had better results in no-till soil in both experiments. In contrast, the same procedure with wheat had a better impact when tillage was applied. It may be concluded that crop rotation and soil cultivation can be essential in reducing LWD, but other factors may affect this approach's benefits in commercial field growth. [ABSTRACT FROM AUTHOR]

The destructive maize late wilt disease (LWD) has heavy economic implications in highly infected areas such as Israel, Egypt, and Spain. The disease outbreaks occur near the harvest, leading to total yield loss in severe cases. Crop rotation has long been known as an effective means to reduce plant diseases. Indeed, agricultural soil conservation practices that can promote beneficial soil and root fungi have become increasingly important. Such methods may have a bioprotective effect against Magnaporthiopsis maydis, the LWD causal agent. In this two-year study, we tested the role of crop rotation of maize with either wheat or clover and the influence of minimum tillage in restricting LWD. In the first experiment, wheat and clover were grown in pots with LWD infected soil in a greenhouse over a full winter growth period. These cultivations were harvested in the spring, and each pot’s group was split into two subgroups that underwent different land processing practices. The pots were sown with LWD-sensitive maize cultivar and tested over a whole growth period against control soils without crop rotation or soil with commercial mycorrhizal preparation. The maize crop rotation with wheat without tillage achieved prominent higher growth indices than the control and the clover crop cycle. Statistically significant improvement was measured in the non-tillage wheat soil pots in sprout height 22 days after sowing, in the healthy plants at the season’s end (day 77), and in shoot and cob wet weight (compared to the control). This growth promotion was accompanied by a 5.8-fold decrease in pathogen DNA in the plant stems. The tillage in the wheat-maize growth sequence resulted in similar results with improved shoot wet-weight throughout the season. In contrast, when maize was grown after clover, the tillage reduced this parameter. The addition of commercial mycorrhizal preparation to the soil resulted in higher growth measures than the control but was less efficient than the wheat crop cycle. These results were supported by a subsequent similar experiment that relied on soil taken from commercial wheat or clover fields. Here too, the wheat-maize growth cycle (without permanent effect for the tillage) achieved the best results and improved the plants’ growth parameters and immunity against LWD and lowered pathogen levels. In conclusion, the results of this study suggest that wheat and perhaps other crops yet to be inspected, together with the adjusted tillage system, may provide plants with better defense against the LWD pathogen. [ABSTRACT FROM AUTHOR]

Magnaporthiopsis maydisis the causal agent of severe maize late wilt disease. Disease outbreak occurs at the maize flowering and fruit development stage, leading to the plugging of the plant’s water vascular system, resulting in dehydration and collapse of the infected host plant.The pathogen is borne by alternative hosts, infected seeds, soil, and plant residues and gradually spreads to new areas and new countries. However, no soil assay is available today that can detect M. maydis infestation and study its prevalence. We recently developed a molecular quantitative Real-Time PCR (qPCR) method enabling the detection of the M. maydis DNA in plant tissues. Despite the technique’s high sensitivity, the direct examination of soil samples can be inconsistent. To face this challenge, the current work demonstrates the use of a soil bioassay involving the cultivation of a hyper-susceptible maize genotype (Megaton cultivar, Hazera Seeds Ltd., Berurim MP Shikmim, Israel) on inspected soils. The use of Megaton cv. may facilitate pathogen establishment and spread inside the plant’s tissues, and ease the isolation and enrichment of the pathogen from the soil. Indeed,this cultivar suffers from severe dehydration sudden death when grown in an infested field. The qPCR method was able to accurately and consistently identify and quantify the pathogen’s DNA in an in vitro seed assay after seven days, and in growth-chamber potted plants at as early as three weeks.These results now enable the use of this highly susceptible testing plant to validate the presence of the maize late wilt pathogen in infested soils and to evaluate the degree of its prevalence. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Late wilt of maize, caused by the fungus Harpophora maydis, is one of the most important fungal diseases in Egypt. The disease has also been reported from India, Hungary, Spain and Portugal. The pathogen survives for long periods in soil. Late wilt is currently controlled using maize varieties with reduced sensitivity, but virulent variants of the fungus may threaten these varieties. Common disease symptoms have been documented over 20 years in Upper Galilee (northern Israel), particularly the Hula Valley. Recently, prevalence of the disease has increased. This is the first confirmed report of the direct and primary cause of the disease in Israel. Isolates of the pathogen obtained from wilting maize plants were morphologically identical to those of strains found in Egypt and India. We modified a molecular method as a diagnostic assay of disease progress in an infested field in northern Israel. The assay identified the pathogen 50 d after seeding, before the emergence of disease symptoms, both in susceptible and partially resistant host plants. DNA assessment was consistent with the disease progress in a susceptible maize variety, and the pathogen also spread in partially resistant plants that showed no symptoms. Seeds of apparently healthy, partially resistant plants may therefore also spread the disease. A modified molecular method presented here is a preliminary step in developing a seed health assay. [ABSTRACT FROM AUTHOR]

Maize late wilt disease caused by the fungus Magnaporthiopsis maydis significantly damages crops in Israel and in other countries. Resistant maize cultivars are the preferred method for disease restraining. However, the pathogen populations of Spain and Egypt have varying aggressiveness, and virulent strains can overcome host resistance. In 2001 and from 2016 to -2019, 17 M. maydis strains were isolated from infected maize fields in Israel. The isolates' effects on seed germination, plant development, and disease symptoms severity were evaluated. The isolates from Israel display a diverse degree of aggressiveness that is not linked to their geographic distribution. The virulent strains are found in mixed populations, whereas less virulent M. maydis isolates exist. Aggressive strains harmed the development of plants and ears and caused severe wilting and death. In contrast, plants inoculated with less virulent strains exhibited only mild dehydration signs, and crop yield was similar to that of the non-infected control. Interestingly, different host cultivars can evoke specific virulence of M. maydis strains. Moreover, some pathogen strains significantly repress plant development, while the impact of other strains was evidenced by wilting symptoms. The current research further increases our understanding of the pathogen and our ability to control it. [Display omitted] • Israel is considered one of the world's most affected by maize late wilt disease. • This is the first work to describe Israel's pathogen dispersal and aggressiveness. • Virulent M. maydis strains scattered throughout the country in mixed populations. • Some strains mainly affect plant growth, while others disease symptoms. • Different host cultivars evoke specific M. maydi s strains' virulence. [ABSTRACT FROM AUTHOR]

Fungal pathogens are a significant threat to crops worldwide. The soil fungus, Magnaporthiopsis maydis, severely affects sensitive maize hybrids by causing the rapid wilting of plants at the maturity stage. Similarly, the soil fungus, Macrophomina phaseolina, develops in a variety of host plants, which leads to rot and plant mortality. The presence of both pathogens together in diseased cotton plants in Israel suggests possible interactions between them. Here, these relationships were tested in a series of experiments accompanied by real-time PCR tracking in maize and cotton. Despite the fact that neither of the pathogens was superior in a growth plate confrontation assay, their co-inoculum had a significant influence under field conditions. In maize sprouts and fully matured plants, infection by both pathogens (compared to inoculation with each of them alone) led to lesser amounts of M. maydis DNA but to increased amounts of M. phaseolina DNA levels. These results were obtained under a restricted water regime, while optimal water irrigation led to less pronounced differences. In water-stressed cotton sprouts, infection with both pathogens led to an increase in DNA amounts of each of the pathogens. Whereas the M. maydis DNA levels in the double infection remain high at the end of the season, a reduction in the amount of M. phaseolina DNA was observed. The double infection caused an increase in growth parameters in maize and cotton and decreased levels of dehydration in maize plants accompanied by an increase in yield production. Dehydration symptoms were minor in cotton under an optimal water supply. However, under a restricted water regime, the double infection abolished the harmful effect of M. phaseolina on the plants' development and yield. These findings are the first report of interactions between these two pathogens in maize and cotton, and they encourage expanding the study to additional plant hosts and examining the potential involvement of other pathogens. [ABSTRACT FROM AUTHOR]

The fungus Magnaporthiopsis maydis is a soil-borne, seed-borne vascular wilt pathogen that causes severe damage to sensitive Zea mays L. (maize) hybrids throughout Egypt, Israel, India, Spain, and other countries. It can undergo virulence variations and survive as spores, sclerotia, or mycelia on plant residues. Maize, Lupinus termis L. (lupine) and Gossypium hirsutum L. (cotton) are the only known hosts of M. maydis. Identification of new plant hosts that can assist in the survival of the pathogen is an essential step in restricting disease outbreak and spread. Here, by field survey and growth chamber pathogenicity test, accompanied by real-time PCR analysis, the presence of the fungal DNA inside the roots of cotton (Pima cv.) plants was confirmed in infested soil. Moreover, we identified M. maydis in Setaria viridis (green foxtail) and Citrullus lanatus (watermelon, Malali cv.). Infected watermelon sprouts had delayed emergence and development, were shorter, and had reduced root and shoot biomass. M. maydis infection also affected root biomass and phenological development of cotton plants but caused only mild symptoms in green foxtail. No M. maydis DNA was detected in Hordeum vulgare (barley, Noga cv.) and the plants showed no disease symptoms except for reduced shoot weight. These findings are an important step towards uncovering the host range and endophytic behavior of M. maydis, encouraging expanding this evaluation to other plant species. [ABSTRACT FROM AUTHOR]

Late wilt, a destructive vascular disease of maize caused by the fungus Magnaporthiopsis maydis, is characterized by relatively fast wilting of maize plants closely before the physiological maturity stage. Previously, traditional microbiology-based methods have been used to isolate the pathogen and to characterize its traits. More recently, several molecular methods have been developed, enabling accurate and sensitive examination of the pathogen spread within the host. Here, we review the methods developed in the past 10 years in Israel, which include new or modified microbial and molecular techniques to identify, monitor, and study M. maydis in controlled environments and in the field. The assays inspected are exemplified with new findings and include microbial isolation methods, microscopic and PCR or qPCR identification, spore germination evaluation, root pathogenicity assay, M. maydis hyphae or filtrate effects on grain germination and sprout development, and a field assay. These diagnostic protocols enable rapid and reliable detection and identification of the pathogen in plants and seeds and studying the pathogenesis of M. maydis in susceptible and relatively resistant maize cultivars in a contaminated field. Moreover, these techniques are important for studying the population structure, and for future development of new strategies to restrict the disease's outburst and spread. [ABSTRACT FROM AUTHOR]