Integrated Management of the Cotton Charcoal Rot Disease Using Biological Agents and Chemical Pesticides.

Charcoal rot disease (CRD), caused by the phytopathogenic fungus Macrophomina phaseolina, is a significant threat to cotton production in Israel and worldwide. The pathogen secretes toxins and degrading enzymes that disrupt the water and nutrient uptake, leading to death at the late stages of growth. While many control strategies were tested over the years to reduce CRD impact, reaching that goal remains a significant challenge. The current study aimed to establish, improve, and deepen our understanding of a new approach combining biological agents and chemical pesticides. Such intervention relies on reducing fungicides while providing stability and a head start to eco-friendly bio-protective Trichoderma species. The research design included sprouts in a growth room and commercial field plants receiving the same treatments. Under a controlled environment, comparing the bio-based coating treatments with their corresponding chemical coating partners resulted in similar outcomes in most measures. At 52 days, these practices gained up to 38% and 45% higher root and shoot weight and up to 78% decreased pathogen root infection (tracked by Real-Time PCR), compared to non-infected control plants. Yet, in the shoot weight assessment (day 29 post-sowing), the treatment with only biological seed coating outperformed (p < 0.05) all other biological-based treatments and all Azoxystrobin-based irrigation treatments. In contrast, adverse effects are observed in the chemical seed coating group, particularly in above ground plant parts, which are attributable to the addition of Azoxystrobin irrigation. In the field, the biological treatments had the same impact as the chemical intervention, increasing the cotton plants' yield (up to 17%), improving the health (up to 27%) and reducing M. phaseolina DNA in the roots (up to 37%). When considering all treatments within each approach, a significant benefit to plant health was observed with the bio-chemo integrated management compared to using only chemical interventions. Specific integrated treatments have shown potential in reducing CRD symptoms, such as applying bio-coating and sprinkling Azoxystrobin during sowing. Aerial remote sensing based on high-resolution visible-channel (RGB), green-red vegetation index (GRVI), and thermal imaging supported the above findings and proved its value for studying CRD control management. This research validates the combined biological and chemical intervention potential to shield cotton crops from CRD.

The cotton charcoal rot causal agent, Macrophomina phaseolina, biological and chemical control.

The fungus Macrophomina phaseolina causes charcoal rot disease (CRD) in cotton, whose symptoms develop in the late stages of growth and result in wilting and death. Despite significant research efforts to reduce disease incidences, effective control strategies against M. phaseolina are an ongoing scientific effort. Today's CRD control tends toward green options to reduce the chemicals' environmental footprint and health risks. Here, different Trichoderma species were examined separately and in combination with Azoxystrobin (AS) in semi-field open-enclosure pots and a commercial field throughout a full season. In the pot experiment, the T. asperellum (P1) excelled and led to improvement in growth (13%-14%, day 69) and crops (the number of capsules by 36% and their weight by 78%, day 173). The chemical treatment alone at a low dose had no significant impact. Still, adding AS improved the effect of T. longibrachiatum (T7507) and impaired P1 efficiency. Real-time PCR monitoring of the pathogen DNA in the plants' roots at the harvest (day 176), revealed the efficiency of the combined treatments: T. longibrachiatum (T7407 and T7507) + AS. In a commercial field, seed dressing with a mixture of Trichoderma species (mix of P1, T7407, and Trichoderma sp. O.Y. 7107 isolate) and irrigation of their secreted metabolites during seeding resulted in the highest yields compared with the control. Applying only AS irrigation at a low dose (2,000 cc/ha), with the sowing, was the second best in promoting crops. The molecular M. phaseolina detection showed that the AS at a high dose (4,000 cc/ha) and the biological mix treatments were the most effective. Reducing the AS chemical treatment dosages by half impaired its effectiveness. Irrigation timing, also studied here, is proven vital. Early water opening during the late spring suppresses the disease outburst and damages. The results demonstrated the benefits of CRD bio-shielding and encouraged to explore the potential of a combined bio-chemo pest control approach. Such interphase can be environmentally friendly (reducing chemical substances), stabilize the biological treatment in changing environmental conditions, achieve high efficiency even in severe CRD cases, and reduce the development of fungicide resistance.

Using computer vision, image analysis and UAVs for the automatic recognition and counting of common cranes (Grus grus).

Long-term monitoring of wildlife numbers traditionally uses observers, which are frequently inefficient and inaccurate due to their variable experience/training, are costly and difficult to sustain over time. Furthermore, there are other inhibiting factors for wildlife counting, such as: inhabiting inaccessible areas, fear of humans, and nocturnal behavior. There is a need to develop new technologies that will automatically identify and count wild animals in order to determine the appropriate management protocol. In this study, an advanced and accurate method for automatically calculating the number of cranes (Grus grus), using thermal cameras at night and visible light (RGB) cameras during the day onboard unmanned aerial vehicles (UAVs), based on image analysis and computer vision, was developed. The cranes congregate at night in a large communal roost, making it possible to count the birds while they are relatively static and all together. Each bird was counted individually by creating a standardized tool to determine population numbers for management, using image analysis and automatic processing. A dedicated algorithm was developed that aimed to identify the cranes based on their spectral characteristics (typical temperature, shape, size) and to effectively separate the cranes from the typical background. The automatic segmentation and counting of roosting common cranes using UAV nighttime thermal images had an Overall Accuracy (OA) of 91.47%, User's Accuracy (UA) of 99.68%, and Producer's Accuracy (PA) of 91.74%. The computer vision and machine learning algorithm based on the YOLO v3 platform of daytime RGB UAV images of common cranes at the feeding station yielded an overall loss accuracy level of 2.25%, with a mean square error of 1.87, OA of 94.51%, UA of 99.91%, PA of 94.59%. These results are highly encouraging, and although the algorithms were developed for the purpose of counting cranes, they could be adapted for other counting purposes for wildlife management.

Cultivars Resistance Assay for Maize Late Wilt Disease.

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.

Crop Rotation and Minimal Tillage Selectively Affect Maize Growth Promotion under Late Wilt Disease Stress.

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.

Trichoderma longibrachiatum and Trichoderma asperellum Confer Growth Promotion and Protection against Late Wilt Disease in the Field.

Late wilt disease (LWD) of maize, caused by Magnaporthiopsis maydis, is considered a major threat to commercial fields in Israel, Egypt, Spain, and India. Today's control methods include chemical and agronomical intervention but rely almost solely on resistant maize cultivars. In recent years, LWD research focused on eco-friendly biological approaches to restrain the pathogen. The current study conducted during two growing seasons explores the potential of three Trichoderma species as bioprotective treatments against LWD. These species excelled in preliminary assays performed previously under controlled conditions and were applied here in the field by directly adding them to each seed with the sowing. In the first field experiment, Trichoderma longibrachiatum successfully rescued the plants' growth indices (weight and height) compared to T. asperelloides and the non-treated control. However, it had no positive effect on yield and disease progression. In the subsequent season, this Trichoderma species was tested against T. asperellum, an endophyte isolated from susceptible maize cultivar. This experiment was conducted during a rainy autumn season, which probably led to a weak disease burst. Under these conditions, the plants in all treatment groups were vivid and had similar growth progression and yields. Nevertheless, a close symptoms inspection revealed that the T. longibrachiatum treatment resulted in a two-fold reduction in the lower stem symptoms and a 1.4-fold reduction in the cob symptoms at the end of the seasons. T. asperellum achieved 1.6- and 1.3-fold improvement in these parameters, respectively. Quantitative Real-time PCR tracking of the pathogen in the host plants' first internode supported the symptoms' evaluation, with 3.1- and 4.9-fold lower M. maydis DNA levels in the two Trichoderma treatments. In order to induce LWD under the autumn's less favorable conditions, some of the plots in each treatment were inoculated additionally, 20 days after sowing, by stabbing the lower stem section near the ground with a wooden toothpick dipped in M. maydis mycelia. This infection method overrides the Trichoderma roots protection and almost abolishes the biocontrol treatments' protective achievements. This study suggests a biological Trichoderma-based protective layer that may have significant value in mild cases of LWD.

Molecular Tracking and Remote Sensing to Evaluate New Chemical Treatments Against the Maize Late Wilt Disease Causal Agent, Magnaporthiopsis maydis.

Late wilt is a destructive disease of corn: outbreaks occur at the advanced growth stage and lead to severe dehydration of susceptible hybrids. The disease's causal agent is the fungus Magnaporthiopsis maydis, whose spread relies on infested soils, seeds, and several alternative hosts. The current study aimed at advancing our understanding of the nature of this plant disease and revealing new ways to monitor and control it. Two field experiments were conducted in a heavily infested area in northern Israel seeded with highly sensitive corn hybrid. The first experiment aimed at inspecting the Azoxystrobin (AS) fungicide applied by spraying during and after the land tillage. Unexpectedly, the disease symptoms in this field were minor and yields were high. Nevertheless, up to 100% presence of the pathogen within the plant's tissues was measured using the quantitative real-time PCR method. The highest AS concentration tested was the most effective treatment, and resulted in a 6% increase in cob yield and a 4% increase in A-class yield. In the second experiment conducted in the following summer of the same year in a nearby field, the disease outbreak was dramatically higher, with about 350 times higher levels of the pathogen DNA in the untreated plots' plants. In this field, fungicide mixtures were applied using a dripline assigned for two coupling rows. The most successful treatment was AS and the Difenoconazole mixture, in which the number of infected plants decreased by 79%, and a 116% increase in crop yield was observed, along with a 41% increase in crop quality. Evaluation of the effectiveness of the treatments on the plants' health using a remote, thermal infra-red sensitive camera supported the results and proved to be an essential research tool.