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Phoma black stem and leaf spot disease (caused by Phoma medicaginis) not only diminishes forage and seed yield but stimulates production of detrimental phytoestrogens in annual Medicago spp. This study aimed to evaluate relationships between disease development from five isolates of P. medicaginis on 16 cultivars with production of coumestrol and 4′-O-methylcoumestrol. In the presence of P. medicaginis, Sava had the highest coumestrol and 4′-O-methylcoumestrol (640 and 85 mg/kg, respectively) followed by Caliph (253 and 15 mg/kg, respectively). In the absence of P. medicaginis, Jemalong and Paragosa showed highest and lowest coumestrol (137 and 0 mg/kg, respectively). 4′-O-methylcoumestrol was not produced in disease-free plants, but coumestrol was. Disease incidence and severity on leaves and on petiole/stems, and consequent leaf yellowing severity ranged from 5%-98.7%, 0%-100%, 4.4%-98.7%, 1.7%-100%, and 0%-85%. Sava, Paraponto, Harbinger, and Serena were most susceptible, while Tornafield and Caliph were least susceptible. There was significant overall positive correlation of disease incidence/severity factors across cultivars (p < 0.01) with both coumestrol and 4′-O-methylcoumestrol. Jemalong was least responsive and Paragosa and Sava most responsive to coumestrol production following P. medicaginis inoculation. Coumestrol in inoculated Paragosa increased to 373 mg/kg in comparison with 0 mg/kg in controls. These findings are of critical importance towards developing less disease-susceptible annual Medicago spp. producing less detrimental phytoestrogens. Least susceptible cultivars like Tornafield and Caliph can be used to manage yield loss, whilst least responsive cultivars to phytoestrogen production like Caliph also can help to reduce phytoestrogen production.

The low availability of micronutrients such as zinc (Zn) and iron (Fe) and macronutrients (phosphorus (P), in particular) in calcareous soils adversely affects plants such as sorghum (Sorghum bicolor) that are highly sensitive to deficiencies of these elements. This study investigated the potential effects of different Fe chelates and plant growth-promoting rhizobacteria (PGPR) on nutrition and growth of sorghum plants growing in a low-fertility calcareous soil. A glasshouse experiment was carried out in a completely randomized design with four PGPR treatments (Pseudomonas putida P159, Pseudomonas fluorescens T17-24, Bacillus subtilis P96, and an uninoculated control), and three Fe treatments (10 mg Fe kg −1 soil as Fe-EDDHA or Fe-EDTA, and no Fe application). Application of both Fe chelates (Fe-EDDHA and Fe-EDTA) increased plant dry weight, shoot content, and concentration of Zn and Cu (copper) compared with the control treatment; however, Fe-EDTAwas found to be more effective than Fe-EDDHA. Fe-EDTA significantly increased the concentration of Cu, Zn, and Mn (manganese) up to 74.7%, 40.3%, and 30.6% in the rhizosphere, respectively. The use of PGPR significantly increased plant growth as well as the content of Mn, Zn, Cu, and K (potassium) compared with the uninoculated plants. Overall, the Fe-EDTA chelate was more effective in increasing nutrient uptake and growth of sorghum plants than Fe-EDDHA because of its low chemical stability at high pH. Combined application of Fe-EDTA and PGPR strains provided the greatest growth and nutritional benefits to sorghum plants growing in the alkaline nutrient-deficient calcareous soil.

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Phoma black stem and leaf spot disease of annual Medicago spp., caused by Phoma medicaginis, not only can devastate forage and seed yield, but also reduces herbage quality by inducing production of phytoestrogens, particularly coumestrol and 4'-O-methylcoumestrol, that can reduce ovulation rates of animals grazing infected forage. We determined the consequent phytoestrogen levels on three different annual Medicago species/cultivars (M. truncatula cv. Cyprus, M. polymorpha var. brevispina cv. Serena and M. murex cv. Zodiac), following inoculation with 35 isolates of P. medicaginis. Across the isolate x cultivar combinations, leaf disease incidence (%LDI), petiole disease incidence (%PDI), leaf disease severity (%LDS), petiole disease severity (%PDS), and leaf yellowing severity (%LYS) ranged up to 100, 89.4, 100, 58.1 and 61.2%, respectively. Cultivars Cyprus and Serena were most susceptible and cv. Zodiac the most resistant to P. medicaginis. Isolates WAC3653, WAC3658 and WAC4252 produced the most severe disease. Levels of phytoestrogens in stems ranged from 25 to 1995 mg/kg for coumestrol and from 0 to 418 mg/kg for 4'-O-methylcoumestrol. There was a significant positive relationship of disease incidence and severity parameters with both coumestrol and 4'-O-methylcoumestrol contents, as noted across individual cultivars, and as across the three cultivars overall where r=0.39 and 0.37 for coumestrol and 4'-O-methylcoumestrol, respectively (P<0.05). Although, cv. Serena was most susceptible to P. medicaginis and produced the highest levels of phytoestrogens in the presence of P. medicaginis, cv. Zodiac contained the highest levels of phytoestrogens in comparison with other cultivars in the absence of P. medicaginis. There was a 15-fold increase in coumestrol in cv. Serena but only a 7-fold increase in cv. Zodiac from infection of P. medicaginis. The study highlights that the intrinsic ability of a particular cultivar to produce phytoestrogens in the absence of the pathogen, and its comparative ability to produce phytoestrogens in the presence of the P. medicaginis, are both important and highly relevant to developing new annual Medicago spp. cultivars that offer improved disease resistance and better animal reproductive outcomes.