Phoma medicaginis Isolate Differences Determine Disease Severity and Phytoestrogen Production in Annual Medicago spp.

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.

Co-inoculation of lucerne (Medicago sativa) with an AM fungus and a rhizobium reduces occurrence of spring black stem and leaf spot caused by Phoma medicaginis

Spring black stem and leaf spot of lucerne (alfalfa, Medicago sativa L.), caused by Phoma medicaginis, is an important disease in temperate regions of the world. It is now a serious disease threatening global lucerne production. This experiment was designed to test the combined effects of the arbuscular mycorrhizal (AM) fungus Funneliformis mosseae and the rhizobium Sinorhizobium medicae on growth, nutrient uptake and disease severity in lucerne. The results showed that F. mosseae increased plant phosphorus and nitrogen uptake and plant dry weight, and this beneficial effect was enhanced when in association with S. medicae. Rhizobial and AM fungal effects were mutually promoting; inoculation with AM fungus significantly increased the formation of root nodules, and inoculation with rhizobium increased the percentage of root length colonised by AM fungus (P < 0.05). After infection with P. medicaginis, typical leaf spot symptoms with the lowest disease incidence and disease index occurred on plants that were host to both F. mosseae and S. medicae. Plants with both symbiotic microorganisms had higher activities (concentrations) of phenylalanine ammonia-lyase, chitinase, β-1,3-glucanase, lignin, hydroxyproline-rich glycoprotein and jasmonic acid. Therefore, the tested AM fungus (F. mosseae) and rhizobium (S. medicae) have the potential to reduce damage and yield loss in lucerne from spring black stem and leaf spot caused by P. medicaginis.

Fungi isolated from phyllosphere of fodder galega (Galega orientalis)

The object of the experiment was fodder galega (<em>Galega orientalis</em> Lam.) cultivated in 2001-2003 as field crop on three plots: 1. without fertilization, 2. 40 kg P₂O₅ × ha^-1 and 80 kg K₂O × ha^-1, 3. 80 kg P₂O₅ × ha^-1 and 160 kg K₂O × ha^-1. During the dry and warm vegetation season of 2002 almost two times fewer isolates were obtained from the leaves than in 2003 that was the most abundant in fungi. Yeasts-like fungi (30% of the total number of isolates) and saprotrophic fungi with dominated species: <em>Acremonium strictum</em> (8.5%), genus <em>Epicoccum</em> (7.8%), <em>Humicola</em> (9.5%) and <em>Penicillium</em> (18.9%) were the fungi most frequently populating the leaves of galega. The share of pathogens in the total number of isolates obtained from the phyllosphere was 10.6%. They were represented by fungi of <em>Ascochyta</em> spp., <em>Botrytis cinerea</em>, genus <em>Fusarium, Phoma medicaginis</em> and <em>Sclerotinia sclerotiorum</em>. Reduction by 1.9 to 4.6% in the number of fungi isolated from the phyllosphere of galega without fertilization as compared to galega cultivated in combinations with fertilization was recorded. Generally, the smallest number of pathogens was recovered from galega fertilized with 40 kg P₂O₅ × ha^-1 and 80 kg K₂O × ha^-1. <em>B. cinerea</em> most frequently populated galega in combination without fertilization, genus <em>Fusarium fungi</em> in combination without fertilization and with fertilization with 80 kg P₂O₅ × ha^-1 and 160 kg K₂O × ha^-1, while <em>Ascochyta</em> spp. were isolated from galega with fertilization only.

Occurrence of pathotypes of Phoma medicaginis var. pinodella (Jones) Boerema in the Olsztyn district

The causal agents of leaves and pods spot-pot of peas and field peas in the Olsztyn district was the fungus <i>Phoma medicaginis</i> var. <i>pinodella</i>. Investigated isolates of <i>P. medicaginis</i> var. <i>pinodella</i> differed by macroscopic and microscopic features. From diseases leaves and pods of peas and field peas obtained also saprophytic fungi among which <i>Alternaria alternata, Botrytis cinerea, Cladosporium cladosporioides, Epicoccum purpurascens, Stemphylium botryosum, Sordaria fimicola</i> and <i>mycelia sterilia</i> were dominated. In the study on the identification and distribution of pathotypes of <i>P. medicaginis</i> var. <i>pinodella</i> six pathotypes were identified among which pathotype 3 were dominated. The identified pathotypes differed by macroscopic and microscopic features.