Leptosphaerulina Leaf Blight of Turfgrasses ... Lion or Lamb? By Steve Abler and Dr. Geunhwa Jung, Turfgrass Diagnostic Lab, Department of Plant Pathology, University of Wisconsin-Madison

eptosphaerulina spp. incite Lseveral diseases of legumes and are often found colonizing necrotic creeping bentgrass, perennial rye- grass, and Kentucky bluegrass plants Figure 1. Pseudothecia of during humid periods of the late Leptosphaerulina australis spring through fall. The fungus pro- on a necrotic creeping duces conspicuous fruiting bodies bentgrass leaf (pseudothecia) which release spores in packets referred to as asci (Figures 1 and 2). Controversy exists regarding the ecological rela- tionship between Leptosphaerulina and the turfgrass hosts. The fungus has been classified as a saprophyte (colonizes only dead tissue), senec- tophyte (colonizes dying or senes- cent tissue), weak pathogen, and pathogen of turfgrasses by various Figure 2. Spores and ascus (pI. plant pathologists. To complicate asci) of Leptosphaerulina australis matters, there have been three dif- fering classification schemes pub- lished in the literature for deter- mining the species within the genus. Because of the frequency with which Leptosphaerulina spp. are found on turfgrasses and because of the uncertainty regarding the virulence was later modified to two by Booth pieces of the fungal genome were and classification of the fungus on and Pirozynski (1967) and back to amplified and sequenced using the turfgrasses, this disease was chosen four by Irwin and Davis (1985) on polymerase chain reaction. By using for my Masters Degree project at the basis of spore morphology powerful computer software, the VirginiaTech (Abler, 2003). alone. Because the delineation of sequences of the DNA were com- Delineation of Leptosphaerulina the species was so controversial pared and the evolutionary related- species when based on spore characteris- ness of each isolate to each. other The previous three descriptions tics, more recent molecular was determined. When the analyses approaches and spore morphology for the classification of species in were completed, the molecular and were utilized for this study. the genus were based mostly on morphological data best supported the size and septation of spores. As many samples of the six species classification of The most thorough work was per- Leptosphaerulina as possible were Graham and Luttrell (1961). formed by Graham and Luttrell collected from turfgrasses and Therefore, the species of (1961), who in addition to spore other hosts (including original iso- Leptosphaerulina that occur on morphology, described ecology, lates and herbarium material from turfgrasses include L. australis host specificity, and growth char- previous taxonomic studies). (most common) and L. argenti- acteristics on artificial media of a Isolates were characterized based nensis. Once it was determined on spore morphology and placed number of isolates. They described which species ofLeptosphaerulina into categories based on the six six species of Leptosphaerulina are found on turfgrasses, experi- species key of Graham and Luttrell that occur on forage plants. The ments to determine their level of number of species in the genus (1961). DNA was extracted from virulence could be performed. the samples and two different

THE GRASS ROOTS JULY/AUGUST 2004 indistinguishable from control turfgrass diseases only appear when plants. Leaves of inoculated plants abiotic stresses predispose the were cleared and stained to verify plants to infection and colonization the presence of spores on the leaves. by fungi that are unable to harm Multiple spores were observed on healthy plants. Therefore, it is impor- each leaf, many of which germinated tant for superintendents as well as and produced germ tubes and researchers to use sound cultural appressoria (specialized structures techniques in order to maintain for penetrating the epidermal cells of healthy turf and reduce the possi- plants). These appressoria failed to bility of problems from these stress- penetrate the epidermis of the plant induced pathogens. (Figure 3). Since L. australis was References unable to infect and colonize non- Abler, S.A., 2003. Ecology and Figu;~-3.~'Germinated spore with appres- stressed and stressed turfgrasses Taxonomy of Leptosphaerulina soria {rounded tip on lowest germ tube} on under conditions favorable for dis- spp. associated with turfgrasses in a creeping bentgrass leaf ease development on legume hosts, the United States. M.s. Thesis, it was determined that VIrginiaPolytechnic Institute and Virulence of Leptosphaerulina Leptosphaerulina species are State University.Blacksburg, VA. to turfgrasses saprophytes of necrotic turf- Booth, C. and Pirozynski, K.A. 1967. To test the virulence of grasses. The practical implication Leptosphaerulina trifolii. C.M.I. Leptosphaerulina to turfgrasses, of these results is that when descriptions of pathogenic fungi healthy and stressed perennial rye- Leptosphaerulina is found on and bacteria 146. grass (cultivars Palmer III and Fiesta dead turfgrasses, the turfgrass Graham J.H., and Luttrell, E.s. 1961. II) and creeping bentgrass (cultivar manager must determine the pri- Species of Leptosphaerulina on Crenshaw) were inoculated with six mary cause of the declining turfgrass forage plants. Phytopathology 51: turfgrass isolates of L. australis. In which could be a multitude of biotic 680-693. order to stress the plants and induce and/or abiotic factors. Determining Irwin, J.A.G., and Davis, R.D. 1985. senescence, perennial ryegrass this underlying cause of the problem Taxonomy of some plants and creeping bentgrass plants may be complicated by the ability of Leptosphaerulina species on were incubated at 38°C (100°F) with Leptosphaerulina spp. to rapidly legumes in eastern Australia. 100% relative humidity in the dark colonize the moribund turfgrasses. Australian Journal of Botany 33: for 48 and 24 hours respectively. Although Leptosphaerulina leaf 233-237.* These plants were then placed back blight is no longer even considered a in the greenhouse for three days pathogen of stressed grasses, several before inoculation. The senescent status of the leaves was measured and verified by the significant reduc- tion in the chlorophyll content of the Rettler Corporation leaves at time of inoculation. After inoculation, both healthy (unstressed) plants and stressed plants were incubated under humidity domes for three days at 22°C (72°F), the optimum tempera- ture for infection and colonization of legume hosts by Leptosphaerulina spp. The plants were inspected for symptoms daily for eight days fol- lowing the inoculation. None of the inoculated plants showed any noticeable symptoms of disease at any time during six inocu- lation experiments. Even the stressed plants which showed mean Middleton. WI Phone: 60g-833~S400 chlorophyll losses of 30-45% were Stevens Point, WI Phone: 715-341-2633 .. WebSite: www.reUler.com

THE GRASS ROOTS JULY/AUGUST 2004