This article is from the October 2014 issue of published by The American Phytopathological Society For more information on this and other topics related to plant pathology, we invite you to visit the APS website at www.apsnet.org Ecology and Epidemiology e-Xtra* A Tripartite Interaction Between Spartina alterniflora, Fusarium palustre, and the Purple Marsh Crab (Sesarma reticulatum) Contributes to Sudden Vegetation Dieback of Salt Marshes in New England Wade H. Elmer The Department of Plant Pathology and Ecology, The Connecticut Agricultural Experiment Station, P.O. Box 1106, New Haven 06504. Accepted for publication 17 March 2014. ABSTRACT Elmer, W. H. 2014. A tripartite interaction between Spartina alterniflora, plants. When the drought treatment was combined with inoculation with Fusarium palustre, and the purple marsh crab (Sesarma reticulatum) F. palustre, plants were significantly more stunted and symptomatic, had contributes to sudden vegetation dieback of salt marshes in New England. less fresh weight, more diseased roots, and a greater number of Fusarium Phytopathology 104:1070-1077. colonies growing from the roots (P < 0.001) than noninoculated plants. The effects were additive, and statistical interactions were not detected Tripartite interactions are common and occur when one agent (an between drought and inoculation. Estimates of herbivory (number of arthropod or pathogen) changes the host plant in a manner that alters the grass blades cut or biomass consumption) by the purple marsh crab were attack of the challenging agent. We examined herbivory from the purple significantly greater on drought-stressed, diseased plants than on healthy marsh crab (Sesarma reticulatum) on Spartina alterniflora following plants irrigated normally. Drought increased attraction to the purple exposure to drought or inoculation with Fusarium palustre in mecocosms marsh crab more than inoculation with F. palustre. However, when only in the greenhouse and in crab-infested creek banks along intertidal salt mild drought conditions were imposed, plant consumption was greater on marshes. Initially, drought stress on S. alterniflora and disease from F. inoculated plants. Healthy, nonstressed transplants set into plots in crab- palustre were examined in the greenhouse. Then, a second challenger, the infested intertidal creek banks were grazed less each year than inoculated purple marsh crab, was introduced to determine how drought and disease plants or plants that were exposed to drought. Several hypotheses relating from F. palustre affected the attraction and consumption of S. alterniflora. to nutrition, chemotaxis, and visual attraction are presented to explain Plant height and shoot and root weights were reduced in plants subjected how stress from drought or disease might favor herbivory. to severe drought treatment when compared with normally irrigated A tripartite interaction occurs when arthropod infestation or Louisiana discovered another pathogenic species called Fusarium pathogen infection changes the host plant in a manner that alters palustre (W. H. Elmer and R. E. Marra) (16). Fungal incidence the attack of the challenging agent (39). Both positive and nega- was higher in plants in SVD sites than in healthy marshes (16% in tive interactions occur, with one agent increasing or decreasing SVD sites and <1% in healthy sites) (15). Root-knot nematodes susceptibility to a second attacker. The role of environment can were also discovered but their role in SVD remains elusive (31). also complicate tripartite interactions and can dramatically shift F. palustre was not implicated as a primary agent of SVD because the ecological costs of the host plant (25). Tripartite interactions pathogenicity tests on healthy plants rarely resulted in mortality. are well documented for many insect–disease–plant systems (4), However, the close association between fungus and host in SVD where the ecological cost to the insect or pathogen is enhanced or sites sparked the hypothesis that, as an endophyte, F. p a l u st re reduced (6,7,19,22,39). Multilayer interactions in natural eco- might function as a pathogen when its host was compromised by systems can collectively affect the structure and composition of stress. Most marsh systems flood daily but tides can occasionally natural plant communities; however, experimentally demon- recede for extended periods of time, causing soil to be aerobic strating the individual contribution of each agent is difficult. and acidic. However, extended drought events, as reported by In the late 1990s, sudden vegetation dieback (SVD) of smooth Palmer Drought Indices, in the northern marshes have not been cordgrass (Spartina alterniflora Loisel.) occurred in salt marshes observed. Short-term droughts (less than 1 week) do occur but on the Atlantic and Gulf Coasts, leaving only remnant peat (2,5, their effect on SVD is not clear. Scientists in Louisiana observed 18,33,38). The patterns associated with SVD vary from marsh to that plants inoculated with species in the G. fujikuroi species marsh and from region to region but usually begin as specific foci complex were significantly more diseased when plants were of dieback that radiate outward and coalesce to create large, drought stressed (42). The relationship between F. palustre and irregular areas within the marsh system, leading scientists to drought was not known. suspect a contagious agent. Surveys in Louisiana found that Another interacting stressor found in SVD sites in New England species in the Gibberella fujikuroi species complex (34) were able are high densities of the herbivorous purple marsh crab (Sesarma to infect S. alterniflora (18,42). Regional surveys from Maine to reticulatum Say) (27) (Fig. 1), and considerable loss of marsh biomass can be attributed to this invertebrate. The degree of herbi- vory in these marshes is variable and certain marshes are grazed Corresponding author: W. H. Elmer; E-mail address: [email protected] more heavily than others. Although release from predation by * The e-Xtra logo stands for “electronic extra” and indicates that Figure 1 appears blue crabs, fin fish, and birds can influence crab densities (3), in color online. creek banks previously denuded by SVD may also provide a better burrowing habitat for marsh crabs than more vegetated http://dx.doi.org/10.1094/PHYTO-08-13-0219-R creek banks. Second, Spartina alterniflora plants on the perimeter © 2014 The American Phytopathological Society 1070 PHYTOPATHOLOGY of SVD sites are commonly more stressed and stunted than plants Four isolates of F. palustre (NRRL 54065, NRRL 54059, more distal to the SVD site. Marsh crabs may preferentially graze NRRL 54054, and NRRL 54056) (16) were grown on 25% potato on stressed plants in SVD sites because of their proximity or dextrose agar (PDA) plates for 7 to 10 days at 20 to 25°C under because of improved palatability. It remained to be demonstrated light for 12-h photoperiods. These isolates were from different whether or not drought stress or colonization by endophytic F. marshes along the Long Island Sound and had been shown in palustre would influence herbivory by purple marsh crab. previous tests to be pathogenic (15,16). Inoculum was prepared The aim of this study was to first examine interactions between two ways, depending on the experiment. In the first set of green- pairwise treatments (drought stress on S. alterniflora and disease house experiments, each isolate was seeded onto twice-autoclaved from F. palustre), then introduce a third challenger to the inter- millet, allowed to grow for 2 weeks, then air dried, ground, and action to determine how drought and inoculation with F. palustre passed through a 0.5-mm sieve. Equal amounts of each inoculum affect attraction and consumption of S. alterniflora by the purple were bulked together and mixed into soil (1:1 peat/sand) at 2 g of marsh crab. millet inoculum per liter of soil. In later experiments on herbi- vory, we wanted the above- and belowground tissue to be exposed MATERIALS AND METHODS to F. palustre; therefore, conidial spray or drenches were used. For conidial inoculations, colonized 25% PDA plates were washed with Production of plants and inoculum. Flower spikes of S. distilled water and spores enumerated to 5 × 104 macroconidia/ alterniflora were collected in fall 2009 from a salt marsh in ml. Using a spray bottle, 30 ml were applied to each plant. Madison, CT where SVD was absent. Seeds were surface dis- Greenhouse drought and disease experiments. To study the infested in 10% bleach (0.53% Na hypochlorite) for 1 min, rinsed effect between drought and inoculation with F. palustre, green- in tap water, spread into the top centimeter of sand in trays, and house experiments were conducted with S. alterniflora trans- set on greenhouse benches. Individual seedlings were transplanted plants. There were six treatments: severe drought-not inoculated, into 36-cell inserts containing a 1:1 mix of sand and peat that was mild drought-not inoculated, normal irrigation-not inoculated, limed to achieve a pH of 6.5 to 7.0. Seedlings were held for 6 to 9 severe drought-inoculated, mild drought-inoculated, and normal months and fertilized with 20-20-20 NPK twice a month. Pre- irrigation-inoculated. Each treatment was replicated 10 times (one liminary studies have shown that S. alterniflora plants were larger plant per replicate) and the experiment was repeated two times. and healthier when exposed to a 50% marsh water irrigation Plants were planted into soil infested with the millet inoculum (approximately 0.14 M NaCl) than continuous tap water; there- and subjected to a severe drought that was imposed by with- fore, plants were exposed to 50% autoclaved marsh water every holding water until at least half of the leaf blades began to curl third or fourth irrigation to help maintain osmoregulation. For inward. Plants were irrigated with tap water until leaf blades each experiment, plants of uniform size were selected and arbi- become turgid, which occurred within 3 days in the summer and trarily assigned to treatment.