Interaction of rhizobacteria with leafy spurge (Euphorbia esula L.) callus tissue cells

The interaction of two rhizobacterial isolates,Pseudomonas fluorescens isolate LS 102 andFlavobacterium balustinum isolate LS 105, with leafy spurge cells at the cellular level was studied using scanning and electron microscopy. Leafy spurge callus tissue inoculated with either isolate showed considerable changes compared to non-inoculated tissue. The attachment of rhizobacteria to cell surfaces was associated with the elaboration of fibrillar material which may anchor bacteria to surfaces and contribute to mediation of the phytotoxic effect caused by rhizobacteria. At the ultracellular level, inoculated callus tissue showed numerous cell alterations including esiculation and convolution of the plasmalemma, cell wall degradation and disorganization of the cytoplasm, similar to those detected in the whole plant. It is concluded that callus tissue may provide an excellent working model to investigate the mode and/or mechanism of action of potential biocontrol agents on their host plants.     

Phytotoxic Allelochemicals From Roots and Root Exudates of Leafy Spurge (Euphorbia esula L.)

Invasive plants are a widespread problem but the mechanisms used by these plants to become invasive are often unknown. The production of phytotoxic natural products by invasive weeds is one mechanism by which these species may become successful competitors. Here we conducted a bioactivity-driven fractionation of root extracts and exudates from the invasive plant leafy spurge (Euphorbia esula L.), and structurally characterized jatrophane diterpenes and ellagic acid derivatives. Ellagic acid derivatives and one of the jatrophane diterpenes, esulone A, have been previously reported from leafy spurge, but another of the jatrophane diterpenes, kasuinine B, has not. We show that these compounds are phytotoxic but affect plants in different ways, either inducing overall plant necrosis or reducing root branching and elongation.Key Words: phytotoxicity, allelochemicals, roots, root exudates, jatrophane diterpenes, kansuinine B, ellagic acid derivatives, leafy spurge, Euphorbia esula, Arabidopsis thaliana     

Hierarchical Bayesian methods estimate invasive weed impacts at pertinent spatial scales

Invasive weed impact estimates are needed to determine whether or not weeds warrant costly control measures. Typically, land managers seek local weed impact estimates (e.g. ranches, parks) and policy-makers want to know how weeds are impacting entire regions. Our goal was to provide local and regional impact estimates for a ubiquitous invasive weed: leafy spurge (Euphorbia esula L.). The specific impacts we looked at related to desired species biomass production, livestock carrying capacities, and grazing land values. Our basic approach was to use an empirical model that characterizes weed biomass across the landscape in combination with another empirical model that predicts weed impact from weed biomass. Our investigation revealed that, without on-site plant biomass data, site-specific leafy spurge impacts are highly uncertain. Supplementing our general predictive model with small quantities of on-site data increased precision considerably. For the 17-state region we considered, 95% Bayesian credibility intervals indicated leafy spurge reduces cattle carrying capacities by 50–217 thousand animals a year and reduces grazing land values by 8–34 million dollars a year. Additional plant biomass data from randomly selected, leafy spurge-infested sites would shrink these fairly wide intervals.     

An efficient method for <Emphasis Type="Italic">in vitro</Emphasis> regeneration of leafy spurge (<Emphasis Type="Italic">Euphorbia esula</Emphasis> L.)

Leafy spurge (Euphorbia esula L.) is a perennial, invasive weed used as a model to study invasive plant behavior, because molecular tools (such as a deep expressed sequence tag database and deoxyribonucleic acid microarrays) have been developed for this species. However, the lack of effective in vitro regeneration and genetic transformation systems has hampered molecular approaches to study leafy spurge. In this study, we describe an efficient in vitro regeneration system. Three highly regenerative lines were selected by screening the in vitro regeneration capabilities of stem explants of 162 seedlings. The effects of various culture conditions on in vitro regeneration were then evaluated based on explant competence to form calluses and shoots. High rates of shoot regeneration can be obtained using a growth medium containing 1× woody plant basal medium and 1× Murashige and Skoog (MS) basal salts, 1× MS vitamins, 1.11 μM 6-benzylaminopurine, 1.97 μM indole-3-butyric acid, and 3% sucrose, pH 5.6–5.8. After 30 d culture, multiple shoots formed either directly from the stem or indirectly from the callus. This method is a requisite for the development of genetic transformation systems for leafy spurge and may be used to develop in vitro regeneration techniques for other species in the Euphorbiaceae.     

Physiological responses of resistant and susceptible plants to diclofop-methyl and its antagonism by 2,4-D and antioxidants

Diclofop-methyl (DM) sprayed onto 6–8-week-old plants of leafy spurge ( Euphorbia esula L.) caused senescence and abscission of older leaves, while the young leaves and apex remained attached. The phytotoxicity of DM was reversed by the antioxidant, α -tocopherol (vitamin E), in leafy spurge and DM-susceptible oat ( Avena sativa L. cv. Gary). DM and 2,4-dichlorophenoxyacetic acid (2,4-D) increased ethylene evolution in mature leaves of leafy spurge. Vitamin E reduced the DM-induced ethylene by ampproximately 50%, but had no effect on the 2,4-D-induced ethylene. DM did not increase ethylene in DM-resistant pea or tobacco, but 2,4-D induced a 3-fold increase in ethylene evolution over controls in DM-resistant tobacco. 2,4-D amppears to act at a site different from that of DM in the pathway of ethylene formation. Ethylene evolution increased in DM-treated susceptible biotypes of annual ryegrass ( Lolium rigidum L.) and wild oat ( Avena fatua L.), but not in resistant biotypes of these species. DM reduced root and shoot formation and dry weight in hypocotyl segments of etiolated leafy spurge seedlings grown in vitro. Organogenesis and dry weights were increased by the combination of DM+antioxidants. Vitamin E was a more effective antioxidant than ascorbic acid. These results sumpport the hypothesis that DM induces oxidative stress in susceptible plant tissues and that antioxidants reduce the damaging action of the phytotoxic free radicals.     

Biology and host specificity ofChamaesphecia hungarica andCh. Astatiformis (Lep.: Sesiidae) two candidates for the biological control of leafy spurge,Euphorbia esula (Euphorbiaceae) in North America

Leafy spurge (Euphorbia esula (s.1.)) is an herbaceous perennial and serious weed of Eurasian origin that has been accidentally introduced into North America. The two European root-boring mothsChamaesphecia hungarica andCh. astatiformis are univoltine and overwinter as mature larvae. Both species have a lower survival rate on leafy spurge than on their field hosts, and thus are not optimal candidates for the biological control of leafy spurge. However, the rate of larval development and larval growth on the target weed and on the two field hosts is nearly the same. The experimental host range of both species is restricted to a few species in the subgenusEsula within the genusEuphorbia. The two species occupy different habitats in the steppe biome and are targeted for similar leafy spurge habitats in North America.     

Energy potential of leafy spurge (Euphorbia esula)

Leafy spurge (Euphorbia esula) is a noxious, perennial weed that infests pastures, rangeland and waste areas in the northern Great Plains. The objective of this study was to determine the productive potential of this species when grown under optimum agronomic conditions. Plants were fertilized and irrigated. Oil, hydrocarbon, total protein, and dry-weight production were measured on 3 harvest dates. Calorimetric analyses were performed to determine the potential of leafy spurge as a fuel crop. The hydrocarbon content of 12 strains of leafy spurge was determined to measure genetic variability for this trait. The addition of fertilizer doubled dryweight production but did not affect percent oil or hydrocarbon content. Oil and hydrocarbon production averaged 6.8 and 0.6% on a plant dry-weight basis. Maximum production of plant biomass, protein, and hydrocarbon was obtained from a mid-July harvest. Oil content increased later in the growing season. The total protein content of leafy spurge averaged 12%. Whole-plant biomass had a caloric value of 4,407 cal/g while the oils contained 10,019 cal/g. Leafy spurge hay can produce 4 times more energy per year than wheat straw; therefore, the immediate potential of leafy spurge whole-plant biomass as a locally grown fuel crop for home-heating purposes is suggested.     

Effect of Leafy Spurge (Euphorbia esula) Genotype on Feeding Damage and Reproduction of Aphthona spp.: Implications for Biological Weed Control

North American leafy spurge (Euphorbia esula L.) is genetically diverse and composed of multiple genotypes introduced from several areas of Europe and Asia. Five species of leafy spurge flea beetle (Aphthona spp.) have been introduced as biological control agents for leafy spurge, but were collected in a relatively small region of Europe. Greenhouse and field experiments were conducted to determine if observed variation in feeding preference and reproduction of Aphthona spp. on North American leafy spurge may be due in part to leafy spurge genotype. Leafy spurge genotypes were collected from Austria; Manitoba in Canada; and the states of Montana, Nebraska, North Dakota, South Dakota, and Wyoming in the United States. Leafy spurge genotype affected feeding but not egg laying by Aphthona spp. adults. Aphthona czwalinae/lacertosa fed slightly less in a free-choice test on a genotype from Manitoba (7%) compared to genotypes from Nebraska and North Dakota (14%). Aphthona flava tended to feed less on the Nebraska genotype than any other genotype evaluated. Reproduction of Aphthona spp. was greatly affected by leafy spurge genotype. For instance, A. czwalinae/lacertosa produced 72 adults per plant from a Nebraska genotype compared to 11 to 32 adults per plant from all other genotypes evaluated. Thus, some observed variation in establishment and reproduction of Aphthona spp. can be attributed to leafy spurge genotype.     

Identification, Pathogenicity and Comparative Virulence of Fusarium spp. Associated with Diseased Euphorbia spp. in Europe

Fusarium spp. isolated from diseased Euphorbia spp. in Europe were assessed for pathogenicity to North American accessions of leafy spurge ( Euphorbia esula/virgata ). Of the nine strains of Fusarium spp. isolated from diseased E. stepposa or E. virgata in the Caucasus region of Russia and E. esula/virgata in southern France, all were pathogenic to leafy spurge. There were significant differences in virulence among strains. Four strains, including the two that were most virulent, were identified as F. oxysporum . Four of the five other strains were identified as F. solani and one was identified as F. proliferatum . Three of the four most virulent strains to leafy spurge were isolated from E. stepposa . The most virulent strain was associated with root damage caused by insect biological control agents, as found earlier with domestic strains of Fusarium spp. pathogenic to leafy spurge. Two strains identified as F. solani were vegetatively compatible. It was concluded that further screening of a larger set of strains of foreign Fusarium spp. under quarantine conditions in the US or in limited overseas facilities would be justified, and could yield promising biological control agents for leafy spurge.     

Modeling Responses of Leafy Spurge Dispersal to Control Strategies

Leafy spurge （Euphorbia esula L.） has substantial negative effects on grassland biodiversity, productivity, and economic benefit in North America. To predict these negative impacts, we need an appropriate plant-spread model which can simulate the response of an invading population to different control strategies. In this study, using a stochastic map lattice approach we generated a spatially explicitly stochastic process-based model to simulate dispersal trajectories of leafy spurge under various control scenarios. The model integrated dispersal curve, propagule pressure, and population growth of leafy spurge at local and short-temporal scales to capture spread features of leafy spurge at large spatial and long-temporal scales. Our results suggested that narrow-, medium-, and fat-tailed kernels did not differ in their ability to predict spread, in contrast to previous works. For all kernels, Allee effects were significantly present and could explain the lag phase （three decades） before leafy spurge spread accelerated. When simulating from the initial stage of introduction, Allee effects were critical in predicting spread rate of leafy spurge, because the prediction could be seriously affected by the low density period of leafy spurge community. No Allee effects models were not able to simulate spread rate well in this circumstance. When applying control strategies to the current distribution, Allee effects could stop the spread of leafy spurge; no Allee effects models, however, were able to slow but not stop the spread. The presence of Allee effects had significant ramifications on the efficiencies of control strategies. For both Allee and no Allee effects models, the later that control strategies were implemented, the more effort had to be input to achieve similar control results.     

Biology and host specificity of Spurgia capitigena (Bremi) (Dipt., Cecidomyiidae), for the biological control of Euphorbia esula L. in North America

Abstract: Spurgia capitigena (Bremi) was evaluated as a potential biological control agent of leafy spurge, Euphorbia esula L. (species complex) in North America. To ascertain the host specificity of this gall midge, tests were conducted in the field at Cavaillon, France, and at laboratories located in Rome, Italy and Bozeman, Montana, USA, in 1994 and 1995. Twenty-nine plant species, including eight native North American euphorbias and biotypes of E . esula , were evaluated. In the laboratory, the gall-midge was able to induce galls on seven species of spurges (all belonging to the subgenus Esula) and the three accessions of leafy spurge. In field tests, S . capitigena infested only E . esula . The biology and host specificity of S . capitigena from France appears to be similar to that observed for Spurgia esulae Gagné from Italy (= Bayeria capitigena Bremi). In France, this gall-midge was commonly observed from early May until October in habitats of very heavy and moist soils located along water channels, roadsides and in fruit orchards. Galls were induced on meristematic tissues, and thereby prevented flowering.     

Attack behavior and host utilization ofAphthona chinchihi (Col.: Chrysomelidae), a potential biological control agent ofEuphorbia esula (Euphorbiaceae, leafy spurge) in North America

Aphthona chinchihi Chen was collected in China feeding on leafy spurge (Euphorbia esula L.). Studies were conducted on its host specificity in the laboratory, using field collected adults and their progeny.Aphthona chinchihi can effectively complement the impact of the other natural enemies of leafy spurge established from Europe in the U.S.A. and Canada. The adults feed on leaves and shoots and the larvae, which cause the main damage to the plant, feed on the hypogeous portion of the plant, seriously stressing the plant and preventing its vegetative spread. The host range ofA. chinchihi was studied with tests on adult feeding and oviposition, larval survival and host suitability, using 40 plant species or varieties distributed in 12 families. The experiments demonstrated that it has a high level of specificity. This flea beetle completed its life cycle only on leafy spurge. Also, because of its ecological valence,A. chinchihi has a very good potential as a biocontrol agent in North America.     

Do invasive plants structure microbial communities to accelerate decomposition in intermountain grasslands?

Invasive plants are often associated with greater productivity and soil nutrient availabilities, but whether invasive plants with dissimilar traits change decomposer communities and decomposition rates in consistent ways is little known. We compared decomposition rates and the fungal and bacterial communities associated with the litter of three problematic invaders in intermountain grasslands; cheatgrass (Bromus tectorum), spotted knapweed (Centaurea stoebe) and leafy spurge (Euphorbia esula), as well as the native bluebunch wheatgrass (Pseudoroegneria spicata). Shoot and root litter from each plant was placed in cheatgrass, spotted knapweed, and leafy spurge invasions as well as remnant native communities in a fully reciprocal design for 6 months to see whether decomposer communities were species‐specific, and whether litter decomposed fastest when placed in a community composed of its own species (referred to hereafter as home‐field advantage–HFA). Overall, litter from the two invasive forbs, spotted knapweed and leafy spurge, decomposed faster than the native and invasive grasses, regardless of the plant community of incubation. Thus, we found no evidence of HFA. T‐RFLP profiles indicated that both fungal and bacterial communities differed between roots and shoots and among plant species, and that fungal communities also differed among plant community types. Synthesis. These results show that litter from three common invaders to intermountain grasslands decomposes at different rates and cultures microbial communities that are species‐specific, widespread, and persistent through the dramatic shifts in plant communities associated with invasions.     

Integration of biological control agents with other weed management technologies: Successes from the leafy spurge (Euphorbia esula) IPM program

An invasive weed can occupy a variety of environments and ecological niches and generally no single control method can be used across all areas the weed is found. Biological control agents integrated with other methods can increase and/or improve site-specific weed control, but such combinatorial approaches have not been widely utilized. The successful leafy spurge (Euphorbia esula L.) control program provides examples for future integrated weed programs that utilize biological control agents with traditional methods. Weed control methods can be used separately, such as when the leafy spurge gall midge (Spurgia esulae Gagné) reduced seed production in wooded areas while herbicides prevented further spread outside the tree line. Traditional methods also can be used directly with biological control agents. Incorporation of Aphthona spp. with herbicides has resulted in more rapid and complete leafy spurge control than either method used alone. Also, the insect population often increased rapidly following herbicide treatment, especially in areas where Aphthona spp. were established for several years but had been ineffective. Incorporation of Aphthona spp. with sheep or goat grazing has resulted in a larger decline in leafy spurge production than insects alone and in weed density than grazing alone. Controlled burns can aid establishment of biological control agents in marginally suitable environments, but timing of the fire must be coordinated to the insect’s life-cycle to ensure survival. Integration of biological control agents with revegetation programs required the agent to be the last method introduced because the cultivation and herbicide treatments necessary to establish desirable grasses and forbs were destructive to the insect. In a practical application, herbicides were combined with Aphthona spp. to help the insect establish and control leafy spurge in the habitat of the western prairie fringed orchid (Platanthera praeclara Sheviak and Bowles), an endangered species. Several experimental designs can be used to evaluate biological control agents with cultural, mechanical, and chemical control methods or with additional biological agents.     

Temporal dynamics of leafy spurge (Euphorbia esula) and two species of flea beetles (Aphthona spp.) used as biological control agents

The goal of this study was to evaluate the biological control program of leafy spurge (Euphorbia esula) in a large natural area, Theodore Roosevelt National Park, western North Dakota, USA. Aphthona lacertosa and Aphthona nigriscutis have been released at more than 1800 points in the 18,600-ha South Unit of the park beginning in 1989; most releases have occurred since 1994. We established permanent vegetation plots throughout the infested area of the park and determined stem counts and biomass of leafy spurge and abundance of the two flea beetle species at these plots each year from 1999 to 2001. Both biomass and stem counts declined over the 3 years of the study. Both species of flea beetle are well established within the park and have expanded into areas where they were not released. A. nigriscutis was more abundant than A. lacertosa in the grassland areas we surveyed, but in all other habitats abundances were similar. Using structural equation models, only A. lacertosa could be shown to have a significant effect on counts of mature stems of leafy spurge. A. nigriscutis numbers were positively correlated with stem counts of mature stems. Previous year’s stem counts had the greatest influence on change in stem counts over each 2-year time step examined with structural equation models.     

Nature,nurture, and vegetation management: Studies with sheep and goats

Diet selection and preference by grazing animals are determined by genetic and environmental factors (i.e., nature and nurture) that interact and affect their efficacy for managing vegetation as targeted grazers. The effect of rearing environment on the consumption of leafy spurge by sheep and goats was investigated. We hypothesized that although rearing environment will affect the preference for chemically defended plants ultimately, the inherent ability to detoxify or eliminate phytotoxins will limit an animal’s preference for them. The objective of this study was to determine if sheep would consume more of the invasive weed leafy spurge (Euphorbia esula) if they were raised by goat compared to sheep raised by sheep and goat raised by goat. Sheep were raised on leafy spurge-infested pastures by either their ewe (S) or a goat doe (FS) on which they were fostered within 24 hours of birth and parturition of lamb and doe, respectively. Does that fostered lambs also raised their own goat offspring (G) such that the same doe raised the FS and G animals. The rearing environment’s effect on leafy spurge consumption was tested the following growing season by simultaneously grazing all animals on the same leafy spurge-infested rangeland and estimating percentage leafy spurge in their diet with either fecal near-infrared spectroscopy (f.NIR) or bite count. Goats consumed more leafy spurge as determined by either f.NIR (62.8%, P < 0.06) or bite count (71.9%, P < 0.01) than FS (35.2 % f.NIR, 39.3% bite count) or S (10.1 % f.NIR, 18.2% bite count). The FS consumed over twice as much leafy spurge as S and were numerically intermediate to G and S for leafy spurge consumption but not significantly different from the S sheep, most likely because one FS sheep did not eat leafy spurge during the evaluation period. Because leafy spurge is aversive to sheep but not goats, higher leafy spurge consumption by FS sheep is hypothesized to result from inoculation of their rumen microbes with microbes from the does capable of denaturing aversive phytotoxins in leafy spurge. The higher consumption of leafy spurge by G compared to FS shows that genetically determined physiological differences influence an animal’s ability to ameliorate phytotoxins and determine the upper limit of an animal’s preference for a chemically defended plant. It also indicated that in addition to the animal’s genome, the genome of an animal’s microbiome, which the mother may influence, can play an important role in diet selection.     

Seasonal variation in nitrogen storage reserves in the roots of leafy spurge (Euphorbia esula) and responses to decapitation and defoliation

Seasonal fluctuations of carbohydrates and nitrogenous components in the roots of the noxious perennial leafy spurge ( Euphorbia esula L.) are strongly associated with overwintering strategy, Amino acids and distinct soluble proteins accumulate during fall and remain at elevated levels throughout winter. The formation of carbohydrate reserves in roots was not significantly affected by decapitation or selective defoliation; however, maximum amino acid and soluble protein contents were markedly reduced. In particular, the accumulation pattern of a 26 kDa protein was altered. This protein may play a role in plant conditioning and regenerative potential.     

Cyanide Production by Rhizobacteria and Potential for Suppression of Weed Seedling Growth

Rhizobacteria strains were characterized for ability to synthesize hydrogen cyanide and for effects on seedling root growth of various plants. Approximately 32% of bacteria from a collection of over 2000 isolates were cyanogenic, evolving HCN from trace concentrations to >30 nmoles/mg cellular protein. Cyanogenesis was predominantly associated with pseudomonads and was enhanced when glycine was provided in the culture medium. Concentrations of HCN produced by rhizobacteria were similar to exogenous concentrations inhibiting seedling growth in bioassays, suggesting that cyanogenesis by rhizobacteria in the rhizosphere can adversely affect plant growth. Growth inhibition of lettuce and barnyardgrass by volatile metabolites of the cyanogenic rhizobacteria confirmed that HCN was the major inhibitory compound produced. Our results suggest that HCN produced in the rhizospheres of seedlings by selected rhizobacteria is a potential and environmentally compatible mechanism for biological control of weeds. Received: 13 December 2000/Accepted: 6 February 2001