Remediation of soil polluted with HMW-PAHs by alfalfa or brome in combination with fungi and starch.

High-molecular weight polycyclic aromatic hydrocarbons (HMW-PAHs) are common pollutants in soil of coal mining areas that affect the safety of crops and the environment. In a pot experiment, we compared the remediation potential of alfalfa (Medicago sativa Linn) and brome (Bromus inermis Leyss.) either alone or in combination with starch or Fusarium sp. strain ZH-H2 for a farmland soil contaminated with 4-6-ring PAHs from a coal mine area. The alfalfa and brome alone treatments reduced the concentrations of most HMW-PAHs. However, when starch was added, the removal rates of indeno(1,2,3-cd)pyrene and benzo(ghi)perylene were significantly higher for brome than for alfalfa. When ZH-H2 was combined with brome, benzo(k)fluoranthene, benzo(a)pyrene, indeno(1,2,3-cd)pyrene, and benzo(ghi)perylene degradation rates were significantly enhanced compared with brome alone. In contrast, an antagonistic effect was observed between alfalfa and Fusarium. The brome, starch and ZH-H2 combination resulted in far better removal rates than the alfalfa combination. Maximum removal rates were obtained with the brome + starch + ZH-H2 combination for benzo(k)fluoranthene (42.64%), benzo(a)pyrene (51.01%), indeno(1,2,3-cd)pyrene (62.29%), and benzo(ghi)perylene (74.85%). These removal rates were 829.78%, 182.34%, 46.13%, and 70.94% higher than the equivalent alfalfa combination treatments. The lignin peroxidase activity was significantly increased in the presence of starch, ZH-H2 and brome, consistent with the increased removal rates of HMW-PAHs.

High carbon dioxide concentration and elevated temperature impact the growth of weeds but do not change the efficacy of glyphosate.

BACKGROUND: Global climate changes may impact the growth and management of weed species. The aim of this work was to evaluate the impact of recent climate changes on the growth of weeds and herbicide efficacy. The effects of temperature, carbon dioxide (CO2 ), and herbicide on growth and control of Bromus tectorum L., Hordeum murinum L., and Lactuca serriola L. were studied. Treatments included: control or ambient environment (CO2 concentration 400-450 ppm; temperature 20/10 °C day/night); elevated temperature (CO2 concentration 400-450 ppm; temperature 25/15 °C day/night); high CO2 and elevated temperature (CO2 concentration 800-900 ppm; temperature 25/15 °C day/night); high CO2 (CO2 concentration 800-900 ppm; temperature 20/10 °C day/night). Glyphosate rates (active ingredient) used in the experiment were: 0 g ha-1 (untreated control); 360 g ha-1 ; 720 g ha-1 ; 1080 g ha-1 ; 1440 g ha-1 (recommended rate), and 2880 g ha-1 . RESULTS: High CO2 concentration and high CO2 concentration plus high temperature improved the biomass and growth parameters of weeds in the studies. In general, high temperature had a neutral, negative or slightly positive effect on the growth of weed species. Climatic conditions did not affect the activity of glyphosate; its application provided equal and effective weed control under both CO2 and temperature levels and their combinations. CONCLUSION: The positive effect of high CO2 concentration on the growth of weeds does not impact the activity of glyphosate. © 2017 Society of Chemical Industry.

First report of Ser653Asn mutation endowing high-level resistance to imazamox in downy brome (Bromus tectorum L.).

BACKGROUND: Bromus tectorum L. is one of the most troublesome grass weed species in cropland and non-cropland areas of the northwestern USA. In summer 2016, a B. tectroum accession (R) that survived imazamox at the field-use rate (44 g ha-1 ) in an imidazolinone-tolerant (IMI-tolerant or Clearfield™) winter wheat field was collected from a wheat field in Carter County, MT, USA. The aim of this study was to determine the resistance profile of the B. tectroum R accession to imazamox and other ALS inhibitors, and investigate the mechanism of resistance to imazamox. RESULTS: The R B. tectorum accession had a high-level resistance (110.1-fold) to imazamox (IMI) and low to moderate-levels cross-resistance to pyroxsulam (TP) (4.6-fold) and propoxycarbazone (SCT) (13.9-fold). The R accession was susceptible to sulfosulfuron (SU) and quizalofop and clethodim (ACCase inhibitors), paraquat (PS I inhibitor), glyphosate (EPSPS inhibitor) and glufosinate (GS inhibitor). Sequence analysis of the ALS gene revealed a single, target-site Ser653Asn mutation in R plants. Pretreatment of malathion followed by imazamox at 44 or 88 g ha-1 did not reverse the resistance phenotype. CONCLUSION: This is the first report of evolution of cross-resistance to ALS-inhibiting herbicides in B. tectorum. A single-point mutation, Ser653Asn, was identified, conferring the high-level resistance to imazamox. © 2017 Society of Chemical Industry.

Indaziflam: a new cellulose-biosynthesis-inhibiting herbicide provides long-term control of invasive winter annual grasses.

BACKGROUND: Indaziflam is a cellulose-biosynthesis-inhibiting (CBI) herbicide that is a unique mode of action for resistance management and has broad spectrum activity at low application rates. This research further explores indaziflam's activity on monocotyledons and dicotyledons and evaluates indaziflam's potential for restoring non-crop sites infested with invasive winter annual grasses. RESULTS: Treated Arabidopsis, downy brome, feral rye and kochia were all susceptible to indaziflam in a dose-dependent manner. We confirmed that indaziflam has increased activity on monocots (average GR50  = 231 pm and 0.38 g AI ha-1 ) at reduced concentrations compared with dicots (average GR50  = 512 pm and 0.87 g AI ha-1 ). Fluorescence microscopy confirmed common CBI symptomologies following indaziflam treatments, as well as aberrant root and cell morphology. Across five application timings, indaziflam treatments resulted in superior invasive winter annual grass control 2 years after treatment (from 84 ± 5.1% to 99 ± 0.5%) compared with imazapic (36% ± 1.2%). Indaziflam treatments significantly increased biomass and species richness of co-occurring species 2 years after treatment. CONCLUSION: Indaziflam's increased activity on monocots could provide a new alternative management strategy for long-term control of multiple invasive winter annual grasses that invade >23 million ha of US rangeland. Indaziflam could potentially be used to eliminate the soil seed bank of these invasive grasses, reduce fine fuel accumulation and ultimately increase the competitiveness of perennial co-occuring species. © 2017 Society of Chemical Industry.

Phytotoxic activity against Bromus tectorum for secondary metabolites of a seed-pathogenic Fusarium strain belonging to the F. tricinctum species complex.

The winter annual grass Bromus tectorum (cheatgrass) has become highly invasive in semiarid ecosystems of western North America. In these areas, a natural phenomenon, complete cheatgrass stand failure ('die-off'), is apparently caused by a complex interaction among soilborne fungal pathogens. Several Fusarium strains belonging to the Fusarium tricinctum species complex were isolated from these soils and found to be pathogenic on B. tectorum seeds. One of these strains was produced in cheatgrass seed culture to evaluate its ability to produce phytotoxins. Six metabolites were isolated and identified by spectroscopic methods (essentially 1D and 2D NMR and ESIMS) as acuminatopyrone (1), blumenol A (2), chlamydosporol (3), isochlamydosporol (4), ergosterol (5) and 4-hydroxybenzaldehyde (6). Upon testing against B. tectorum in a seedling bioassay, (6) the coleoptile and radicle length of cheatgrass seedlings were significantly reduced. Compounds 1 and 2 showed moderate activity, while 3-5 were not significantly different from the control.

Viral pathogen production in a wild grass host driven by host growth and soil nitrogen.

Nutrient limitation is a basic ecological constraint that has received little attention in studies on virus production and disease dynamics. Nutrient availability could directly limit the production of viral nucleic acids and proteins, or alternatively limit host growth and thus indirectly limit metabolic pathways necessary for viral replication. In order to compare direct and indirect effects of nutrient limitation on virus production within hosts, we manipulated soil nitrogen (N) and phosphorus (P) availability in a glasshouse for the wild grass host Bromus hordeaceus and the viral pathogen Barley yellow dwarf virus-PAV. We found that soil N additions increased viral concentrations within host tissues, and the effect was mediated by host growth. Specifically, in statistical models evaluating the roles of host biomass production, leaf N and leaf P, viral production depended most strongly on host biomass, rather than the concentration of either nutrient. Furthermore, at low soil N, larger plants supported greater viral concentrations than smaller ones, whereas at high N, smaller plants supported greater viral concentrations. Our results suggest that enhanced viral productivity under N enrichment is an indirect consequence of nutrient stimulation to host growth rate. Heightened pathogen production in plants has important implications for a world facing increasing rates of nutrient deposition.

Pyrenophoric acids B and C, two new phytotoxic sesquiterpenoids produced by Pyrenophora semeniperda.

Two new phytotoxic sesquiterpenoid acids, named pyrenophoric acids B and C, were isolated together with the related pyrenophoric and abscisic acids from solid Bromus tectorum (cheatgrass) seed culture of the seed pathogen Pyrenophora semeniperda. This fungus has been proposed as a mycoherbicide for biocontrol of cheatgrass (Bromus tectorum), a Eurasian annual grass that has become invasive in rangelands and is also a serious agricultural weed in the western U.S. Pyrenophoric acids B and C were characterized by spectroscopic methods (NMR and HR ESIMS) as (2Z,4E)-5-[(1R*,4R*,6R*)-1,4-dihydroxy-2,2,6-trimethylcyclohexyl]-3-methylpenta-2,4-dienoic and (2Z,4E)-5-[(1S*,3S*,4R*,6S*)-3,4-dihydroxy-2,2,6-trimethylcyclohexyl]-3-methylpenta-2,4-dienoic acids, respectively. Cytochalasins A, B, F, and Z3, as well as deoxaphomin and pyrenophoric acid, all previously isolated from P. semeniperda grown on wheat seed, were also isolated from cheatgrass seed culture. In a cheatgrass seedling bioassay at 10(-3) M, pyrenophoric acid B showed higher coleoptile toxicity than pyrenophoric acid, while pyrenophoric acid C showed lower phytotoxicity. Abscisic acid was by far the most active compound.

Abscisic acid induced freezing tolerance in chilling-sensitive suspension cultures and seedlings of rice.

BACKGROUND: The role of abscisic acid (ABA) as a possible activator of cold acclimation process was postulated since endogenous levels of ABA increase temporarily or constitutively during cold-hardening. Exogenous application of ABA has been known to induce freezing tolerance at ambient temperatures in in vitro systems derived from cold hardy plants. Yet, some cell cultures acquired much greater freezing tolerance by ABA than by cold whilst maintaining active growth. This raises questions about the relationships among ABA, cold acclimation and growth cessation. To address this question, we attempted to 1) determine whether exogenous ABA can confer freezing tolerance in chilling-sensitive rice suspension cells and seedlings, which obviously lack the mechanisms to acquire freezing tolerance in response to cold; 2) characterize this phenomenon by optimizing the conditions and compare with the case of cold hardy bromegrass cells. RESULTS: Non-embryogenic suspension cells of rice suffered serious chilling injury when exposed to 4°C. When incubated with ABA at the optimal conditions (0.5-1 g cell inoculum, 75 µM ABA, 25-30°C, 7-10 days), they survived slow freezing (2°C/h) to -9.0 ~ -9.3°C (LT50: 50% killing temperature) while control cells were mostly injured at -3°C (LT50: -0.5 ~ -1.5°C). Ice-inoculation of the cell suspension at -3°C and survival determination by regrowth confirmed that ABA-treated rice cells survived extracellular freezing at -9°C. ABA-induced freezing tolerance did not require any exposure to cold and was best achieved at 25-30°C where the rice cells maintained high growth even in the presence of ABA. ABA treatment also increased tolerance to heat (43°C) as determined by regrowth. ABA-treated cells tended to have more augmented cytoplasm and/or reduced vacuole sizes compared to control cultures with a concomitant increase in osmolarity and a decrease in water content. ABA-treated (2-7 days) in vitro grown seedlings and their leaves survived slow freezing to -3°C with only marginal injury (LT50: -4°C) whereas untreated seedlings were killed at -3°C (LT50: -2°C). CONCLUSIONS: The results indicate that exogenous ABA can induce some levels of freezing tolerance in chilling-sensitive rice cells and seedlings, probably by eliciting mechanisms different from low temperature-induced cold acclimation.

Elevated CO2 spurs reciprocal positive effects between a plant virus and an arbuscular mycorrhizal fungus.

Plants form ubiquitous associations with diverse microbes. These interactions range from parasitism to mutualism, depending partly on resource supplies that are being altered by global change. While many studies have considered the separate effects of pathogens and mutualists on their hosts, few studies have investigated interactions among microbial mutualists and pathogens in the context of global change. Using two wild grass species as model hosts, we grew individual plants under ambient or elevated CO(2), and ambient or increased soil phosphorus (P) supply. Additionally, individuals were grown with or without arbuscular mycorrhizal inoculum, and after 2 wk, plants were inoculated or mock-inoculated with a phloem-restricted virus. Under elevated CO(2), mycorrhizal association increased the titer of virus infections, and virus infection reciprocally increased the colonization of roots by mycorrhizal fungi. Additionally, virus infection decreased plant allocation to root biomass, increased leaf P, and modulated effects of CO(2) and P addition on mycorrhizal root colonization. These results indicate that plant mutualists and pathogens can alter each other's success, and predict that these interactions will respond to increased resource availability and elevated CO(2). Together, our findings highlight the importance of interactions among multiple microorganisms for plant performance under global change.

Non-target-site-based resistance to ALS-inhibiting herbicides in six Bromus rigidus populations from Western Australian cropping fields.

BACKGROUND: Bromus rigidus is a common weed species that has increased in cropping fields owing to limited control options. During a random field survey in Western Australia, six B. rigidus populations that had survived in-crop weed control programmes were collected. The study aimed to determine the resistance profile of these six populations. RESULTS: Based on dose-response studies, all six B. rigidus populations had a low-level resistance to sulfosulfuron and sulfometuron (both sulfonylurea herbicides) while remaining susceptible to herbicides with other modes of action. ALS in vitro activity assays revealed no differences in enzyme sensitivity between susceptible and resistant populations, while the use of malathion (a cytochrome P450 inhibitor) in combination with sulfosulfuron caused the resistant populations to behave like the susceptible population. CONCLUSION: This study established that these six B. rigidus populations have a low-level resistance to the ALS-inhibiting sulfonylurea herbicides, but are able to be controlled by other herbicide modes of action. The low-level, malathion-reversible resistance, together with a sensitive ALS, strongly suggest that a non-target-site enhanced metabolism is the mechanism of resistance.

Pyrenophora bromi, causal agent of brownspot of bromegrass, expresses a gene encoding a protein with homology and similar activity to Ptr ToxB, a host-selective toxin of wheat.

Ptr ToxB, encoded by ToxB, is one of multiple host-selective toxins (HST) produced by the wheat pathogen Pyrenophora tritici-repentis. Homologs of ToxB are found in several ascomycetes, including sister species Pyrenophora bromi, causal agent of brownspot of bromegrass. Due to the close evolutionary relatedness of P. tritici-repentis and P. bromi and that of their grass hosts, we hypothesized that homologs of ToxB in P. bromi may act as HST in the disease interaction between P. bromi and bromegrass. A representative set of transcriptionally active P. bromi ToxB genes were heterologously expressed in Pichia pastoris and the resultant proteins tested for their ability to act as HST on bromegrass. The tested Pyrenophora bromi ToxB (Pb ToxB) proteins were not toxic to bromegrass; thus, Pb ToxB does not appear to function as an HST in the P. bromi-bromegrass interaction. Instead, we revealed that the Pb ToxB proteins can be toxic to Ptr ToxB-sensitive wheat, at levels similar to Ptr ToxB, and the corresponding P. bromi ToxB genes are expressed in P. bromi-inoculated wheat. Our data suggest that P. bromi possesses the potential to become a wheat pathogen and highlights the importance of investigating the interaction between P. bromi and wheat.

Variability in C(3)-plant cell-wall biosynthesis in a high-CO(2) atmosphere by solid-state NMR spectroscopy.

We have used a frequency-selective rotational-echo double-resonance (REDOR) solid-state NMR experiment to measure the concentrations of glycine-glycine pairs in proteins (and protein precursors) of intact leaves of plants exposed to both high- and low-CO(2) atomospheres. The results are interpreted in terms of differences in cell-wall biosynthesis between plant species. We illustrate this variability by comparing the assimilation of label in cheatgrass and soybean leaves labeled using (15)N-fertilizer and (13)CO(2) atmospheres. Cheatgrass and soybean are both C(3) plants but differ in their response to a high-CO(2) environment. Based on REDOR results, we determined that cheatgrass (a plant that seems likely to flourish in future low-water, high-CO(2) environments) routes 2% of the assimilated carbon label that remains in the leaf after 1 h in a 600-ppm (13)CO(2) atmosphere to glycine-rich protein (or its precursors), a structural component of cell walls cross-linked to lignins. In contrast, soybean under the same conditions routes none of its assimilated carbon to glycine-rich protein.

Herbicidal activity of glucosinolate degradation products in fermented meadowfoam ( Limnanthes alba ) seed meal.

Meadowfoam ( Limnanthes alba ) is an oilseed crop grown in western Oregon. After extraction of the oil from the seeds, the remaining seed meal contains 2-4% of the glucosinolate glucolimnanthin. This study investigated the effect of fermentation of seed meal on its chemical composition and the effect of the altered composition on downy brome ( Bromus tectorum ) coleoptile emergence. Incubation of enzyme-inactive seed meal with enzyme-active seeds (1% by weight) resulted in complete degradation of glucolimnanthin and formation of 3-methoxybenzyl isothiocyanate in 28% yield. Fermentation in the presence of an aqueous solution of FeSO(4) (10 mM) resulted in the formation of 3-methoxyphenylacetonitrile and 2-(3-methoxyphenyl)ethanethioamide, a novel natural product. The formation of the isothiocyanate, the nitrile, and the thioamide, as a total, correlated with an increase of herbicidal potency of the seed meal (r(2) = 0.96). The results of this study open new possibilities for the refinement of glucosinolate-containing seed meals for use as bioherbicides.

Stomatal conductance of semi-natural Mediterranean grasslands: implications for the development of ozone critical levels.

Intra-genus and intra-specific variation and the influence of nitrogen enrichment on net assimilation and stomatal conductance of some annual Trifolium species of Mediterranean dehesa grasslands were assessed under experimental conditions. Also gas exchange rates were compared between some Leguminosae and Poaceae species growing in the field in a dehesa ecosystem in central Spain. The results showed that the previously reported different O3 sensitivity of some Trifolium species growing in pots does not seem to be related to different maximum g(s) values. In addition, no clear differences on gas exchange rates could be attributed to Leguminosae and Poaceae families growing in the field, with intra-genus variation being more important than differences found between families. Further studies are needed to increase the database for developing a flux-based approach for setting O3 critical levels for semi-natural Mediterranean species.

Ophiobolin E and 8-epi-ophiobolin J produced by Drechslera gigantea, a potential mycoherbicide of weedy grasses.

Drechslera gigantea, a fungal pathogen isolated from large crabgrass (Digitaria sanguinalis) and proposed as a potential mycoherbicide of grass weeds, produces phytotoxic metabolites in liquid and solid cultures. Ophiobolin A and three minor ophiobolins i.e., 6-epi-ophiobolin A, 3-anhydro-6-epi-ophiobolin A and ophiobolin I were obtained from the liquid culture broths. Interestingly and unexpectedly, ophiobolins also appeared in cultures of this fungus and they were isolated together with the known ophiobolins B and J, and designed as ophiobolin E and 8-epi-ophiobolin J. They were characterized using essentially spectroscopic methods. It is noteworthy that D. gigantea produces such a plethora of bioactive organic substances. Some structure-activity relationship results are also discussed in this report.

[Allelopathy of aqueous extract from Ligularia virgaurea, a dominant weed in psychro-grassland, on pasture plants].

Ligularia virgaurea is a noxious weed widely distributed in the alpine grassland of east Qinghai-Tibet Plateau of China. This paper studied the allelopathy of its aqueous extract on the pasture plants Festuca sinensis, Bromus magnus, Elymus nutans, Poa annua, and F. ovina in the region. The mean response index (RI) values of the pasture plants were calculated, and used to quantitatively assess the allelopathic sensitivity of the receptors at three levels, i. e., growth items, development stages, and species. Corresponding values of the weed were also treated in similar way to assess the allelopathic potential of the donor. The results showed that the allelopathic sensitivity was in the order of P. annua > B. magnus > F. sinensis > F. ovina > E. nutans. Both the seed germination and the seedling growth of test pasture plants were inhibited at species level, suggesting that rain eluviation was one of the means by which the weed released allelochemicals. The aqueous extracts from L. virgaurea root and leaf had a significant inhibitory effect at species level, and the effect of root extract was stronger than that of leaf extract, suggesting the competition among species on the underground resources in natural grassland. Allelopathy played an important role in L. virgaurea invasion, and might be responsible to the formation of mono-dominant community and the degeneration of grassland.

Effect of growth phase on survival of bromegrass suspension cells following cryopreservation and abiotic stresses.

BACKGROUND AND AIMS: Cryopreservation is a practical method of preserving plant cell cultures and their genetic integrity. It has long been believed that cryopreservation of plant cell cultures is best performed with cells at the late lag or early exponential growth phase. At these stages the cells are small and non-vacuolated. This belief was based on studies using conventional slow prefreezing protocols and survival determined with fluorescein diacetate staining or 2,3,5-triphenyltetrazolium chloride assays. This classical issue was revisited here to determine the optimum growth phase for cryopreserving a bromegrass (Bromus inermis) suspension culture using more recently developed protocols and regrowth assays for determination of survival. METHODS: Cells at different growth phases were cryopreserved using three protocols: slow prefreezing, rapid prefreezing and vitrification. Stage-dependent trends in cell osmolarity, water content and tolerance to freezing, heat and salt stresses were also determined. In all cases survival was assayed by regrowth of cells following the treatments. KEY RESULTS: Slow prefreezing and rapid prefreezing protocols resulted in higher cell survival compared with the vitrification method. For all the protocols used, the best regrowth was obtained using cells in the late exponential or early stationary phase, whereas lowest survival was obtained for cells in the late lag or early exponential phase. Cells at the late exponential phase were characterized by high water content and high osmolarity and were most tolerant to freezing, heat and salt stresses, whereas cells at the early exponential phase, characterized by low water content and low osmolarity, were least tolerant. CONCLUSIONS: The results are contrary to the classical concept which utilizes cells in the late lag or early exponential growth phase for cryopreservation. The optimal growth phase for cryopreservation may depend upon the species or cell culture being cryopreserved and requires re-investigation for each cell culture. Stage-dependent survival following cryopreservation was proportionally correlated with the levels of abiotic stress tolerance in bromegrass cells.

A lipid transfer protein gene BG-14 is differentially regulated by abiotic stress, ABA, anisomycin, and sphingosine in bromegrass (Bromus inermis).

The objective was to investigate the expression of a lipid transfer protein gene (LTP) both in bromegrass (Bromus inermis) cells and seedlings after exposure to abiotic stresses, abscisic acid (ABA), anisomycin, and sphingosine. A full-length cDNA clone BG-14 isolated from bromegrass suspension cell culture encodes a polypeptide of 124 amino acids with typical LTP characteristics, such as a conserved arrangement of cysteine residues. During active stages of cold acclimation LTP expression was up-regulated, whereas at the final stage of cold acclimation LTP transcript level declined to pre-acclimation level. A severe drought stress induced the LTP gene; yet, LTP expression doubled 3 d after re-hydration. Both temperature and heat shock duration influence LTP induction; however temperature is the primary factor. Treatment with NaCl stimulated accumulation of LTP mRNA within 15 min and the transcripts remained at elevated levels for the duration of the salinity stress. Most interestingly, Northern blots showed LTP was rapidly induced not only by ABA, but also by anisomycin and sphingosine in suspension cell cultures. Of the three chemicals, ABA induced the most rapid and highest response in LTP expression as well as highest freezing tolerance, whereas sphingosine was the least active for both LTP expression and freezing tolerance.