Diversity of Claviceps paspali reveals unknown lineages and unique alkaloid genotypes.

Claviceps species affecting Paspalum spp. are a serious problem, as they infect forage grasses such as Paspalum dilatatum and P. plicatulum, producing the ergot disease. The ascomycete C. paspali is known to be the pathogen responsible for this disease in both grasses. This fungus produces alkaloids, including ergot alkaloids and indole-diterpenes, that have potent neurotropic activities in mammals. A total of 32 isolates from Uruguay were obtained from infected P. dilatatum and P. plicatulum. Isolates were phylogenetically identified using partial sequences of the genes coding for the second largest subunit of RNA polymerase subunit II (RPB2), translation elongation factor 1-α (TEF1), ß-tubulin (TUB2), and the nuc rDNA 28S subunit (28S). Isolates were also genotyped by randomly amplified polymorphic DNA (RAPD) and presence of genes within the ergot alkaloid (EAS) and indole-diterpene (IDT) biosynthetic gene clusters. This study represents the first genetic characterization of several isolates of C. paspali. The results from this study provide insight into the genetic and genotypic diversity of Claviceps paspali present in P. dilatatum and suggest that isolates from P. plicatulum could be considered an ecological subspecies or specialized variant of C. paspali. Some of these isolates show hypothetical alkaloid genotypes never reported before.

Contrasting P acquisition strategies of the bacterial communities associated with legume and grass in subtropical orchard soil.

Phosphorus (P) cycling is a fundamental process driven by microorganisms, and plants can regulate P cycling directly or via their influence on the soil microbial community. However, the differential P cycling patterns associated with legumes and grass are largely unknown. Therefore, we investigated the microbial community involved in P cycling in subtropical soil grown with stylo (Stylosanthes guianensis, legume) or bahiagrass (Paspalum notatum, grass) using metagenomic sequencing. P fractionation indicated that sparingly soluble inorganic P (Pi) accounted for approximately 75% of P pool. Bacteria involved in sparingly soluble Pi solubilization (pqq, gad, JEN) were more abundant in bahiagrass soil, with Candidatus Pelagibacter, Trichodesmium, Neorickettsia, Nitrobacter, Paraburkholderia, Candidatus Solibacter, Burkholderia as major contributors. In contrast, bacteria involved in organic P (Po) mineralization (php, glpQ, phn) were more abundant in stylo soil, consistent with phosphatase activity and Frankia, Kyrpidia, Thermobispora, Streptomyces, Rhodococcus were major contributors. Bacteria taking up low molecular-weight Po were more abundant in stylo soil than in bahiagrass soil, while those taking up Pi were less abundant. These data suggest that bacterial communities associated with legumes and grass develop contrasting P acquisition strategies, highlighting the possibility of intercropping with legumes and grass for better P cycling.

Cellulase activity and dissolved organic carbon release from lignocellulose macrophyte-derived in four trophic conditions

Abstract Considering the importance of lignocellulose macrophyte-derived for the energy flux in aquatic ecosystems and the nutrient concentrations as a function of force which influences the decomposition process, this study aims to relate the enzymatic activity and lignocellulose hydrolysis in different trophic statuses. Water samples and two macrophyte species were collected from the littoral zone of a subtropical Brazilian Reservoir. A lignocellulosic matrix was obtained using aqueous extraction of dried plant material (≈40 °C). Incubations for decomposition of the lignocellulosic matrix were prepared using lignocelluloses, inoculums and filtered water simulating different trophic statuses with the same N:P ratio. The particulate organic carbon and dissolved organic carbon (POC and DOC, respectively) were quantified, the cellulase enzymatic activity was measured by releasing reducing sugars and immobilized carbon was analyzed by filtration. During the cellulose degradation indicated by the cellulase activity, the dissolved organic carbon daily rate and enzyme activity increased. It was related to a fast hydrolysable fraction of cellulose that contributed to short-term carbon immobilization (ca. 10 days). After approximately 20 days, the dissolved organic carbon and enzyme activity were inversely correlated suggesting that the respiration of microorganisms was responsible for carbon mineralization. Cellulose was an important resource in low nutrient conditions (oligotrophic). However, the detritus quality played a major role in the lignocelluloses degradation (i.e., enzyme activity) and carbon release.

Cellulase activity and dissolved organic carbon release from lignocellulose macrophyte-derived in four trophic conditions.

Considering the importance of lignocellulose macrophyte-derived for the energy flux in aquatic ecosystems and the nutrient concentrations as a function of force which influences the decomposition process, this study aims to relate the enzymatic activity and lignocellulose hydrolysis in different trophic statuses. Water samples and two macrophyte species were collected from the littoral zone of a subtropical Brazilian Reservoir. A lignocellulosic matrix was obtained using aqueous extraction of dried plant material (≈40°C). Incubations for decomposition of the lignocellulosic matrix were prepared using lignocelluloses, inoculums and filtered water simulating different trophic statuses with the same N:P ratio. The particulate organic carbon and dissolved organic carbon (POC and DOC, respectively) were quantified, the cellulase enzymatic activity was measured by releasing reducing sugars and immobilized carbon was analyzed by filtration. During the cellulose degradation indicated by the cellulase activity, the dissolved organic carbon daily rate and enzyme activity increased. It was related to a fast hydrolysable fraction of cellulose that contributed to short-term carbon immobilization (ca. 10 days). After approximately 20 days, the dissolved organic carbon and enzyme activity were inversely correlated suggesting that the respiration of microorganisms was responsible for carbon mineralization. Cellulose was an important resource in low nutrient conditions (oligotrophic). However, the detritus quality played a major role in the lignocelluloses degradation (i.e., enzyme activity) and carbon release.

Microdochium paspali, a new species causing seashore paspalum disease in southern China.

A new species of Microdochium was identified as the causal agent of leaf blight of seashore paspalum (Paspalum vaginatum), a turf grass widely used in tropical and subtropical golf courses. In 2010 foliar necrosis and canopy thinning were observed on 11 surveyed golf courses in Hainan province, China, especially on fairways and putting greens. The infected leaves initially appeared water-soaked and dark green, rapidly faded to yellow or became chlorotic and quickly died, resulting in a sparse appearance in infected areas, leading to the disease name "sparse leaf patch." Isolates with rich and light pink to yellow mycelia and salmon-colored pionnotes were cultured from diseased turf foliage. Pathogenicity was demonstrated by inoculating these isolates onto "seaspray" seashore paspalum. Phylogenetic analysis based on the nuc rDNA internal transcribed spacer 1-5.8S-internal transcribed spacer 2 region (ITS), translation elongation factor 1-α (TEF1-α) and ß-tubulin (BenA) indicated these isolates formed as a distinct clade within Microdochium (Xylariales). Further microscopic examination demonstrated that the species was morphologically distinct from three similar species of Microdochium. The name Microdochium paspali sp. nov. is proposed for this novel fungal pathogen.

Fly ash mycorrhizoremediation through Paspalum scrobiculatum L., inoculated with Rhizophagus fasciculatus.

Fly ash is the residue produced during the combustion of coal, and its disposal is a major environmental concern worldwide. However, fly ash can ameliorate soils by improving their physical, chemical, and biological properties. Hence, we conducted a study to understand the mycorrhizoremediation of different levels of fly ash (2%, 4%, and 6%) by using kodo millet (Paspalum scrobiculatum L.) inoculated with arbuscular mycorrhizal (AM) fungus Rhizophagus fasciculatus under greenhouse conditions. Fly ash amendment at a low level (2%) significantly enhanced AM colonization, spore number, plant growth, nutrient uptake, nutrient-use efficiencies and grain yield of kodo millet. Nevertheless, inoculation of soils amended with 2% fly ash with the AM fungus further enhanced the AM fungal, plant growth, nutrient uptake and yield parameters. Arbuscular mycorrhizal colonization decreased with increasing concentrations of fly ash amendment; however, such decrease was not linear. Our results also revealed a significantly higher plant growth, root/shoot ratios and nutrient contents in kodo millet shoots raised on 2% fly ash amendment and inoculated with the AM fungus at both harvests. Both fly ash amendment and AM fungus inoculation also significantly influenced the number of grains produced as well as the grain weight. Arbuscular mycorrhizal inoculation and fly ash amendment affected K, Ca, Mg, Na use efficiencies. Plant growth and nutrient parameters were strongly related to the extent of AM fungal colonization in the roots. These observations suggest that the inoculation of AM fungi along with low levels of fly ash amendment could be effectively used for the reclamation of low fertile or marginal soils and in turn fly ash could aid in crop production.

Mercury and flooding cycles in the Tapajós River basin, Brazilian Amazon: the role of periphyton of a floating macrophyte (Paspalum repens).

Methylmercury (MeHg) increases mercury (Hg) toxicity and is biomagnified in the trophic chain contaminating riverine Amazon populations. Freshwater macrophyte roots are a main site of Hg methylation in different Brazilian environments. Paspalum repens periphyton was sampled in four floodplain lakes during the dry, rainy and wet seasons for measurement of total Hg (THg), MeHg, Hg methylation potentials, %C, %N, δ(13)C, δ(15)N and bacterial heterotrophic production as (3)H-leucine incorporation rate. THg concentration varied from 67 to 198 ng/g and the potential of Me(203)Hg formation was expressive (1-23%) showing that periphyton is an important matrix both in the accumulation of Hg and in MeHg production. The concentration of MeHg varied from 1 to 6 ng/g DW and was positively correlated with Me(203)Hg formation. Though methylmercury formation is mainly a bacterial process, no significant correlation was observed between the methylation potentials and bacterial production. The multiple regressions analyses suggested a negative correlation between THg and %C and %N and between methylation potential and δ(13)C. The discriminant analysis showed a significant difference in periphyton δ(15)N, δ(13)C and THg between seasons, where the rainy season presented higher δ(15)N and the wet period lighter δ(13)C, lower THg values and higher Me(203)Hg formation. This exploratory study indicates that the flooding cycle could influence the periphyton composition, mercury accumulation and methylmercury production.

Exploring soil bacterial communities in different peanut-cropping sequences using multiple molecular approaches.

Soil bacterial communities have significant influence on soilborne plant pathogens and, thus, crop health. The present study focuses on ribotyping soil bacterial communities in different peanut-cropping sequences in Alabama. The objective was to identify changes in microbial assemblages in response to cropping sequences that can play a role in managing soilborne plant pathogens in peanut. Four peanut-cropping sequences were sampled at the Wiregrass Research Station, Headland, AL in 2006 and 2007, including continuous peanut, 4 years of bahiagrass followed by peanut, peanut-cotton, and peanut-corn-cotton. Soil sampling was done at early and mid-season and at harvest. Bacterial community structure was assessed using ribosomal intergenic spacer analysis (RISA) combined with 16S rRNA cloning and sequencing. RISA results indicated >70% dissimilarities among different cropping sequences. However, 90% similarities were noticed among replicated plots of the same cropping sequences. Cropping sequences and time of soil sampling had considerable effect on soil microbial community structure. Bahiagrass rotation with peanut was found to have the highest bacterial diversity, as indicated by a high Shannon Weaver Diversity index. Overall, higher bacterial diversity was observed with bahiagrass and corn rotations compared with continuous peanut. The bacterial divisions Proteobacteria, Acidobacteria, Firmicutes, Bacteroidetes, and Actinomycetes were the predominant bacterial phyla found in all peanut-cropping sequences. The Proteobacteria taxa in these soils were negatively correlated with the abundance of members of division Firmicutes but, conversely, had a significant positive correlation with Gemmatimonadetes taxa. The prevalence of the division Actinomycetes was negatively correlated with the relative abundance of members of division Verrucomicrobia. These results indicate complex interactions among soil bacteria that are important contributors to crop health.

Staggers in horses grazing paspalum infected with Claviceps paspali.

Invasion of the flowering heads of grasses by Claviceps spp. can produce sclerotia (ergots) containing several toxins. Ingestion of these toxins, through the consumption of paspalum (Paspalum dilatatum), can induce a range of clinical symptoms, including staggers. Cattle are the most commonly affected species, but although sheep and horses have been reported affected there are no published descriptions of paspalum staggers in horses. We describe two occurrences of paspalum staggers, the first in three Australian Stockhorse foals and the second in mature Standardbred horses. All three foals presented with ataxia in all limbs after consuming infected paspalum. One foal died from misadventure and the other two recovered within 1 week of removal from the infected paddock. In the second case, two of eight mares and geldings grazing in an irrigation channel developed hindquarter paresis. After removal of all horses from the area, one of the affected horses continued to deteriorate. Both horses were treated with antibiotics. The more severely affected horse was also treated with fluids and electrolytes, but had to be euthanased. The second affected horse recovered after 2 days. Paspalum pastures should inspected for Claviceps paspali infection before the introduction of horses.

On-farm production of inoculum of indigenous arbuscular mycorrhizal fungi and assessment of diluents of compost for inoculum production.

On-farm production of arbuscular mycorrhizal [AM] fungus inoculum can be employed to make the benefits of the symbiosis more available to vegetable farmers. Experiments were conducted to modify an existing method for the production of inoculum in temperate climates to make it more readily adoptable by farmers. Perlite, vermiculite, and peat based potting media were tested as diluents of yard clippings compost for the media in which the inoculum was produced using bahiagrass (Paspalum notatum Flugge) as host plant. All produced satisfactory concentrations of AM fungus propagules, though vermiculite proved to be better than potting media (89 vs. 25 propagules cm(-3), respectively). Two methods were tested for the growth of AM fungi indigenous to the farm: (1) adding field soil into the vermiculite and compost mixture and (2) pre-colonizing the bahiagrass seedlings in media inoculated with field soil prior to transplant into that mixture. Adding 100 cm(3) of field soil to the compost and vermiculite produced 465 compared to 137 propagules cm(-3) for the pre-colonization method. The greater flexibility these modifications give will make it easier for farmers to produce inoculum of AM fungi on-the-farm.

Toxic effects of chlorate on three plant species inoculated with arbuscular mycorrhizal fungi.

Pot experiments were conducted to examine the toxic effects of chlorate on bermudagrass, bahiagrass, and longan seedling with a focus on arbuscular mycorrhizal fungi-plant associations. The results show that application of chlorate could cause slight soil acidification, but the resulting pH was still around 5.5, which is unlikely to adversely affect plant growth. Increase in the application rate of chlorate resulted in a decrease in colonization rate of arbuscular mycorrhizal fungi in plant roots, P uptake by the plants and plant biomass. This appears to suggest that the reduction in plant growth may be related to impeded uptake of P by the plants due to the failure of the plants to form sufficient mycorrhizal associations when chlorate is in sufficient amounts to cause toxicity to arbuscular mycorrhizal fungi. Under the experimental conditions set for this study, bermudagrass suffered from stronger chlorate stress than bahiagrass and longan seedling did in terms of plant-arbuscular mycorrhizal fungi (AMF) symbiosis development.

On-farm production of AM fungus inoculum in mixtures of compost and vermiculite.

On-farm production of arbuscular mycorrhizal (AM) fungus inoculum can reduce the cost of the inoculum and increase utilization of this symbiosis in plant production. Bahiagrass (Paspalum notatum Flugge) seedlings, colonized by AM fungi, were transplanted into raised bed enclosures. Media within the enclosures was vermiculite mixed with either field soil or yard clippings compost in Experiment I and vermiculite mixed with yard clippings compost or dairy manure/leaf compost in Experiment II. Compost and vermiculite mixtures yielded more propagules of AM fungi than soil-based mixtures in Experiment I. Growth of plants in a 1:4 (v/v) mixture of yard clippings compost and vermiculite produced more inoculum (503 propagules cm(-3)) than growth in 1:9 and 1:99 (v/v) mixtures (240 and 42 propagules cm(-3), respectively). Water, inorganic nutrient solution minus P, and fish protein digest were added to inoculum production enclosures in Experiment II. Results indicated that supplemental nutrient addition was unnecessary. This method produces a concentrated inoculum of AM fungi in a form readily used as an amendment to horticultural potting media for the production of vegetable seedlings.

Action spectrum for the induction of hyphal branches of an arbuscular mycorrhizal fungus: exposure sites versus branching sites.

The first action spectrum for a photo-induced response of an arbuscular mycorrhizal fungus is reported. At low light intensity, the responsive wavelengths for light-induced hyphal branching of the primary germ tube of Gigaspora gigantea were determined to be in the blue to uv-A range. The action spectrum showed the greatest stimulation of branching occurred around 390 nm although a shoulder was observed between 360-370 nm. A second major peak of light-induced branching occurred at 430 nm. The exposure of specific areas of the germ tube to high intensity blue light for a short period led to several interesting observations. By exposing 2 mm segments (0-2, 2-4, 4-6, etc.) or 3 mm segments away from the tip, it was determined that photoinduction of hyphal branches could occur anywhere along the axis of a growing germ tube except in the apical 2 mm. When 3 mm segments were exposed at greater distances from the tip (6-9, 9-12, and up to 33-36 mm), branches frequently formed in areas not directly exposed to light. The branches were usually in clusters which were spaced approximately 3 or 6 mm apart. Since light scattering could be ruled out, these results indicated that the exposure sites and sites of hyphal branching did not necessarily coincide and suggested the probable involvement of a second messenger during this blue light-induced event.

Potential risk of acute hepatotoxicity of kodo poisoning due to exposure to cyclopiazonic acid.

Kodo millet (Paspalum scrobiculatum L.) is a staple food of some sections of people of North India. Consumption of Kodo millet is often found to cause intoxication and poisoning. The grains are frequently infested with Aspergillus tamarii Kita, which produced substantial amount of a mycotoxin, cyclopiazonic acid (CPA). Investigations were carried out to evaluate the hepatotoxic/preneoplastic changes in rat liver following single and multiple dose administration of CPA. Results showed a marked increase in the activity of glutamate pyruvate transaminase (GPT) and glutamate oxaloacetate transaminase (GOT) following CPA exposures, suggesting acute hepatotoxicity. Significant increase was also observed in gamma glutamyl transpeptidase (GGT) activity following CPA exposures, indicating preneoplastic changes in the liver. The results reveal that Kodo poisoning might cause acute hepatotoxicity in men and animals. The findings thus suggest that the consumption of contaminated Kodo millet is a serious health hazard due to exposure to CPA produced by Aspergillus tamarii associated with the millet.