Biosynthesis of Depsipeptide Mycotoxins in Fusarium

The cyclic hexadepsipeptide enniatin is known as a phytopathogenic compound from Fusaria causing necrosis and wilt. The molecule consists of three alternating residues each of a branched chain amino acid and D-hydroxyisovaleric acid (D-Hiv). Enniatins are synthesized by a 347kDa multienzyme (enniatin synthetase) via a thiol template mechanism. The corresponding gene esyn1 has an open reading frame of 9393 nucleotides and harbours two modules, one responsible for D-hydroxy acid activation and one for L-amino acid activation with an integrated N-methyltransferase domain. Such methyltransferases build an homologous group among N-methyl peptide synthetases. Enniatins are synthesized by step-wise condensation of dipeptidol building blocks in an iterative manner resembling fatty acid synthesis. A key enzyme in enniatin biosynthesis is the NADPH-dependent D-2-hydroxyisovalerate dehydrogenase, that supplies enniatin synthetase with D-Hiv. Enniatins contribute to the wilt toxic character of Fusaria. Virulence was significantly reduced in F. avenaceum after disruption of the esyn1 gene.     

The Effect of Inoculation Treatment and Long-term Application of Moisture on Fusarium Head Blight Symptoms and Deoxynivalenol Contamination in Wheat Grains

Fusarium head blight (FHB) is an important disease of wheat, which can result in the contamination of grains with mycotoxins such as deoxynivalenol (DON). Artificial inoculation of flowering ears with conidial suspensions is widely used to study FHB diseases. Our goal was to compare four inoculation treatments in which a conidial suspension was sprayed on flowering ears and to study the effect of the application of moisture during kernel setting and filling with a mist-irrigation system. Ten wheat genotypes were inoculated with a DON-producing Fusarium culmorum strain. Inoculation treatments varied in time of application of the inoculum (morning or evening) and in the method of controlling humidity during inoculation (bagging or mist irrigation). A wet season was simulated with a mist-irrigation system, keeping the crop canopy wet for at least 26 days after flowering. The severity of FHB symptoms (area under disease progress curve (AUDPC)), yield loss and DON contamination in the grains were determined. AUDPC data obtained with the different inoculation treatments were highly correlated (r=0.85–0.95). Mist irrigation after inoculation resulted in a higher mean disease severity, but in a overall lower toxin contamination as compared to the non-irrigated treatments. Genotypic differences in DON accumulation were present: for one wheat line toxin contamination significantly increased when irrigated, while two genotypes accumulated significantly less toxin. The closest relationships (r=0.73–0.89) between the visual symptoms and the DON content were obtained under moderate mean infection pressure. This relation between visual symptoms and the DON content deteriorated at higher infection levels.     

Evaluation of a combined transcutaneous carbon dioxide pressure and pulse oximetry sensor in adult sheep and dogs

OBJECTIVE: To evaluate a combined transcutaneous carbon dioxide pressure (tcPCO(2)) and pulse oximetry sensor in sheep and dogs. ANIMALS: 13 adult sheep and 11 adult dogs. PROCEDURES: During inhalation anesthesia, for the first 10 minutes following sensor placement, arterial blood gas was analyzed and tcPCO(2) was recorded every 2 minutes. Subsequently, the animals were hyper-, normo-, and hypoventilated. The simultaneously obtained tcPCO(2) and PaCO(2) values were analyzed by use of Bland-Altman statistical analysis. RESULTS: Mean +/- SD overall difference between tcPCO(2) and PaCO(2) 10 minutes after sensor application was 13.3 +/- 8.4 mm Hg in sheep and 8.9 +/- 12 mm Hg in dogs. During hyper-, normo-, and hypoventilation, mean difference (bias) and precision (limits of agreement [bias +/- 2 SD]) between tcPCO(2) and PaCO(2) values were 13.2 +/- 10.4 mm Hg (limits of agreement, -7.1 and 33.5 mm Hg) in sheep and 10.6 +/- 10.5 mm Hg (limits of agreement, -9.9 and 31.2 mm Hg) in dogs, respectively. Changes in PaCO(2) induced by different ventilation settings were detected by the tcPCO(2) sensor with a lag (response) time of 4.9 +/- 3.5 minutes for sheep and 6.2 +/- 3.6 minutes for dogs. CONCLUSIONS AND CLINICAL RELEVANCE: The tcPCO(2) sensor overestimated PaCO(2) in sheep and dogs and followed changes in PaCO(2) with a considerable lag time. The tcPCO(2) sensor might be useful for noninvasive monitoring of changes but cannot be used as a surrogate measure for PaCO(2).     

Biochemical and toxicological differences in the modes of action of the benzoylureas

The two insecticidal benzoylurea compounds, diflubenzuron and chlorfluazuron, show large differences in their toxicity against the larvae of insects like the tobacco budworm, Heliothis virescens, or the Egyptian cotton leafworm, Spodoptera littoralis, chlorfluazuron being about 100 times more toxic. This difference is due mainly to a much faster metabolism of diflubenzuron. Its half-life within the larvae is about 5 h, compared to about 50 h for chlorfluazuron. Chlorfluazuron is also the much better ovicide of the two, following injection of the compounds into the females of H. virescens. Again the difference in the rate of metabolism is the main cause. The rate of excretion of the parent benzoylureas is relatively low, but their metabolites are excreted very quickly.     

Restriction endonuclease and monoclonal antibody analysis of Brazilian isolates of bovine herpesviruses types 1 and 5

Twelve Brazilian isolates and three reference strains of bovine herpesviruses (BHVs) were subjected to restriction endonuclease analysis (REA) and monoclonal antibody (MAb) analysis. Viral DNA was cleaved with BamHI, BstEII, EcoRI, HindIII and PstI. The monoclonal antibody panel allowed the differentiation between types 1 and 5 viruses, while REA with BstEII and HindIII showed the distinction between BHV-1 and -5 subtypes. Typical 1.1 and 1.2a patterns were observed with two isolates from respiratory disease. An isolate from semen of a clinically healthy bull displayed 1.2b profile, whereas another displayed a clear 5a pattern, which was never reported before. Seven out of nine Brazilian type 5 (BHV-5) isolates displayed REA patterns similar to the Australian BHV-5 strain N569 (BHV-5a), and differing from the Argentinean A663 strain (BHV-5b) virus. Another two BHV-5 isolates, which displayed an unusual MAb pattern of reactivity, showed a BstEII profile different from both reference strains of BHV-5. These two viruses were considered BHV-5 "non-a/non-b" subtype.     

Evaluation of the i-STAT portable clinical analyzer in chickens (Gallus gallus).

The i-STAT portable clinical analyzer (PCA) was evaluated for performance in avian species. With the EG7+ cartridge, which provided results for hydrogen ion concentration, oxygen tension, carbon dioxide tension, sodium, potassium, ionized calcium, hematocrit, and various calculated parameters, analytical accuracy and precision were tested by comparing obtained values to those of established traditional blood gas and chemistry analyzers. Deming's regression and bias plots were used to compare i-STAT results with those obtained by laboratory professionals using benchtop analyzers. The reliability of the i-STAT PCA with EG7+ cartridges was good, with 0-5.7% system failures in measured values. Regression statistics were good for all blood gas analytes and acceptable for electrolytes and calculated parameters, except for potassium and base excess, for which the regression data or the discrepancy between the methods was too large. The system was reliable and easy to use and had an overall acceptable accuracy in avian species. These features, together with portability and small required blood volumes, make the i-STAT suitable for point-of-care use in critical avian patients, although single values require careful interpretation.     

Age effects on Norway spruce (Picea abies) susceptibility to ozone uptake: a novel approach relating stress avoidance to defense

Cumulative ozone (O3) uptake and O3 flux were related to physiological, morphological and biochemical characteristics of Norway spruce (Picea abies (L.) Karst.) trees of different ages. Under ambient CO2 conditions, photosynthetic capacity (Amax) declined in mature trees when cumulative O3 uptake into needles, which provides a measure of effective O3 dose, exceeded 21 mmol m-2 of total needle surface area. A comparable decline in Amax of seedlings occurred when cumulative O(3) uptake was only 4.5 mmol m-2. The threshold O3 flux causing a significant decline in Amax ranged between 2.14 and 2.45 nmol m-2 s-1 in mature trees and seedlings subjected to exposure periods of > or = 70 and > or = 23 days, respectively. The greater O3 sensitivity of young trees compared with mature trees was associated with needle morphology. Biomass of a 100-needle sample increased significantly with tree age, whereas a negative correlation was found for specific leaf area, these changes parallel those observed during differentiation from shade-type to sun-type needles with tree ontogeny. Age-dependent changes in leaf morphology were related to changes in detoxification capacity, with area-based concentrations of ascorbate increasing during tree ontogeny. These findings indicate that the extent of O3-induced injury is related to the ratio of potentially available antioxidants to O3 influx. Because this ratio, when calculated for ascorbate, increased with tree age, we conclude that the ratio may serve as an empirical basis for characterizing age-related differences in tree responses to O3.     

Estimating “autotrophic” belowground respiration in spruce and beech forests: decreases following girdling

Seasonal fluxes of CO₂ from soil and the contribution of autotrophic (root + mycorrhizal) to total soil respiration (SR) were estimated for a mixed stand of European beech (Fagus sylvatica) and Norway spruce (Picea abies) in Central Europe. Mature trees of each species were girdled in August 2002 to eliminate carbohydrate allocation to roots. SR was measured at distances of 0.5, 1.0, and 1.5/2.0 m from the bole of each tree at 1–2 weeks intervals throughout the fall of 2002 and monthly during the spring and summer of 2003. The contribution of roots and mycorrhizae to total SR was estimated by the decrease in SR compared to ungirdled control trees to account for seasonal patterns evident in controls. SR decreased with soil temperature in the fall 2002 and increased again in 2003 as soil warmed. During most of the study period, SR was strongly related to soil temperature. During the dry summer of 2003, however, SR appeared to be uncoupled from temperature and was strongly related to soil water content (SWC). Mean rates of SR in beech and spruce control plots as well as root densities did not show a clear pattern with distance from the bole. SR decreased to levels below controls in beech within a few days after girdling, whereas spruce did not show a significant decrease until October 2002, 6 weeks after girdling. In both beech and spruce, decreased SR in response to girdling was greatest closest to the bole, possibly reflecting increased mycorrhizal activity close to the bole. Autotrophic respiration was estimated in beech to be as much as 50% of the total SR in the stand. The contribution of autotrophic respiration was less certain for spruce, although close to the bole, the autotrophic fraction may contribute to total SR as much as in beech. The large fraction of autotrophic respiration in total SR requires better understanding of tree level stresses that affect carbon allocation below ground.