Antifungal mode of action of triforine

Rapidly growing mycelia of Aspergillus fumigatus treated with 10 μg/ml triforine (N,N′-bis-(1-formamido-2,2,2-trichloroethyl)-piperazine) showed little or no inhibition in dry weight increase prior to 2 h. By 2.5–3 h, triforine inhibited dry weight increase by 85%. The effects of triforine on protein, DNA, and RNA syntheses corresponded to the effect on dry weight increase both in time of onset and magnitude. Neither glucose nor acetate oxidation were inhibited by triforine.Ergosterol synthesis was almost completely inhibited by triforine even in the first hour after treatment. Inhibition of ergosterol synthesis was accompanied by an accumulation of the ergosterol precursors 24-methylenedihydrolanosterol, obtusifoliol, and 14α-methyl-Δ^{8, 24 (28)}-ergostadienol. Mycelia treated with 5 μg/ml of triarimol (α-(2,4-dichlorophenyl)-α-phenyl-5-pyrimidinemethanol) also accumulated the same sterols as well as a fourth sterol believed to be Δ^{5, 7}-ergostadienol.Identification of 4,4-dimethyl-Δ^{8, 24 (28)}-ergostadienol in untreated mycelia indicates that the C-14 methyl group is the first methyl group removed in the biosynthesis of ergosterol by A. fumigatus. The lack of detectable quantities of 4,4-dimethyl-Δ^{8, 24 (28)}-ergostadienol in triforine or triarimol-treated mycelia and the accumulation of C-14 methylated sterols in treated mycelia suggests that both fungicides inhibit sterol C-14 demethylation. The accumulation of Δ^{5, 7}-ergostadienol in triarimol-treated mycelia further implies that triarimol also inhibits the introduction of the sterol C-22(23) double bond.Two strains of Cladosporium cucumerinum tolerant to triforine and triarimol were also tolerant to the fungicide S-1358 (N-3-pyridyl-S-n-butyl-S′-p-t-butylbenzyl imidodithiocarbonate).     

Trichoderma harzianum produces nonanoic acid, an inhibitor of spore germination and mycelial growth of two cacao pathogens

An isolate of Trichoderma harzianum Rifai from an infected cacao pod produces and secretes nonanoic (pelargonic) acid into a liquid culture medium. Nonanoic acid (NA) was very inhibitory to spore germination and mycelial growth of two cacao pathogens, Crinipellis perniciosa Stahel and Moniliophthora roreri Cif. H.C. Evans. It was highly active causing 75% inhibition of spore germination in an in vitro assay at a rate as low as 0.09 μM for M. roreri and 0.92 μM for C. perniciosa. Mycelial growth was comparatively less sensitive to inhibition, but still there was a 75% reduction in growth with 0.62 μM in M. roreri and 151 μM NA in C. perniciosa. In contrast, NA did not affect Trichoderma mycelial growth or spore germination at concentrations that were inhibitory to the pathogens. 6-pentyl-α-pyrone was also produced and secreted into the medium by T. harzianum, however; it was not antagonistic to the cacao pathogens. Although a number of metabolites produced by Trichoderma spp. have been identified in the past, this is the first report of NA production and secretion by any Trichoderma. The results suggest that NA may play a role in the successful use of some Trichoderma spp. isolates in the biocontrol of fungal diseases of plants.     

Distribution and Activity of Fatty Acid Amide Hydralase (FAAH) in the Gastrointestinal Neurons of Mouse

The endocannabinoid anandamide may regulate intestinal motility through activation of CB1 receptors. Anandamide is then inactivated by fatty acid amide hydrolase (FAAH), a membrane bound enzyme. Under pathological conditions, inactivation of such enzymatic activity may lead to inhibition of the intestinal motility. Here, preliminary reports on the distribution of Fatty Acid Amide Hydrolase (FAAH) immunoreactivity in the mouse gastrointestinal neurons, and the pharmacological effects of N‐arachidonoylserotonin (AA‐5HT), a selective inhibitor of FAAH, are reported. FAAH was revealed by an indirect immunofluorescence. Laminar preparations containing the myenteric or the submucous plexus adhered, were peeled off after the whole gut wall had been stretched out and fixed in 4% paraformaldehyde. They were subsequently incubated with a polyclonal anti‐serum directed against a region near the N‐terminus of the human FAAH and revealed by a FITC‐conjugated goat anti‐rabbit secondary anti‐serum. FAAH‐immunoreactive neurons were observed within the myenteric ganglia throughout the GIT. The positive nerve cells varied in size and density of immunoreactivity. Stomach and large intestine showed the highest neuronal density. AA‐5HT significantly reduced both gastric emptying and gastrointestinal tract transit. Such inhibitory effect was reduced by the C1 receptor antagonist SR141716A. Both morphological and pharmacological results suggest that FAAH may play a critical role in controlling gut anandamide levels.     

Serodiagnosis of Burkholderia mallei Infections in Horses: State‐of‐the‐art and Perspectives

Burkholderia mallei causes glanders or farcy in solipeds, a disease that must be reported to the OIE (Office International des Epizooties, Paris, France). The number of reported outbreaks has increased steadily during the last decade. Serodiagnosis is hampered by the considerable number of false‐positives and ‐negatives of the internationally prescribed tests. The major problem leading to low sensitivity and specificity of complement fixation test (CFT) and enzyme‐linked immunosorbent assay (ELISA) has been linked to the test antigens currently used, i.e. crude preparations of whole cells. Future perspectives for the development and evaluation of serological test kits using well‐characterized single antigens are discussed in the light of recent molecular research on B. mallei and the closely related saprozoonotic agent B. pseudomallei.     

Prepenetration stages in infection of clematis by Phoma clematidina

A detailed study of conidial germination, germ-tube growth and the formation of infection structures in Phoma clematidina , the causal agent of clematis wilt, is described for two clematis varieties differing in disease resistance. On both the resistant and susceptible varieties, the fungus entered leaves and stems by direct penetration of the cuticle, often, but not always, following the formation of infection structures. More germ tubes per conidium were formed on the susceptible host, but these germ tubes were on average shorter than on the resistant host. Although germ tubes regularly entered the plant via trichomes, stomata were not found to be sites of entry. Following penetration of the cuticle of resistant plants, germ-tube growth was sometimes restricted to the subcuticular region, and halo formation occurred at the sites where penetration was attempted. Subcuticular growth and halo formation were not observed on susceptible plants. These observations may partly explain the resistance of small-flowered clematis varieties to P. clematidina .