Fusaproliferin production by Fusarium subglutinans and its toxicity to Artemia salina, SF-9 insect cells, and IARC/LCL 171 human B lymphocytes.

Fusarium subglutinans is an important pathogen of maize and other commodities worldwide. We examined MRC-115 and 71 other F. subglutinans strains from various geographic areas for their ability to synthesize fusaproliferin, a novel toxic sesterterpene recently isolated from F. proliferatum. Fusaproliferin production ranged from 30 to 1,500 micrograms/g of dried ground substrate, with 33 strains producing more than 500 micrograms/g. In particular, strain MRC-115 produced as much as 1,100 to 1,300 micrograms/g. In toxicity studies of two invertebrate models, fusaproliferin was toxic to Artemia salina (50% lethal dose, 53.4 microM) and to the lepidopteran cell line SF-9 (50% cytotoxic concentration, approximately 70 microM, after a 48-h exposure). Fusaproliferin was also toxic to the human nonneoplastic B-lymphocyte cell line IARC/LCL 171 (50% cytotoxic concentration, approximately 55 microM in culture in stationary phase after a 48-h exposure). Experiments performed will cells exposed at seeding suggested a possible cytostatic effect at subtoxic concentrations.     

Two new diketopiperazines and a new glucosyl sesterterpene from Alternaria alternata,an endophytic fungi from Ceratostigma griffithii

Two diketopiperazine derivatives, altenarizines A (1) and B (2), and a new glucosyl sesterterpene, 24-α-d-glucosyl-(−)-terpestacin (3), together with two known phytotoxic sesterterpenes, (−)-terpestacin (4) and fusaproliferin (5), were isolated from the fermentation broth of an endophytic fungus Alternaria alternata, which was obtained from the fresh root of Ceratostigma griffithii. Structures of all the isolates were identified by spectroscopic data.     

Effects of temperature, incubation period and substrate on production of fusaproliferin by Fusarium subglutinans ITEM 2404

The kinetics of the production of fusaproliferin by Fusarium subglutinans ITEM 2404 in maize and rice cultures was investigated at various incubation temperatures. The growth rate of F. subglutinans was highest at 20 degrees C and 25 degrees C in maize cultures and at 15 degrees C in rice cultures. Although the growth rate was higher in rice than in maize, the maximal production of fusaproliferin was obtained in maize cultures, with a maximum yield (4309 microg g(-1)) at 20 degrees C for 6 weeks. In rice cultures the optimal incubation regimen was at 15 degrees C for 6 weeks, with a fusaproliferin level of 1557 microg g(-1). The production of fusaproliferin at 25 degrees C and 30 degrees C in both substrates was very low, with maximal yield at 25 degrees C of 979 microg g(-1) after 2 weeks and 143 microg g(-1) after 3 weeks in maize and rice cultures, respectively.     

Inhibition of infection of the rice blast fungus by halisulfate 1, an isocitrate lyase inhibitor

Halisulfate 1, a sesterterpene sulfate and an isocitrate lyase (ICL) inhibitor that is isolated from tropical sponge Hippospongia spp., reduces both appressorium formation and infection of rice plants by the fungus Magnaporthe grisea. Rice plants infected with wild-type M. grisea Guy 11 exhibited significantly lower disease severity after halisulfate 1 treatment than without, and the treatment effect was comparable to the behavior of the Delta icl knockout mutant I-10. The protection observed upon applying halisulfate 1 to rice plants suggests that the ICL inhibitor may be a promising candidate for crop protection, particularly to protect rice plants against M. grisea.     

Species diversity of and toxin production by Gibberella fujikuroi species complex strains isolated from native prairie grasses in Kansas

Fusarium species from agricultural crops have been well studied with respect to toxin production and genetic diversity, while similar studies of communities from nonagricultural plants are much more limited. We examined 72 Fusarium isolates from a native North American tallgrass prairie and found that Gibberella intermedia (Fusarium proliferatum), Gibberella moniliformis (Fusarium verticillioides), and Gibberella konza (Fusarium konzum) dominated. Gibberella thapsina (Fusarium thapsinum) and Gibberella subglutinans (Fusarium subglutinans) also were recovered, as were seven isolates that could not be assigned to any previously described species on the basis of either morphological or molecular characters. In general, isolates from the prairie grasses produced the same toxins in quantities similar to those produced by isolates of the same species recovered from agricultural hosts. The G. konza isolates produce little or no fumonisins (up to 120 micro g/g by one strain), and variable but generally low to moderate amounts of beauvericin (4 to 320 micro g/g) and fusaproliferin (50 to 540 micro g/g). Toxicity to Artemia salina larvae within most species was correlated with the concentration of either beauvericin or fusaproliferin produced. Organic isolates from some cultures of G. moniliformis were highly toxic towards A. salina even though they produced little, if any, beauvericin or fusaproliferin. Thus, additional potentially toxigenic compounds may be synthesized by G. moniliformis strains isolated from prairie grasses. The Fusarium community from these grasses appears to contain some species not found in surrounding agricultural communities, including some that probably are undescribed, and could be capable of serving as a reservoir for strains of potential agricultural importance.     

The biosynthesis of 23,24-dinor-5-cholene-3beta,20-diol and 23,24-dinor-5-cholene-3beta,21-diol

An alternative pathway for steroidogenesis, via a sesterterpene, has been proposed. This communication presents evidence that two of the proposed compounds with the 23,24-dinorcholane carbocyclic system, 23,24-dinor-5-cholene-3β,20-diol and 23,24-dinor-5-cholene3β,21-diol, can be biosynthesised from sodium [³H]acetate in a bovine adrenal preparation.     

Biosynthesis of adrenal corticosteroids by the sesterterpene pathway via 23,24-dinor-4-cholen-3-one

Rat adrenal gland preparations were incubated with 23,24-dinor-5-cholen-3 beta-ol (Guneribol), a proposed intermediate in the sesterterpene pathway for steroid biosynthesis. Steroids were isolated, purified by thin layer and high-performance liquid chromatographies and crystallized to constant specific activity. These preparations converted the substrate to 23,24-dinor-4-cholen-3-one. Radioactive 23,24-dinor-4-cholen-3-one was synthesised and incubated with further tissue preparations and shown to be converted to corticosteroids. These findings suggest that 23,24-dinor-4-cholen-3-one is an intermediate on the sesterterpene pathway for steroidogenesis.     

Species Diversity of and Toxin Production by Gibberella fujikuroi Species Complex Strains Isolated from Native Prairie Grasses in Kansas

Fusarium species from agricultural crops have been well studied with respect to toxin production and genetic diversity, while similar studies of communities from nonagricultural plants are much more limited. We examined 72 Fusarium isolates from a native North American tallgrass prairie and found that Gibberella intermedia (Fusarium proliferatum), Gibberella moniliformis (Fusarium verticillioides), and Gibberella konza (Fusarium konzum) dominated. Gibberella thapsina (Fusarium thapsinum) and Gibberella subglutinans (Fusarium subglutinans) also were recovered, as were seven isolates that could not be assigned to any previously described species on the basis of either morphological or molecular characters. In general, isolates from the prairie grasses produced the same toxins in quantities similar to those produced by isolates of the same species recovered from agricultural hosts. The G. konza isolates produce little or no fumonisins (up to 120 μg/g by one strain), and variable but generally low to moderate amounts of beauvericin (4 to 320 μg/g) and fusaproliferin (50 to 540 μg/g). Toxicity to Artemia salina larvae within most species was correlated with the concentration of either beauvericin or fusaproliferin produced. Organic isolates from some cultures of G. moniliformis were highly toxic towards A. salina even though they produced little, if any, beauvericin or fusaproliferin. Thus, additional potentially toxigenic compounds may be synthesized by G. moniliformis strains isolated from prairie grasses. The Fusarium community from these grasses appears to contain some species not found in surrounding agricultural communities, including some that probably are undescribed, and could be capable of serving as a reservoir for strains of potential agricultural importance.     

The conversion of 23,24-dinor-5-cholen-3β-ol to cortisol by the canine adrenal

An alternative pathway for steroidogenesis, via a sesterterpene, has been proposed. This communication presents evidence that the canine adrenal can utilise 23,24-dinor-5-cholen-3β-ol to biosynthesise cortisol.It has been proposed that steroid hormones could be biosynthesised by a pathway other than that through cholesterol, possibly by a sesterterpene pathway (1,2).The previous studies were carried out using bovine adrenal tissue. This communication extends these studies to include the canine adrenal gland.     

Enzymatic cyclization of all-trans pentaprenyl and hexaprenyl methyl ethers by a cell-free system from the protozoon Tetrahymena pyriformis. The biosynthesis of scalarane and polycyclohexaprenyl derivatives

A cell-free system from the protozoon Tetrahymena pyriformis capable of cyclizing squalene into tetrahymanol cyclizes all-trans pentaprenyl methyl ether to a scalarane-type sesterterpene and all-trans hexaprenyl methyl ether to bicyclo-, tricyclo-, tetracyclo- and pentacyclohexaprenyl methyl ethers, each corresponding to a possible cationic intermediate. The structures of the cyclization products have been determined by spectroscopic methods and are compatible with a biogenetic scheme involving polyprenyl ether cyclization. This is the first direct proof of an enzymatic cyclization of higher isoprenic alcohol derivatives, and we assume it was performed by the squalene-to-hopane cyclase of the protozoon. The formation of a scalarane-type sesterterpene from C25 polyprenyl methyl ether suggests that these terpenoids, whose presence is restricted to a few sponges, might be in fact microbial metabolites. Tricyclopolyprenyl derivatives have been identified in the organic matter from numerous sediments and they were interpreted as being chemical fossils of still unidentified microorganisms. The cyclization of hexaprenyl methyl ether is the first attempt of identification of these tricyclopolyprenol derivatives in living organisms.     

Adrenal androgen biosynthesis by a sesterpene pathway

Rat and human adrenal gland preparations were incubated with radioactive cholesterol and 23,24-dinor-5-cholen-3 beta-ol, the latter being a proposed intermediate in the sesterterpene pathway for steroid biosynthesis. Steroids were isolated, purified by TLC and crystallised to constant specific activity. It was found that rat and human adrenal glands can utilise 23,24-dinor-5-cholen-3 beta-ol to produce androstenedione and dehydroepiandrosterone. Also, it was found that the conversion of 23,24-dinor-5-cholen-3 beta-ol to androgens occurs in the microsomal fraction. It was concluded that the sesterterpene pathway for steroid biosynthesis can function in the rat and human adrenal glands to produce androgens and that the intermediates are converted to androgens in the microsomal fraction.     

Inhibition of Candida albicans isocitrate lyase activity by sesterterpene sulfates from the tropical sponge Dysidea sp

Seven sesterterpene sulfates (1-7) were isolated from the tropical sponge Dysidea sp. and their inhibitory activities against isocitrate lyase (ICL) from Candida albicans were evaluated. Among the isolated natural products compound 6 and 7 were found to be strong ICL inhibitors. The isolated compounds (1-7) also showed potent antibacterial effect against Bacillus subtilis and Proteus vulgaris, but did not display antifungal activity.     

Biosynthesis of androgens by the sesterterpene pathway via 23,24-dinor-4-cholen-3-one

Rat testicular and adrenal gland microsomal preparations were incubated with 23,24-dinor-5-cholen-3β-ol (Guneribol) a proposed intermediate in the sesterterpene pathway for steroid biosynthesis. Steroids were isolated, purified by thin layer and high-performance liquid chromatography and crystallized to constant specific radioactivity. These preparations converted the substrate to 23,24-dinor-4-cholen-3-one. Radioactive 23,24-dinor-4-cholen-3-one was synthesized and incubated with further tissue preparations and shown to be converted to steroid hormones. These findings suggest that 23,24-dinor-4-cholen-3-one is an intermediate on the sesterterpene pathway for steroidogenesis.     

Production of beauvericin,moniliformin, fusaproliferin,and fumonisins b(1), b(2), and b(3) by fifteen ex-type strains of fusarium species

Fifteen Fusarium species were analyzed by high-performance liquid chromatography for the production of six mycotoxins in corn grits cultures. Production of mycotoxins ranged from 66 to 2,500 micro g/kg for fumonisin B(1), 0.6 to 1,500 micro g/g for moniliformin, 2.2 to 720 micro g/g for beauvericin, and 12 to 130 micro g/g for fusaproliferin. Fumonisin B(2) (360 micro g/kg) was produced by two species, fumonisin B(3) was not detected in any of the 15 species examined, and Fusarium bulbicola produced none of the six mycotoxins that we analyzed.     

丝状真菌二倍半萜化合物及其合成酶

相比其他萜类化合物,丝状真菌来源的二倍半萜化合物数量较少,但具有广泛的生理活性和药用价值。已经发现的丝状真菌二倍半萜合成酶均由萜类环化酶和异戊烯基转移酶两个结构域组成,表现出底物的非特异性和环化方式的多样性。本文重点叙述了丝状真菌来源的二倍半萜化合物以及其合成酶的结构与功能特征,并对丝状真菌二倍半萜化合物及其合成酶研究现状作简要概述。     

A new neolignan and a new sesterterpenoid from the stems of Picrasma quassioides Bennet

A new dihydrobenzofuran-type neolignan, picrasmalignan A (1), and a new sesterterpene lactone, 2'-isopicrasin A (4), were isolated from the stems of Picrasma quassioides Bennet, along with four known compounds, comprising two neolignans, 2 and 3, a sesterterpene lactone, 5, and a flavonol, 6. The structures of these compounds were determined by detailed analysis of NMR and MS data, and comparison with the literature data. Compounds 1-6 were tested for their anti-inflammatory activity, and 1-3 and 6 showed potent inhibitory activities on nitric oxide, tumor necrosis factor-α, and interleukin-6 production in mouse monocyte-macrophage RAW 264.7 stimulated by lipopolysaccharide (LPS).     

Occurrence of Fusaproliferin and Beauvericin in Fusarium-Contaminated Livestock Feed in Iowa

Fusarium fungal contaminants and related mycotoxins were investigated in eight maize feed samples submitted to the Iowa State University Veterinary Diagnostic Laboratory. Fusarium moniliforme, F. proliferatum, and F. subglutinans were isolated from seven, eight, and five samples, respectively. These strains belonged to mating populations A, D, and E of the teleomorph Gibberella fujikuroi. Fusaproliferin was detected at concentrations of 0.1 to 30 μg/g in four samples, and beauvericin was detected (0.1 to 3.0 μg/g) in five samples. Fumonisins were detected in all eight samples (1.1 to 14 μg/g). Ten of 11 strains of F. proliferatum and all 12 strains of F. subglutinans isolated from the samples produced fusaproliferin in culture on whole maize kernels (4 to 350 and 100 to 1,000 μg/g, respectively). Nine F. proliferatum strains also produced beauvericin in culture (85 to 350 μg/g), but none of the F. subglutinans strains produced beauvericin. Fumonisin B₁ was produced by all nine F. moniliforme strains (50 to 2,000 μg/g) and by 10 of the F. proliferatum strains (1,000 to 2,000 μg/g). This is the first report of the natural occurrence of fusaproliferin outside Italy and of the natural occurrence of beauvericin in North America.     

Sesterterpene sulfates as isocitrate lyase inhibitors from tropical sponge Hippospongia sp

Two sesterterpene sulfates (1-2) were isolated from tropical sponge Hippospongia sp. and their inhibitory activities against isocitrate lyase (ICL) from the rice blast fungus Mgnaporthe grisea were evaluated. Compound 3 was obtained by hydrolysis of compound 1. Compounds 1 and 3 were found to be potent ICL inhibitors, which inhibited appressorium formation and C(2) carbon utilization in M. grisea. Our results suggest that ICL plays crucial role in appressorium formation of M. grisea and is a new target for the protection of rice blast disease.     

Effect of<Emphasis Type="Italic">Trichoderma</Emphasis> species on growth of Fusarium<Emphasis Type="Italic">proliferatiom</Emphasis> and production of fumonisins,fusaproliferin and beauvericin

Trichoderma species has been suggested as potential biocontrol agent forFusarium verticillioides on maize. In this cereal,F. verticillioides and F. proliferatum contributed to fumonisin accumulation. In addition,F. proliferatum could produce beauvericin and fusaproliferin. The aim of this work was to evaluate the effect ofTrichoderma spp. on growth and fumonisin B¹ fusaproliferin and beauvericin production byF. proliferatum. Dual cultures of F.proliferatum andT. harzianum ITEM 3636 andT. longibrachiatum ITEM 3635 on maize meal agar at 0.995 aw were done. The effect ofTrichoderma spp. on the lineal growth ofF. proliferatum was determined. The effect ofTrichoderma species on fumonisin B¹, fusaproliferin and beauvericin production byF. proliferatum was determined on co-inoculated maize kernels by HPLC.T. harzianum suppressedF. proliferatum growth once contact between the colonies occurred.T. longibrachiatum showed a less antagonistic effect againstF. proliferatum. A reduction on fumonisin B¹ production of 98% and 88% was observed in the co-incubation ofF. proliferatum withT. harzianum andT. longibrachiatum, respectively. The decrease of FB¹ production was significant even in maize kernels on whichF. proliferatum had been growing 7 days prior to the addition ofTrichoderma spp. The concentration of beauvericin and fusaproliferin produced during 30 days coincubation ofF. proliferatum with bothTrichoderma spp. did not differ to those produced byF. proliferatum alone. These mycotoxins might enter the food chain causing so far unknown consequences to the health of domestic animals and humans. For this reason it is important, when a potential biocontrol agent is under study, to test the effect on the fungal growth and on the putative mycotoxin produced. Part of the information was presented at the Mycotoxin Prevention Cluster Dissemination Day and Mycoglobe Launch Conference, Brussels, Belgium, Oct 20–21, 2004 Financial support: Agenda Córdoba Ciencia, grant N^o 0279–000431/00