Natural occurrence of the mycotoxin penitrem a in moldy cream cheese

The occurrence of the mycotoxin penitrem A in refrigerated cream cheese that was involved in the intoxication of two dogs is described. Penitrem A and the fungus Penicillium crustosum were isolated from the cream cheese and identified. This is believed to be the first definitive case of the natural occurrence of penitrem A.     

Moldy walnut toxicosis in a dog,caused by the mycotoxin,penitrem A

Penitrem A was found in moldy walnuts that were involved in an intoxication of a dog in California.Penicillium crustosum was isolated from the walnuts and penitrem A was isolated from extracts of the mycelium of this fungus when it was grown on a synthetic medium.     

Moldy walnut toxicosis in a dog, caused by the mycotoxin, penitrem A

Penitrem A was found in moldy walnuts that were involved in an intoxication of a dog in California.Penicillium crustosum was isolated from the walnuts and penitrem A was isolated from extracts of the mycelium of this fungus when it was grown on a synthetic medium.     

Tremorgenic toxins produced by soil fungi.

Penitrem A or an unknown tremorgenic toxin, "X," was produced by 10 of 60 fungal isolates obtained from a pasture involved in an outbreak in cattle and sheep resembling migram and ryegrass staggers. Tremorgenic properties of extracts containing penitrem A or toxin X were confirmed by bioassay.     

Biosynthesis of penitrems and roquefortine by Penicillium crustosum

Roquefortine and the penitrems were biosynthesised concurrently at an approximately equimolar rate by Penicillium crustosum after growth and sporulation. [14C]mevalonic acid was incorporated (15% efficiency) into the isoprenoid regions of the penitrem and roquefortine molecules to an extent consistent with their 6:1 molar ratio of isoprenoid components. [14C]penitrem A (specific activity, 3.4 X 10(2) mu Ci mmol-1) and 14C-penitrems B, C, and E readministered to young cultures were metabolically interconverted, indicating considerable metabolic flux, though generally directed towards penitrem A as the end product and suggesting a metabolic grid for the penitrem metabolites. Addition of bromide to the medium preferentially favored the production of bromo-analogs rather than the usual chloropenitrems.     

Biosynthesis of penitrems and roquefortine by Penicillium crustosum.

Roquefortine and the penitrems were biosynthesised concurrently at an approximately equimolar rate by Penicillium crustosum after growth and sporulation. [14C]mevalonic acid was incorporated (15% efficiency) into the isoprenoid regions of the penitrem and roquefortine molecules to an extent consistent with their 6:1 molar ratio of isoprenoid components. [14C]penitrem A (specific activity, 3.4 X 10(2) mu Ci mmol-1) and 14C-penitrems B, C, and E readministered to young cultures were metabolically interconverted, indicating considerable metabolic flux, though generally directed towards penitrem A as the end product and suggesting a metabolic grid for the penitrem metabolites. Addition of bromide to the medium preferentially favored the production of bromo-analogs rather than the usual chloropenitrems.     

Thomitrems A and E,two indole-alkaloid isoprenoids from Penicillium crustosum Thom

Two indole-alkaloid isoprenoids were isolated from extracts of Penicillium crustosum Thom grown on rice. Their structures were elucidated on the basis of various NMR experiments and by comparison to the structurally related penitrems. The two compounds, designated thomitrem A and thomitrem E, contain a 18(19)-double bond and lack the characteristic penitrem 17(18)-ether linkage.     

In vitro neuropharmacological evaluation of penitrem-induced tremorgenic syndromes: importance of the GABAergic system

The effects of the fungal neurotoxin penitrem A on the GABAergic and glutamatergic systems in rat brain were evaluated. Penitrem A inhibited binding of the GABA_A-receptor ligand [³H]TBOB to rat forebrain and cerebellar membrane preparations with IC₅₀ (half maximal inhibitory concentration) values of 11 and 9 μM, respectively. Furthermore, penitrem A caused a concentration-dependent increase of [³H]flunitrazepam and [³H]muscimol binding in rat forebrain, but not in cerebellar preparations. The stimulation of [³H]flunitrazepam binding by penitrem A was abolished by the addition of GABA. In cerebellar preparations, a different pharmacological profile was found, with penitrem A allosterically inhibiting [³H]TBOB binding by interacting with a bicuculline-sensitive site. Moreover, penitrem A inhibited the high affinity uptake of GABA and glutamate into cerebellar synaptosomes with IC₅₀ values of 20 and 47 μM, respectively. The toxin showed no effect on NMDA or AMPA glutamate receptor binding. In conclusion, our results suggest that penitrem A exerts region-specific effects in the brain, leading to positive modulation of GABA_A-receptor function in forebrain. Conversely, penitrem A may act as a bicuculline-like convulsant in cerebellum.     

Tremorgenic mycotoxins produced by strains of Penicillium spp. isolated from toxic Poa huecu parodi

Seventeen strains of Penicillium spp. have been isolated from Poa huecu Parodi from the Zapala zone, exhibiting toxicity to sheet. The following strains have been identified: P. crustosum, cyclopium, notatum, palitans, puberulum, verrucosum, viridicatum and Penicillium spp. The toxigenic capacity of the strains was studied after growing them under suitable conditions. Toxins produced were analysed by thin layer chromatography (TLC). Penitrem A (PA) and Penitrem B (PB) neurotoxins were identified and quantitated in twelve strains; verruculogen (VERR) and fumitremorgen B (FTB) being present in one of them. The effect of these mycotoxins was studied in mice. Neurological symptoms characteristic of the intoxication by tremorgenic toxins and similar to those observed in sheep suffering from huecu's disease were observed. The possible role of these toxins as causative agents of huecu's disease is discussed.     

Effects of penitrem A on rat's performances in passive avoidance and Morris water maze tests

Intraperitoneal administration of the mycotoxin penitrem A 30 min before a training session in passive avoidance task, impaired performance of rats subjected to a test-session 24 h after. This effect was not antagonised by pretraining administration of physostigmine or bicuculline. Administration of penitrem A 20 min before a training session or 30 min before a test-session did not impair performance. In the Morris water maze, doses of penitrem A that induces slight to moderate tremors, but not a lower dose, disrupted place learning. These results suggest that penitrem A disrupts the processes that take place at the time of acquisition, but not those just after acquisition, and does not alter the restitution of information. This effect would not be related to a decrease of cholinergic neurotransmission nor to a stimulation of GABA A receptors. Nevertheless, it could not be totally excluded that the performance impairments induced by penitrem A would be secondary to a motor disruption. This revised version was published online in June 2006 with corrections to the Cover Date.     

Characterization of outward K(+) currents in isolated smooth muscle cells from sheep urethra

The perforated-patch techniquewas used to measure membrane currents in smooth muscle cells from sheepurethra. Depolarizing pulses evoked large transient outward currentsand several components of sustained current. The transient current anda component of sustained current were blocked by iberiotoxin, penitremA, and nifedipine but were unaffected by apamin or 4-aminopyridine,suggesting that they were mediated by large-conductanceCa²⁺-activated K⁺ (BK) channels. When the BKcurrent was blocked by exposure to penitrem A (100 nM) andCa²⁺-free bath solution, there remained a voltage-sensitiveK⁺ current that was moderately sensitive to blockade withtetraethylammonium (TEA; half-maximal effective dose = 3.0 ± 0.8 mM) but not 4-aminopyridine. Penitrem A (100 nM) increasedthe spike amplitude and plateau potential in slow waves evoked insingle cells, whereas addition of TEA (10 mM) further increased theplateau potential and duration. In conclusion, bothCa²⁺-activated and voltage-dependent K⁺currents were found in urethral myocytes. Both of these currents arecapable of contributing to the slow wave in these cells, suggesting that they are likely to influence urethral tone under certain conditions.

     

Classification of terverticillate penicillia based on profiles of mycotoxins and other secondary metabolites

Strains of available terverticillate penicillium species and varieties were analyzed for profiles of known mycotoxins and other secondary metabolites produced on Czapek yeast autolysate agar (intracellular metabolites) and yeast extract-sucrose agar (extracellular metabolites) by using simple thin-layer chromatography screening techniques. These strains (2,473 in all) could be classified into 29 groups based on profiles of secondary metabolites. Most of these profiles of secondary metabolites were distinct, containing several biosynthetically different mycotoxins and unknown metabolites characterized by distinct colors and retardation factors on thin-layer chromatography plates. Some species (P. italicum and P. atramentosum) only produced one or two metabolites by the simple screening methods. The 29 groups based on profiles of secondary metabolites were known species or subgroups thereof. These species and subgroups were independently identifiable by using morphological and physiological criteria. The species accepted, the number of isolates in each species investigated, and the mycotoxins they produced were: P. atramentosum, 4; P. aurantiogriseum, 510 (group I: penicillic acid and S-toxin and group II: penicillic acid, penitrem A [low frequency], terrestric acid [low frequency], viomellein, and xanthomegnin); P. brevicompactum, 81 (brevianamid A and mycophenolic acid); P. camembertii group I, 38, and group II, 114 (cyclopiazonic acid); P. chrysogenum, 87 (penicillin, roquefortine C, and PR-toxin); P. claviforme, 4 (patulin and roquefortine C); P. clavigerum, 4 (penitrem A); P. concentricum group I, 10 (griseofulvin and roquefortine C), and group II, 3 (patulin and roquefortine C); P. crustosum, 123 (penitrem A, roquefortine C, and terrestric acid); P. echinulatum, 13; P. expansum, 91 (citrinin, patulin, and roquefortine C); P. granulatum, 6 (patulin, penitrem A, and roquefortine C [traces]); P. griseofulvum, 21 (cyclopiazonic acid, griseofulvin, patulin, and roquefortine C); P. hirsutum, 100 (group I: terrestric acid; group II: citrinin, penicillic acid , roquefortine C, and terrestric acid; and group III: roquefortine C and terrestric acid), P. hirsutum group IV, 2 (chaetoglobosin C); P. isariiforme, 1; P. italicum, 41; P. mali, 104; P. roquefortii, 78 (group I: mycophenolic acid, PR-toxin, and roquefortine C and group II: mycophenolic acid, patulin, penicillic acid [low frequency], and roquefortine C); P. viridicatum group I, 634 (brevianamid A [low frequency], penicillic acid, viomellein, and xanthomegnin), P. viridicatum group II and III, 494 (citrinin and ochratoxin A), P. viridicatum group IV, 12 (griseofulvin and viridicatumtoxin). It is proposed that profiles of secondary metabolites be strongly emphasized in any future revision of the penicillia.     

Physiological criteria and mycotoxin production as AIDS in identification of common asymmetric penicillia

The taxonomy of the asymmetric (predominantly terverticillate) penicillia is based on morphological differences that leave identification difficult. The application of physiological criteria facilitated the identification of the common asymmetric penicillia investigated. Changes in the placement of some strains of these penicillia made the connection to mycotoxin-producing ability clearer. The classical criterion of conidium color was deemphasized and replaced by the following criteria: (i) growth on nitrite-sucrose agar and (ii) growth and acid (and subsequent base) production on creatine-sucrose agar (containing bromocresol purple). Other criteria used or developed were: (iii) growth on sorbic acid plus benzoic acid agar (50 + 50 ppm, pH 3.8), (iv) growth on an agar containing 1,000 ppm propionic acid (pH 3.8), (v) growth on an agar containing 0.5% acetic acid, (vi) growth at 37 degrees C, (vii) growth rate on an agar containing 0.1% pentachloronitrobenzene, (viii) production of extracellular tricaproinase, and (ix) fasciculation on a medium containing 10 ppm botran (2,6-dichloro-4-nitroanilin). The pattern of extracellular metabolites after thin-layer chromatography was used as a chemotaxonomic criterion. The species investigated, the number of isolates investigated, and the toxins which some of these isolates produce were: Penicillium roqueforti (18) (patulin), P. citrinum (11) (citrinin), P. patulum (9) (patulin and griseofulvin), P. expansum (patulin and citrinin), P. hirsutum (13), P. brevicompactum (19), and P. chrysogenum (12). Widespread species of the P. cyclopium, P. viridicatum, and P. expansum series of Raper and Thom (A Manual of the Penicillia, 1949) were subdivided into four new groups: "P. crustosum pA" (29) (penitrem A), "P. melanochlorum" (29), "P. cyclopium p" (119) (penicillic acid and infrequently penitrem A), and "P. viridicatum o-c" (43) (ochratoxin A and citrinin). "P. viridicatum o-c" was separated from "P. cyclopium p" due to its ability to grow on nitrite as sole nitrogen source. The species and groups investigated were related to the new taxonomic classification of the genus Penicillium according to Pitt.     

Experimental intoxication of guinea pigs with multiple doses of the mycotoxin,penitrem A

Young female guinea pigs were fed various doses of penitrem A every 3 days for 3 weeks. Guinea pigs fed penitrem A had muscle tremors, seizures, and ataxia, and total weight gains were less than those of control guinea pigs. Histologic examination of multiple tissues and electron microscopic examination of liver and kidney revealed no differences between guinea pigs fed penitrem A and controls. Sera harvested at necropsy from guinea pigs fed penitrem A and control guinea pigs did not differ significantly in mean values of two liver enzymes (ornithine carbamoyltransferase and sorbital dehydrogenase), complement, total protein, and differential proteins (albumin; ₁, ₂,, and gobulins). Results of this study indicate that penitrem A causes only central nervous system dysfunction; evidence of cytotoxicity for extraneural tissues was not found.     

南方红豆杉内生真菌的抗肿瘤产物震颤毒素的研究

从红豆杉内生真菌青霉菌Penicillum sp. H⑥.1发酵产物中自然析出一种化合物,经图谱检测和文献比对,鉴定为震颤毒素Penitrem A。四唑盐（MTT）比色法研究表明Penitrem A对恶性黑色素瘤细胞A375有一定的抑制作用,且抑制作用随浓度的增大而增强,IC₅₀为27.24μg/mL。本研究是首次从红豆杉内生真菌发酵产物中分离得到震颤毒素Penitrem A,为抗肿瘤药物开发提供了理论参考。     

Penitrem A and Roquefortine Production by Penicillium commune

Extracts of Penicillium commune, a fungus isolated from cottonseed, showed biological activity in day-old cockerels. Two neurotoxic metabolites were isolated and identified as penitrem A and roquefortine. This is the first report of roquefortine being produced by a fungus other than Penicillium roqueforti as well as the first report of penitrem A and roquefortine being produced in the same culture. Production of these toxins on liquid media and cottonseed was determined.     

Actions of Tremorgenic Fungal Toxins on Neurotransmitter Release

The neurochemical effects of the tremorgenic mycotoxins Verruculogen and Penitrem A, which produce a neurotoxic syndrome characterised by sustained tremors, were studied using sheep and rat synaptosomes. The toxins were administered in vivo, either by chronic feeding (sheep) or intraperitoneal injection 45 min prior to killing (rat), and synaptosomes were subsequently prepared from cerebrocortical and spinal cord/medullary regions of rat, and corpus striatum of sheep. Penitrem A (400 mg mycelium/kg) increased the spontaneous release of endogenous glutamate, GABA (gamma-aminobutyric acid), and aspartate by 213%, 455%, and 277%, respectively, from cerebrocortical synaptosomes. Verruculogen (400 mg mycelium/kg) increased the spontaneous release of glutamate and aspartate by 1300% and 1200%, respectively, but not that of GABA from cerebrocortical synaptosomes. The spontaneous release of the transmitter amino acids or other amino acids was not increased by the tremorgens in spinal cord/medullary synaptosomes. Penitrem A pretreatment reduced the veratrine (75 microM) stimulated release of glutamate, aspartate, and GABA from cerebrocortical synaptosomes by 33%, 46%, and 11%, respectively, and the stimulated release of glycine and GABA from spinal cord/medulla synaptosomes by 67% and 32% respectively. Verruculogen pretreatment did not alter the veratrine-induced release of transmitter amino acids from cerebrocortex and spinal cord/medulla synaptosomes. Penitrem A pretreatment increased the spontaneous release of aspartate, glutamate, and GABA by 68%, 62%, and 100%, respectively, from sheep corpus striatum synaptosomes but did not alter the synthesis and release of dopamine in this tissue. Verruculogen was shown to cause a substantial increase (300-400%) in the miniature-end-plate potential (m.e.p.p.) frequency at the locust neuromuscular junction. The response was detectable within 1 min, rose to a maximum within 5-7 min, and declined to the control rate over a similar period. No change in the amplitude of the m.e.p.p.'s was observed. These effects of the tremorgens on transmitter release are interpreted in terms of their mode of action.     

Ethylene production by soil microorganisms.

Ethylene-producing strains of Penicillium cyclopium and P. crustosum were isolated from soil. These isolates produced ethylene on a variety of carbon growth substrates including phenolic acids. The quantities of ethylene produced on the various substrates varied, and the subtrate-ethylene prosuction pattern for P. cyclopium strains differed significantly from that of P. crustosum strains.     

A novel process for the production of penitrem mycotoxins by submerged fermentation of Penicillium nigricans

A strain of Penicillium nigricans, which produces both the antifungal antibiotic griseofulvin and tremorgenic penitrem mycotoxins concurrently in static liquid culture, also elaborated both metabolites in submerged culture when stimulated by calcium chloride to sporulate. Maximum yield of penitrems (60 mg l-1) occurred within 5 d in a 60 l stirred fermenter, thus constituting the first significant process for penitrem production in submerged culture.     

Alternariol derivatives from Alternaria alternata,an endophytic fungus residing in red sea soft coral,inhibit HCV NS3/4A protease

Applied Biochemistry and Microbiology - Three fungal strains, Alternaria alternata, Eurotium chevalieri and Penicillium crustosum were isolated from the normal tissues of the soft coral Litophyton...