The occurrence of Fusarium varieties and their mycotoxins in silo corn. 2. The formation of zearalenone in the field by artificial infection of silo corn with Fusarium culmorum (W. G. Smith) Sacc

The formation of zearalenone in a maize plot artificially infected with Fusarium culmorum was studied. The zearalenone concentration steeply increased only in the 8th week after inoculation and reached a maximum value of ca. 7 ppm, whereas zearalenone could not be detected in the control variants. The crude nutrient and dry matter content was not significantly influenced by the fungal infection. The infected crop showed average ear dry weights distinctly lower than that of the control variants (P less than or equal to 0.001). Apart from zearalenone, the mycotoxin deoxynivalenol was qualitatively detected in the infected maize. The toxicological relevance of the ascertained zearalenone content with regard to the health of dairy cattle and pigs was discussed.     

The occurrence of Fusarium varieties and their mycotoxins on silo corn. 1. A method for the determination of zearalenone in corn and corn silage by high performance liquid chromatography (HPLC) with fluorescence detection

A method for the determination of zearalenone in maize and maize silage was developed which distinguishes itself by the effective and fast cleaning of the extracts with the help of a silica gel minicolumn. The samples were extracted with chloroform/methanol (9 + 1) and cleaned on a silica gel minicolumn after acid-base partition. The zearalenone was quantitatively determined optionally by means of high-performance liquid chromatography (HPLC) with fluorescence detection (excitation wavelength 236 nm, emission filter 418 nm) or thin-layer chromatography (TLC), p-methoxybenzene diazonium fluoroborate and aluminium chloride were used as detection chemicals. The limits of detection are 0.01 mg/kg (HPLC) and 0.1 mg/kg resp. (TLC), the average recovery is 81%. The method was used for the determination of zearalenone in grain maize, CCM silage and silage from whole maize plants.     

The occurrence of Fusarium strains and their mycotoxins in corn silage. 4. The occurrence of zearalenone in the first cut zone of horizontal silos

Studies of the occurrence of zearalenone in the cutting surface of a horizontal silo were carried out over 12 days. On all sampling days the silage contained less than 0.1 mg/kg zearalenone (detection limit of the thin-layer chromatographic procedure). Zearalenone could not be detected in silage put into interim store over a period of 17 days either. The results document that zearalenone-producing Fusarium species have no considerable share in the decay of maize silage.     

Occurrence of Fusarium strains and their mycotoxins in corn silage. 7. Formation of deoxynivalenol (DON) in a silage corn plot artificially inoculated with Fusarium culmorum and the effect of silaging on the stability of the DON formed

The formation of deoxynivalenol in a maize plot inoculated with Fusarium culmorum was studied over a growing season. Already three weeks after inoculation 4.9 mg/kg of DON were measured in the infected ears. The toxin concentration increased continuously up to harvest after eight weeks to a value of 261 mg/kg. Ensilage experiments in laboratory scale silos have shown that the DON content of naturally contaminated corn-cob-mix was not reduced during the ensilage process. It was concluded that infection of maize plants by toxin-producing Fusarium species followed by DON production in the field seems to be the most probable way of contamination of maize silage with this mycotoxin.     

Occurrence of Fusarium species and their mycotoxins in corn silage. 5. Fusarium infestation in corn silage

The internal mycoflora of silage maize plants was examined in several years. Average Fusarium infection of the maize plants was 10.4% and leaf sheaths and stalks were preferably colonized by the fungi. During the growing season an increase in total colonization of the maize plant by field fungi as well as by species of the genus Fusarium was observed 17 different Fusarium species were isolated from silage maize plants. Most frequently occurring species (more than 85%) in descending order were F. avenaceum, F. culmorum, F. equiseti, F. sporotrichioides, F. crookwellense, F. oxysporum, F. sambucinum var. coeruleum.     

Occurrence of Fusarium strains and their mycotoxins on corn silage. 6. Formation of zearalenone and trichothecenes (type A) by indigenous Fusarium isolates

A total of 399 indigenous Fusarium strains mainly isolated from silage maize were tested for the production of zearalenone and type A trichothecenes by thin-layer chromatography and biological assays. About 45% of the isolates examined were capable of producing different levels of zearalenone and trichothecene toxins on a cracked corn substrate. The majority of these strains (75%) produced zearalenone only and no trichothecenes type A. The results of the biological tests indicated a higher rate of toxin-positive extracts than chemical analysis. Isolates of nine out of seventeen Fusarium species examined produced one or several mycotoxins looked for. The most important toxin producers were F. culmorum and F. crookwellense (zearalenone) and F. sporotrichioides (trichothecenes type A), respectively. F. avenaceum, the species most frequently isolated from silage maize, produced neither zearalenone nor trichothecenes but avenacein Y a antibiotic compound. First results of a study of the production of type B trichothecenes have shown that indigenous F. culmorum isolates were capable of producing high levels of deoxynivalenol.     

Studies on the application of enzyme immunoassays for the Fusarium mycotoxins deoxynivalenol, 3-acetyldeoxynivalenol, and zearalenone.

Polyclonal antisera against zearalenone (ZEA) were produced in rabbits after immunization with ZEA-oxime coupled to human serum albumin. Using these antibodies and a ZEA-oxime-horseradish peroxidase conjugate in a competitive direct enzyme immunoassay (EIA), the detection limit for ZEA was 70 pg/ml. The relative cross-reactivities of the assay with ZEA, alpha-zearalenol, beta-zearalenol, zearalanone, alpha-zearalanol, and beta-zearalanol, respectively, were 100%, 37.3%, 7.2%, 59.2%, 5.3%, and 3.9%, respectively. This EIA and two EIAs for deoxynivalenol (DON) and 3-acetyldeoxynivalenol(3-AcDON) (Usleber et al., 1991) were used to analyze wheat samples. The limits of determination for DON, 3-AcDON, and ZEA in wheat were 200 ppb, 50 ppb, and 20 ppb, respectively. The analysis of reference materials (wheat flour) containing DON by EIA showed good agreement with the nominal values. The EIA for ZEA was in addition used to analyze biological fluids, obtained during a feeding trial. Two lactating cows were administered 25 mg and 100 mg ZEA per day, respectively, over a period of 6 days. Serum, milk, urine, and feces were assayed in the ZEA-EIA with and without sample treatment with beta-glucuronidase prior to the analysis. Maximum toxin levels (ZEA-equivalents) found in milk were 0.4 and 1.2 ppb (glucuronides). The toxin concentration in milk decreased rapidly after the last toxin administration. In the urine, maximum levels of toxin-glucuronide conjugates were 23 ppb and 24 ppb, respectively. The serum toxin levels corresponded to those found in milk. In the feces, mean values were 150 ppb and 500 ppb, respectively, no conjugated toxins were found in feces.     

Effect of Fusarium roseum corn culture containing zearalenone on early pregnancy in swine

A corn culture of Fusarium roseum was added to a standard corn-soybean swine gestation ration. Low, middle, and high dosage mixed feeds contained 7, 38, and 64 mg of zearalenone/kg of feed (7, 38, and 64 ppm) and 0.5, 2.5, and 4.5 mg of deoxynivalenol/kg, respectively. Control feed was the standard ration without added F roseum corn culture. Mature gilts were bred by natural service and fed control or F roseum molded feed from 3 to 34 days after breeding. The main effect of the molded feed was an inhibition of fetal development, with decreased numbers of fetuses present in treated animals at slaughter (38 to 43 days after breeding). Normal litters were present in 7 of 8 control animals, in 2 of 4 gilts given the low-dosage feed, in 1 of 4 gilts given the medium dosage, and in 0 of 4 given the high-dosage feed. Corpora lutea were maintained in all treated animals, as evidenced by serum progesterone concentrations. Serum estradiol concentrations were decreased in gilts in the middle- and high-dosage groups. The genital system of the gilts fed low- and middle-dosage feeds had a gross and microscopic appearance similar to that of the pregnant controls and reflected prolonged progesterone stimulation. Morphologic changes in the genital system of the high-dosage group were intermediate between changes induced by progesterone and those induced by estrogen. Clinical signs of hyperestrogenism and partial feed refusal were noticed in only some of the high-dosage group animals.     

青贮玉米品种筛选研究

研究4种玉米青贮品种在不同生育期干物质产量、青贮品质表明:在作物干物质产量上,蜡熟期,完熟期产量最高(P<0.05);其中品种京科9号,生产性能表现优异(P<0.05)。在蜡熟期进行玉米青贮,其青贮品质好于乳熟期和完熟期,达到优质标准。     

The occurrence of zearalenone in domestic grains

The occurrence of zearalenone had not been tested n Czechoslovakia; out of a number of analytic procedures for the determination of this mycotoxin in grains and some feed mixtures, the method described by Mirocha et al. (1974) was chosen for this study. The tests were performed with 61 samples of grain and 22 samples of other feeds. In barley 58.3% of the samples were positive in the range from 0.19 to 0.82 micromol.kg-1 in wheat 46.4% of samples in the range from 0.19 to 0.57 micromol. kg-1, in maize 6.7% of samples, containing 0.33 mumol.kg-1. Oat samples contained no zearalenone. The positive findings in the samples of the commercial mixtures COS 1, COS 2, SOL and KPB ranged from 0.19 to 0.32 mumol. kg-1. Zearalenone concentrations above 0.32 mumol. kg-1 were found in seven samples of wheat, in four samples of barley, in one sample of maize and in one sample of complete feed mixture for early-weaned piglets. The described method is recommended for screening examination of grains for zearalenone in Czechoslovakia.     

The occurrence of mycotoxins in cereals and animal feed-stuffs

During the period 1976–1979, just over 4000 samples of animal feedstuffs comprising cereals, compound feeds, hay and silage were examined for moulds and mycotoxins. Examination was carried out in the course of routine advisory and investigational work undertaken by the Agricultural Development and Advisory Service (Ministry of Agriculture, Fisheries and Food) Microbiology Laboratories in England and Wales in connection with livestock health and production problems and defects in grain storage. Mouldy cereals, mostly invaded byPenicillium andAspergillus species, were often found contaminated with ochratoxin A (12.8% positive) and also with citrinin, sterigmatocystin and zearalenone to a lesser extent. Aflatoxin B1 was detected in barley which had been inadequately treated with propionic acid in 3 instances.Commerically-produced compound feedstuffs and protein concentrates were found to contain aflatoxin B1 at levels ranging from less than 10 to 750 g/kg due to the inclusion of contaminated constituents such as ground-nut, palm kernels, cottonseed and copra. There was no evidence that any significant spoilage byAspergillus flavus in compound feedstuffs after manufacture had led to the formation of aflatoxin.Mould spoilage of hay, as indicated by mould counts exceeding 100,000 per g, was common withAspergillus glaucus andPenicillium spp. frequently present, though mycotoxins were not detected, with the exception of sterigmatocystin in just one sample. Silage which had been exposed to air was frequently colonised byPaecilomyces species, giving rise to the formation of patulin in several samples. Despite the frequent detection of mycotoxins in animal feedingstuffs, there was rarely any conclusive evidence implicating them in causing impaired performance or disease in animals.     

"饲用玉米-黑麦草”草地农业系统的研究Ⅰ饲用玉米的品种比较与生产性能

研究表明，在饲用玉米－黑麦草草地农业系统中采用优良的玉米品种。适宜的种植密度，适时的刈割时间和科学的栽培技术，能获得19－27t/hm^2的饲料干物质产量。通过测定饲用玉米乳熟期后整株水分动态变化，为适时刈割和青贮提供了依据。同时还对饲用玉米与黑麦草的时空配置、营养价值和生产成本进行了分析和评价。     

The effects of rumen fluid on the content of zearalenone in animal fodder

The object of the study was to clarify whether rumination has a bearing on the decomposition of zearalenone in the metabolism of the toxin or not. The experiment was performed using rumen fluid in vitro. Rumen fluid samples containing various amounts of zearalenone were incubated in a carbon dioxide atmosphere at 38 degrees C for two days. The zearalenone content was determined by liquid chromatography after 0, 4, 24 and 48 hours incubation periods. The incubation of 4 hours decreased the toxin content by 11%, that of 24 h by 29% and that of 48 h by 37.5% on average. These results indicate that the quantity of zearalenone and the quality of rumen fluid have effects on the ratio of the toxin decomposed.     

The significance of the mycotoxins desoxynivalenol, zearalenone and ochratoxin A for agricultural domestic animals]

Mycotoxins are metabolic products of mycotoxins which have various chemical structures and show various toxic effects. Deoxynivalenol (vomitoxin) is an important economic factor in pig production due to growth depression and suppression of the immune system. Previous studies have shown that the 1-ppm limit in the sole feed for pigs should not be exceeded. Studies of methods of detoxification have as yet not produced conclusive results. Zearalenone has an tolerable effect and may lead to fertility disturbances on the oestrogen production in pigs and can cause remarkable economic damage even in the ppb range. Recommendations of upper limits cannot be made on the basis of the available results. The kidney toxin ochratoxin A is of importance in pig and poultry breeding and--due to its accumulation in the tissue--represents a possible source of danger to man. Since a possible carcinogenic effect of the toxin cannot be excluded, its content in animal rations should be kept as low as possible. For ruminants mycotoxins as a whole do not represent a particular source of danger as these substances can be degraded or converted by rumen flora.     

The occurrence of zearalenone and its derivatives in standard and therapeutic feeds for companion animals

Weather anomalies are the cause of the more and more frequent occurrence of the rezorcycyklic acid derivatives--zearalenone--in the fodders of the animal origin. This mycotoxine induces in the organism of gilts an oestrus-like condition. According to the latest reports the zearalenone derivatives were found in the human food-stuffs in the retail market. It was considered important to assay the concentration of zearalenone and its derivatives in the standard and therapeutic feeds for dogs. Zearalenone and its derivatives were found in 48 commercial formulations for a total number of 57 brands analysed.     

The effect of undecanones and their derivatives on tumor angiogenesis and VEGF content

The in vivo effects of some derivatives of aliphatic ketones (2-undecanone, 3-undecanone, 4-undecanone and their derivatives) on L-1 sarcoma tumor angiogenesis and VEGF content were studied in Balb/c mice. Mice that inhaled 10% solution of 3-undecanone(3-on) or 1% solution of 2-undecanone propylene acetal (Acpr2) for 3 days after tumor cells implantation, presented lower neovascular response measured by tumor-induced cutaneous angiogenesis test (TIA) and lower tumor VEGF content in 5-days tumors, than non-inhaled controls. Other substances presented various effects on tumor VEGF concentration and angiogenesis. Histological examination of lesions collected from mice inhaled Acpr2, or non-inhaled controls, revealed small diffused areas of necrosis in the former group. In both groups, slight to moderate inflammatory infiltrations were seen at the tumor's margin. In Acpr2 group, there were less small blood vessels at tumor's margin than in the control group.     

间接竞争ELISA法检测玉米中ZEN的提取方法研究

应用抗玉米赤霉烯酮单克隆抗体(ZEN-McAb)建立间接竞争ELISA,用以检测玉米中的玉米赤霉烯酮(ZEN)。检测限为1ng/ml,检测范围为1～20ng/ml。在提取玉米赤霉烯酮的过程中,提取液中的甲醇含量对检验结果的影响很大。当提取液中甲醇含量为60%时,加标回收率最理想,平均回收率为92.4%。