Effect of zearalenone on female White Leghorn chickens.

Acute toxic effects of purified zearalenone were studied in growing female White Leghorn chickens. In the first experiment, zearalenone in gelatin capsules was administered to 10 chickens (zearalenone-treated chickens [ZC]) in a single oral dose of 15.0 g/kg. Another 10 control chickens (CC) received empty gelatin capsules. All chickens survived the 10-day experiment and did not show any noticeable gross or histopathological lesions. There were no differences between CC and ZC in weight gain, oviduct, comb and liver weights, hematological parameters, and serum cholesterol. ZC had significantly less (P less than 0.05) serum calcium but significantly greater (P less than 0.01) serum phosphorus than CC. In the second experiment, zearalenone was administered orally or intramuscularly (pectoral muscle) at levels of 0, 50, 200, 400, and 800 mg/kg for 7 consecutive days. The oviduct weight increased with increasing toxin levels in both orally (OZC) and intramuscularly (IZC) administered groups: there were more pronounced effects in the IZC. The liver weight increased and comb weight decreased in IZC. The relative estrogenic biopotency of zearalenone in IZC, using estradiol dipropionate as a standard, was 1.37%. The results of this experiment demonstrate that chickens are highly tolerant to zearalenone and that the estrogenic effects of the toxin are greater when it is administered in multiple doses than in a single dose and in IZC than in OZC.     

The effect of method of administration on ovarian responses of seasonally anestrous ewes administered gonadotrophin releasing hormone

Mature ewes were treated during the anestrous season with saline (I) or GnRH either intramuscularly in saline (II), subcutaneously in carboxymethylcellulose (CMC) (III) or subcutaneously in gelatin capsules (IV). Fifty μg of GnRH or 1 ml of saline were administered to 22 ewes in experiment 1. In experiments 2 and 3, forty-seven and 10 ewes received 250 μg GnRH or 1 ml of saline. Ewes were bled for progesterone determination prior to treatment and up to 12 or 13 days after treatment. In experiment 3, ovaries were observed via mid-ventral laparotomy 4 days after treatment and ovarian structures recorded. Ewes were classified into one of four progesterone response categories: cyclic, transient, prolonged or no response. The only treatment that changed the progesterone response from the saline-treated controls was GnRH in gelatin capsules. More ewes in this group were classified with a prolonged progesterone response (40%) than in the saline control group (0%). GnRH (in gelatin capsules)-treated ewes in the prolonged progesterone response category had higher concentrations of plasma progesterone than GnRH (in saline or CMC)-treated ewes with a prolonged progesterone response. For the GnRH (in gelatin capsule)-treated ewes, the prolonged progesterone response was similar to progesterone in ewes during the estrous cycle and all ewes in the prolonged progesterone category had corpora lutea (experiment 3). In summary, implanting the GnRH in gelatin capsules subcutaneously in seasonally anestrous ewes increased the ovulation response and enhanced corpus luteum function over ewes administered GnRH in saline intramuscularly.     

The effect of method of GnRH administration and short-term calf removal on ovarian function and reproductive performance in postpartum suckled beef cows administered PGF(2)alpha for estrous synchronization

The first experiment was a 2 x 2 factorial experiment with calf removal (none or short-term) and method of GnRH administration (intramuscularly in saline or subcutaneously in gelatin capsules) as main effects. The durations of the GnRH-induced LH surges were similar among groups but the LH surges were delayed in the cows that received GnRH subcutaneously in gelatin capsules. Calf removal enhanced the GnRH-induced LH release for cows administered GnRH subcutaneously in a gelatin capsule but not for cows administered GnRH intramuscularly in saline. In the second experiment, 191 postpartum suckled beef cows were administered two injections of prostaglandin F(2)alpha(PGF(2)alpha) 11 days apart. After the second PGF(2)alpha injection, the cows were assigned to a 2 x 2 factorial experiment as in Experiment 1 plus one control group. Short-term calf removal (47 h) began 28 h after the second PGF(2)alpha injection. GnRH was administered 30 h after the time of calf removal. The number of cows that ovulated following the time of the GnRH treatment, the number that had abnormal luteal phases and the first-service pregnancy rates among treatment groups within the anestrous and cyclic cows classifications were not significantly different. However, several effects were detected and are reported.     

alpha-Zearalenol, a major hepatic metabolite in rats of zearalenone, an estrogenic mycotoxin of Fusarium species

The chemical and biological properties of the hepatic metabolite of zearalenone, an estrogenic and non-steroidal fungal toxin produced by Fusarium species, were investigated by employing TLC, GC/MS, high pressure liquid chromatography and fluorospectral analyses, as well as uterine weight bioassay in immature mice. All the chemical and physical data supported the view that the major metabolite, obtained by incubating zearalenone with S-9 and microsomes of rat liver in the presence of NADPH, is C-6'-alpha-hydroxylated zearalenone (alpha-zearalenol). The estrogenic activity of this metabolite was several times higher than that of the parent zearalenone, and the results of biological and toxicological evaluations of alpha-zearalenol are discussed.     

Ovarian responses of seasonally anestrous ewes administered progesterone,PMS, HCG and(or) GnRH

One hundred and sixty ewes were assigned to sixteen groups in a 2 × 2 × 4 factoral design and were treated during the anestrous season. The main effects were progesterone pretreatment (non-implanted and implanted for 14 days), PMS pretreatment (no pretreatment and pretreatment with 500 IU at the time of progesterone implant removal) and treatments (none, GnRH in saline, GnRH in gelatin capsules and HCG). GnRH in saline (250 μg) and HCG (500 IU) were administered intramuscularly and GnRH in gelatin capsules (250 μg) was administered subcutaneously 24 hours after the time of progesterone implant removal.Ewes were classified into one of four progesterone response categories: cyclic, transient, prolonged and no response. An injection of GnRH in saline induced a prolonged progesterone response in only one ewe (13%) which was similar to the response in the untreated ewes (0%). More ewes administered GnRH in gelatin capsules (56%) and more ewes administered HCG (89%) had a prolonged progesterone response than GnRH (in saline) treated or untreated ewes. A higher percentage of ewes that were pretreated with PMS and treated with GnRH in saline (78%) had a prolonged progesterone response than ewes treated with either PMS (22%) alone or with GnRH (in saline; 13%) alone.     

Influence of neuropeptide Y (NPY) on food intake and growth of penaeid shrimps Marsupenaeus japonicus and Penaeus semisulcatus (Decapoda: Penaeidae)

The effects of neuropeptide Y (NPY) administered intramuscularly or orally on postlarvae (PLs) of two penaeid species were investigated in this study. In experiment 1, food intake (FI) of Marsupenaeus japonicus PLs (0.96 g), injected with NPY at 0.6 microg per g BW, was investigated within 48 h posttreatment. In experiment 2, oral administration of NPY (at doses of 0, 0.125, 0.25, 0.5 microg g(-1) food) on feed intake and growth performance of Penaeus semisulcatus PLs (0.27 g) was examined for 6 weeks. In experiment 1, NPY injection significantly increased average daily FI of M. japonicus PLs within the first 24 h compared to the control (P<0.05), but its stimulatory effect decreased on the second day (P>0.05). The increase in FI was 33% during the first 24 h and 17% during the next 24 h. In experiment 2, significant (P<0.05) differences were found among the groups in terms of weight gain and food utilization (P<0.05). Mean FI significantly increased (as much as 1.3-fold over the control) when NPY was orally administered at doses from 0.125 to 0.5 microg g(-1) feed. There was a positive relationship between FI and final weight (y=-0.972+2.098x, R(2)=0.81) and between FCE and NPY doses in the diets (y=45.37+3.46x, R(2)=0.91). The present findings indicated for the first time that NPY is a potent stimulator of food intake when administered either intramuscularly or orally to penaeid shrimps.     

Rotavirus virus-like particles administered mucosally induce protective immunity.

We have evaluated the immunogenicity and protective efficacy of rotavirus subunit vaccines administered by mucosal routes. Virus-like particles (VLPs) produced by self-assembly of individual rotavirus structural proteins coexpressed by baculovirus recombinants in insect cells were the subunit vaccine tested. We first compared the immunogenicities and protective efficacies of VLPs containing VP2 and VP6 (2/6-VLPs) and G3 2/6/7-VLPs mixed with cholera toxin and administered by oral and intranasal routes in the adult mouse model of rotavirus infection. VLPs administered orally induced serum antibody and intestinal immunoglobulin A (IgA) and IgG. The highest oral dose (100 microg) of VLPs induced protection from rotavirus challenge (> or = 50% reduction in virus shedding) in 50% of the mice. VLPs administered intranasally induced higher serum and intestinal antibody responses than VLPs administered orally. All mice receiving VLPs intranasally were protected from challenge; no virus was shed after challenge. Since there was no difference in immunogenicity or protective efficacy between 2/6- and 2/6/7-VLPs, protection was achieved without inclusion of the neutralization antigens VP7 and VP4. We also tested the immunogenicities and protective efficacies of 2/6-VLPs administered intranasally without the addition of cholera toxin. 2/6-VLPs administered intranasally without cholera toxin induced lower serum and intestinal antibody titers than 2/6-VLPs administered with cholera toxin. The highest dose (100 microg) of 2/6-VLPs administered intranasally without cholera toxin resulted in a mean reduction in shedding of 38%. When cholera toxin was added, higher levels of protection were achieved with 10-fold less immunogen. VLPs administered mucosally offer a promising, safe, nonreplicating vaccine for rotavirus.     

红三叶草总异黄酮对小公鸡生长及血清睾酮水平的影响

日粮中添加红三叶草总异黄酮（４．３８ｍｇ／ｋｇ饲料）,饲喂４５－５６日龄红布罗肉用公仔鸡,与对照组比较,总异黄酮实验组公仔鸡日增重提高５．１％,料重比下降９．２％,提高了饲料的利用率。屠宰结果表明,腹脂重、腹脂／胴体显著降低,鸡冠重、睾丸重增加,鸡冠、睾丸和体重间的相关分析,睾丸石蜡切片和血清睾酮含量的放射免疫测定,均表明总异黄酮能提高血清睾酮水平,促进雄性动物生殖系统发育。     

Production of zearalenone,nivalenol, moniliformin,and wortmannin from toxigenic cultures ofFusarium obtained from pasture soil samples collected in New Zealand

One culture ofF avenaceum, 4 cultures ofF oxysporum, and 11 cultures of Fsambucinum were isolated from soil samples of pasture in New Zealand in 1987. All cultures, when grown on rice media and fed to rats caused a weight loss in rats as well as toxic signs including hemorrhaging and congestion, uterine enlargement, and hematuria. 6 out of 16 cultures caused death in rat feeding tests.F oxysporum #1 killed rats (feeding test) within 5-12hrs. 10 cultures produced zearalenone (19 to 8,849 ppm), 8 cultures produced nivalenol (32 to 117 ppm), 1 culture,F sambucinum #8, produced wortmannin (40 ppm), and 5 cultures produced moniliformin (19 to 9,000ppm). We report for the first time the co-occurrence of zearalenone, nivalenol, and moniliformin produced byF sambucinum #3 in culture.F avenaceum #1 andF oxysporum cultures (nos 1, 2, and 3) produced moniliformin alone.F oxysporum #4 produced zearalenone alone as well.F sambucinum #5 caused erythema in the small intestine of rats and 100% mortality and did not produce any known toxin(s). Nivalenol when administered to the stomach of rats orally at levels 10, 20, and 40mg/kg body weight caused inflammation in the intestines, coma, and death. The mycotoxins T-2 toxin, HT-2 toxin, T-2 tetraol, diacetoxyscirpenol (DAS), monoacetoxyscirpenol (MAS), deoxynivalenol (DON), 3-acetyl-and 15-acetyldeoxynivalenol, depoxynivalenol, fusarenon-X, alpha-and beta-zearalenone, and fusarochromanone (TDP-1) were not detected in the extracts of these cultures.     

Reproductive performance of Coopworth ewes following oral doses of zearalenone before and after mating

The reproductive performance of Coopworth ewes after administration of zearalenone was determined in two trials. In Trial I, zearalenone was administered to groups of 33 ewes at rates of 0, 1.5, 3.0, 6.0, 12.0 and 24.0 mg/ewe/day for 10 days, starting on Day 7 of the oestrous cycle before mating. There was a linear decline (P less than 0.001) in ovulation rate with dose of zearalenone; also cycle length decreased and duration of oestrous increased with increasing dose levels. Reductions in the incidence of ovulation and in fertilization were seen only at doses of 12 and 24 mg. In Trial 2, groups of 50 ewes were given the same range of doses of zearalenone for 10 days, starting 5 days after mating to entire rams. There was no effect of zearalenone treatment after mating on pregnancy rate or embryonic loss. These results indicate that the effects of zearalenone, administered orally, on ewe reproduction, at the dose levels examined, were restricted to ewes exposed before mating. Intakes of zearalenone of 3 mg/ewe/day or more during this period would be reflected as depressed ovulation rates and lower lambing percentages.     

Tremorgenic Toxin from Penicillium verruculosum

A new mycotoxin that produces severe tremors and acute toxicity when administered orally or intraperitoneally (ip) to mice and 1-day-old cockerels was obtained from a strain of Penicillium verruculosum Peyronel isolated from peanuts. The ip 50% lethal dose (LD(50)) of this tremorgen was 2.4 mg/kg in mice and 15.2 mg/kg in chickens. Orally administered LD(50) values for the toxin were 126.7 mg/kg in mice and 365.5 mg/kg in chickens. The trivial name "verruculogen" is proposed for this tremorgenic mycotoxin. Physical and chemical characteristics of the mycotoxin are described.     

The effects of gossypol on spermatogenesis and development of the eyefluke, Philophthalmus gralli, and its chicken host

To determine the effects of gossypol, a male antifertility drug, on the eyefluke, P. gralli, this chemical was administered orally to chickens in long-term and short-term regimens. Gossypol acetic acid (GAA), fed to juvenile chickens from 1 to 35 days, caused a decreased weight gain when compared to controls on untreated feed. An FeSO4 supplement to the GAA-fed chickens provided partial protection from the toxic effects of GAA. Worms from GAA-fed chickens were significantly larger than controls, while those from chickens fed GAA + FeSO4 were intermediate in size. Sperm development in these worms was unaffected by GAA. In a second experiment, GAA was administered either in the feed of the hosts from days 35 to 70 or by capsule from days 63 to 77. Worms were exposed to [3H] thymidine, transplanted to the host's eyes, removed on a timed schedule, and processed for autoradiography to determine the rate of spermatogenesis in both GAA-feed and GAA-capsule groups. Early stages of spermatogenesis in both groups were unaffected by GAA and later stages developed at a slightly faster rate than reported for worms from chickens on untreated feed. Higher frequencies of testicular anomalies were observed in both groups including 3 testes, 1 testis, no testes, fused testes, degenerating testes, ovarian tissue in the testes, deformed sperm, and encapsulated sperm. Testes from chickens in both groups showed a significantly lower weight and no signs of spermatogenesis when compared to control chickens.     

Opposite effects of glucocorticoid on estrogen-induced growth and differentiation of quail oviduct: demonstration by sequential treatments

Control of the development and functions of avian oviduct is monitored by four classes of steroid hormones, including glucocorticoids. The effects of dexamethasone (DEX), a synthetic glucocorticoid, were studied via sequential treatments with estradiol benzoate, paying special attention to changes in estrogenic oviduct responses involving DNA synthesis and cell proliferation, ovalbumin accumulation and cell differentiation. DEX exerted an antagonistic effect upon estrogen stimulation when administered separately before or after estradiol benzoate (EB). Given before EB, DEX was more strongly antagonistic for DNA synthesis than when given simultaneously with EB. Administered after EB, DEX reversed EB-induced cell proliferation: the DNA content declined and the oviduct regressed. In the same way, protein and ovalbumin synthesis was inhibited and delayed by first intervention of DEX, and accelerated catabolism of ovalbumin and proteins was observed when DEX followed EB. DEX, which was ineffective alone, but synergistic on ovalbumin synthesis when given concomitantly with EB, prevented or dissipated the estrogenic effects, cell proliferation and secretory process when administered in sequential treatments.     

Indirect enzyme-linked immunosorbent assay for the mycotoxin zearalenone

A competitive indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of zearalenone, an estrogenic mycotoxin. Zearalenone was converted to zearalenone-6'-carboxymethyloxime and conjugated to bovine serum albumin and poly-L-lysine for use as immunogen and solid-phase marker, respectively. Immunization of rabbits with the bovine serum albumin conjugate resulted in zearalenone antibody titers of 20,480 in 11 weeks. A competitive indirect ELISA was conducted by simultaneously incubating zearalenone with zearalenone antiserum over zearalenone-6'-carboxymethyloxime poly-L-lysine solid phase and then determining the bound rabbit immunoglobulin with goat anti-rabbit peroxidase conjugate. Response range for zearalenone in the resulting competition curve was between 1 and 50 ng/ml. Reactivities of this antiserum for alpha-zearalenol, beta-zearalenol, alpha-zearalanol, and beta-zearalanol were, respectively, 50, 12, 6, and 3% of that found for zearalenone. By using the competitive indirect ELISA, zearalenone was detectable in methanol-water extracts of corn, wheat, and pig feed samples.     

Indirect enzyme-linked immunosorbent assay for the mycotoxin zearalenone.

A competitive indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of zearalenone, an estrogenic mycotoxin. Zearalenone was converted to zearalenone-6'-carboxymethyloxime and conjugated to bovine serum albumin and poly-L-lysine for use as immunogen and solid-phase marker, respectively. Immunization of rabbits with the bovine serum albumin conjugate resulted in zearalenone antibody titers of 20,480 in 11 weeks. A competitive indirect ELISA was conducted by simultaneously incubating zearalenone with zearalenone antiserum over zearalenone-6'-carboxymethyloxime poly-L-lysine solid phase and then determining the bound rabbit immunoglobulin with goat anti-rabbit peroxidase conjugate. Response range for zearalenone in the resulting competition curve was between 1 and 50 ng/ml. Reactivities of this antiserum for alpha-zearalenol, beta-zearalenol, alpha-zearalanol, and beta-zearalanol were, respectively, 50, 12, 6, and 3% of that found for zearalenone. By using the competitive indirect ELISA, zearalenone was detectable in methanol-water extracts of corn, wheat, and pig feed samples.     

鸡apoA5基因单核苷酸多态性及其与屠体性状的关联研究

以丝羽乌骨鸡和隐性白洛克正反交产生的F2代为实验群体, 采用PCR-SSCP和DNA测序的方法检测鸡载脂蛋白A5(apoA5)基因的单核苷酸多态性(SNPs), 并将所发现的SNPs与体重、胸肌重、腿肌重、心脏重、肝脏重和腹脂重等屠体性状进行关联分析。结果发现, 鸡apoA5基因5′-调控区C-169T, 外显子2 C600T、T635C, 外显子3 C841G、C914T、C1142G、C1394T共7个突变位点。其中外显子2突变位点C600T、T635C对12周龄腹脂重、腹脂率、肝脏重和心脏重有显著影响(P<0.05), 根据PCR-SSCP的结果将其分为6种基因型(AA, AB, AC, BB, BC, CC): 其中CC型个体的腹脂重和腹脂率显著高于AA型、AB型、AC型、BB型、BC型 (P<0.05); AC型个体的肝脏重显著低于AA型、AB型、BB型、BC型和CC型的肝脏重(P<0.05); BC型个体的心脏重显著低于BB型的心脏重(P<0.05)。     

Effect of F-2 and T-2 fusariotoxins on experimental Cryptosporidium baileyi infection in chickens

The course of Cryptosporidium baileyi infection in chickens fed with different doses of fusariotoxins was compared with that of control groups. F-2 toxin levels of 0.187–1.5 mg kg⁻¹ and T-2 toxin levels of 0.187–6.0 mg kg⁻¹ were investigated. The experimental amimals were orally infected with 6 × 10⁵ C. baileyi oocysts at 1 week of age. Total daily oocyst output was monitored by a quantitative method. Acquired immunity was tested at the age of 4 weeks, by ELISA and by a challenge infection with an equal number of oocysts, upon recovery from the primary infection. The results show that in chickens kept on the lower doses of F-2 and T-2 toxins, the parasite infection ran a similar course to that in the control groups, and the animals became resistant to re-infection. However, when higher doses (2.0–6.0 mg kg⁻¹) of T-2 toxin were used, a depression of weight gain was observed with some other physiological parameters (PCV, weight of bursa, weight of thymus, skin thickness in PHA-P skin test) also indicating toxic effect and, simultaneously, the oocyst output decreased significantly and the patent period was slightly prolonged. Although certain modifications of the immune response could be revealed, the chickens became resistant to re-infection. Only early (1 week of age) parasite infection and 6 mg kg⁻¹ T-2 toxin in the feed significantly depressed body weight gain and immunity.     

T-2 metabolites in the excreta of broiler chickens administered 3H-labeled T-2 toxin.

A method for the detection of T-2 metabolites was developed and applied to analysis of metabolites in excreta of broiler chickens administered 3H-labeled T-2 toxin. The method used acetonitrile extraction and partitioning with petroleum ether followed by chromatography on Amberlite XAD-2, Florisil, and Sep-Pak C18. The recovery of T-2 toxin added to the chicken excreta was 73% at a concentration of 0.2 microgram/g. About 80% of orally administered 3H-labeled T-2 toxin was rapidly metabolized to more polar derivatives and eliminated in the excreta within 48 h. T-2 toxin, HT-2 toxin, neosolaniol, and T-2 tetraol were detected at 0.06 to 1.13% of the total dose, 48 h after administration. Eight unknown derivatives, named TB-1 to TB-8, were quantitatively more significant than the metabolites above. TB-3 and TB-9 represented about 12 and 25% of the total dose, respectively. One of the metabolites (TB-6), 1.5% of the total dose, was identified as 4-deacetylneosolaniol (15-acetyl-3 alpha, 4 beta, 8 alpha-trihydroxy-12, 13-epoxytrichothec-9-ene).     

Uterotropic activity of cis and trans isomers of zearalenone and zearalenol.

The cis and trans isomers of zearalenone [2,4-dihyroxy-6-(10-hydroxy-6-oxo-1-undecenyl)-benzoic acid mu-lactone] and zearalenol [2,4-dihydroxy-6-(6,10-dihydroxy-1-undecenyl)-benzoic acid mu-lactone] were tested for uterotropic activity in the white rat. The metabolites were administered through the oral route (per os) and by topical application to the freshly shaven skin on the back. cis-Zearalenone was significantly more active than trans when fed orally to the rats in the diet or when applied topically by skin application. However, the cis isomer of zearalenol was not significantly different than its trans isomer. trans-Zearalenone was less active than trans-zearalenol.     

Estrogen-controlled gene expression in tissue culture cells by zearalenone

In two estrogen-sensitive cell lines, Le42 and MCF-7, the estrogenic potential of the nonsteroidal mycotoxin zearalenone has been investigated. The chloramphenicol acetyltransferase (CAT) gene expression in Le42 cells is induced by zearalenone after transfection with a CAT-gene construct controlled by an estrogen responsive element [(1986) Cell 46, 1053-1061]. In MCF-7 cells zearalenone induces at least 2 exoproteins (52 and 160 kDa) which are estrogen-specific [(1980) Cell 20, 353-362). These data suggest that zearalenone acts by activating the estrogen receptor. Due to the high sensitivity of these cell lines for zearalenone both test systems are proposed as assays for a quantitative estimation of the biological (estrogenic) activity of this widespread mycotoxin.