TOXICITY OF CHLORPYRIPHOS CONTAINING FORMULATION AND HEAVY ELEMENTS (Cd, Pb) TO CHICKEN EMBRYOS.

The phytotoxic effect of cadmium sulphate, lead acetate and chlorpyriphos containing insecticide Pyrinex 48 EC were investigated on chicken embryos on late developmental stage (day 19 of incubation). The eggs were injected by 0.1 ml amount of lead acetate (0.1%), of cadmium sulphate (0.01%), of Pyrinex 48 EC (chlorpyriphos, 480 g/l; 1%) and as a control 0.1 ml of bird-physiologic saline solution (0.75%). The treatments were performed on day 0 of incubation, and the embryos were examined on day 19. Number of embryonic death, developmental abnormalities and body weight of embryos were recorded on day 19. The body weight of embryos reduced significantly, the rate of embryo mortality increased, but teratogenic effect was not realised by the single treatment of lead acetate and cadmium sulphate. By the single administration of PYRINEX 48 EC the average body weight of embryos resulted in a significant decrease and the rate of embryo mortality and developmental anomalies were increased and it was found to be embryo toxic and teratogenic in the embryos. The increased embryo toxic effect can occur by the additive effect of the heavy elements (cadmium sulphate and lead acetate) and the PYRINEX 48 EC.

TOXIC INTERACTION OF CHLORPYRIPHOS AND COPPER SULPHATE ON CHICKEN EMBRYO.

The single and simultaneous phytotoxic effect of copper sulphate and Pyrinex 48 EC insecticide were investigated on chicken embryos. The eggs were injected by 0.1 ml of copper sulphate solution (0.05%) and/or by 0.1 ml of Pyrinex 48 EC (chlorpyriphos, 480 g/l; 1%). The treatments were performed on day 0 of incubation, and the embryos were examined on day 3 and 19 of it. Germinal disc was prepared to study the early stage of development. Number of embryonic death, developmental abnormalities and body weight of embryos were recorded on day 19. Liver samples were taken for histology and the skeleton was stained by Dawson method. The embryo mortality was not influenced by single treatment of copper sulphate, however, Pyrinex 48 EC and the combination of the test items significantly increased it on day 3. Same tendency was observed in the case of developmental aberration. Single administration of both test items and their combination significantly increased the embryo mortality on day 19. Frequency of abnormalities was not influenced by copper sulphate but single and simultaneous application of insecticide increased it significantly. Pyrinex 48 EC alone and in combination with copper sulphate significantly reduced the body weight, however, the copper sulphate alone did not influence it. Developmental abnormalities were observed sporadically in all cases. There were no findings of drug-induced hepatopathy, however, the ratio of the mitotic cells were markedly reduced. Based on the results, addition and synergistic toxic interaction may be between the copper sulphate and Pyrinex 48 EC that can highly reduce the viability of the embryos.

Toxicity of pendimethalin containing formulation and copper sulphate to chicken embryos.

The aim of this study was to determine the individual and combined toxic effects of STOMP 330 EC herbicide (33% pendimethalin) and copper sulphate on the development of chicken embryos. On the first day of incubation chicken eggs were dipped in the solution or emulsion of the test materials for 30 minutes. Applied concentration of copper sulphate was 0.01% and of herbicide STOMP 330 EC was 1.25%. The chicken embryos were examined for the followings: rate of embryo mortality, body weight, type of developmental anomalies, macroscopic examination. Our teratogenicity study revealed that, the individual toxic effect of copper sulphate and pendimethalin containing herbicide formulation (STOMP 330 EC) were embryotoxic but not teratogenic in chicken. The combined administration of STOMP 330 EC and copper sulphate did not increase the embryotoxic effect.

In vitro eye corrosion study of agrochemicals on isolated chicken eye.

Agrochemicals must undergo numberless toxicological tests before marketing. The eye irritation test is part of this test packet. Nowadays, OECD 405 can be used to classify the irritation potential of substances, the base of the OECD 405 guideline is the Draize test, which is one of the most criticized in vivo methods because of the injuries of the test animals and subjective nature of the test in recording the results. Therefore, several in vitro tests have been developed to replace totally or partly the in vivo eye irritation testing. The isolated chicken eye test method (OECD 438), which was used, is one of these alternative methods. Five different agrochemicals were examined in the following way: All test compounds were applied in a single dose onto the cornea of isolated chicken eyes in order to potentially classify the test compounds as ocular corrosive and/or severe irritant. The damages caused by the test substances were assessed by the determination of corneal swelling, opacity, fluorescein retention and morphological effects. These parameters were evaluated pre-treatment and starting at approximately 30, 75, 120, 180, and 240 minutes after the post-treatment rinse. The endpoints evaluated were corneal opacity, swelling, fluorescein retention and morphological effects. All of the endpoints, with the exception of fluorescein retention (which was determined only at pre-treatment and 30 minutes after test substance exposure) were determined at each of the above time points. Positive and negative controls were used and they showed the expected results in each study. In these in vitro eye corrosives and severe irritants studies, using the Isolated Chicken Eye model with five different products, no ocular corrosion or severe irritation potential were observed. These results correspond to the available information about the tested agrochemicals, so these studies with isolated chicken eye are considered to be successful.

Toxicity of chlorothalonil containing formulation and Cu-sulphate to chicken.

The agrochemicals are being used increasing amounts in plant protection. The plant protection must pay close attention to the decrease or complete elimination of the damage to the environment which affects wildlife, because the cultivated plants provided not only feedstuffs for wild birds but a living and hatching territory as well. The pesticides sprayed during the plant protection activities might have an influence not only on mature birds but also on the embryos in the eggs. The eggs may be exposed to pesticides, the toxic effect of which may appear as embryo lethality and development anomalies. The aim of this study was to determine the individual and combined toxic effects of a chlorothalonil containing fungicide formulation (BRAVO 500) and copper sulphate on the development of chicken embryos. The test materials were injected directly into the air-chamber in a volume of 0.1 ml/egg on the first day of incubation. Applied concentration of copper sulphate was 0.01% and the concentrations of fungicide BRAVO 500 were 1% and 10%. The chicken embryos were examined for the followings: rate of embryo mortality, body weight, type of developmental anomalies, light microscopic examination. Our teratogenicity study revealed that, the combined administration of injected copper sulphate and chlorothalonil containing fungicide formulation (BRAVO 500) caused a significant reduction in the body weight of embryos, markedly increased the rate of embryonic mortality and the incidence of developmental anomalies. The joint toxic effect of copper sulphate and chlorothalonil containing fungicide formulation (BRAVO 500) is an additive effect compared to the individual toxicity of the test materials.

Toxicity of S-metolachlor containing formulation and heavy metals to chicken embryos.

Environmental pollution of metal modelled by copper sulphate, cadmium sulphate and a 960 g/l S-metolachlor containing herbicide formulation (Dual Gold 960 EC) were studied on chicken embryos after administration as a single compounds or in combination. The test materials were injected into the air-chamber in a volume of 0.1 ml/egg on day 0 of incubation. The concentration of copper, and cadmium sulphate was 0.01%. The applied concentration of Dual Gold 960 EC was 0.375%. Evaluation was done on day 19 of the hatching period. In comparison with the values of the control group, the rate of embryomortality also increased significantly in the groups which were treated by herbicide and heavy metals individually. When the herbicide (Dual Gold 960 EC) was used individually, one quarter of the treated animals died in comparison with the values measured in the control group. In the groups which were treated with cadmium sulphate and Dual Gold 960 EC embryomortality increased significantly in comparison with both the control and individually treated groups. In the groups, which were treated together, the occurrence of development disorders remained as low level as in the case individual treatments. Looking at the types of development disorders, the most frequent problems were the oedema, the shortening of the beak mandible, and the incorrect posture of the feet and the neck. As the collective result of the treatment with cadmium sulphate and Dual Gold 960 EC, the body mass of the treated embryos decreased significantly in comparison with the control and individually treated groups, therefore we can conclude that the combined treatment resulted in increased embryo toxic effect in comparison with the individual embryo damaging effect of the used components.

Model study on the clinical signs and residue concentrations of sublethal carbofuran poisoning in birds.

The incidence of fatal poisoning of birds of prey caused by carbofuran has increased markedly in Hungary since 2007. An experimental model with broiler chickens was used to study clinical signs of sublethal carbofuran poisoning in birds and to measure the residue concentrations of carbamate in tissues after exposure. Eight chickens were treated with a carbofuran-containing insecticide orally by gastric tube at a single dose of 2.5 mg/kg body weight, and clinical signs of poisoning were observed. Gas chromatography was used to determine carbofuran concentrations in the blood, muscle, and liver samples, and in stomach contents. Poisoning was characterized by typical muscarinic and nicotinic clinical signs without mortality. Carbofuran in the stomach and edible tissues of acutely poisoned birds may lead to secondary poisoning of predators and may also present risks to human health.

Investigation on possible ecotoxicological risk of carbofuran insecticides.

Carbofuran-containing insecticides are widely used agents in plant protection. Their use may pose considerable environmental risk for both the protected and non-protected predator and plantivorous birds. For defence of wild birds a model experiment was carried out on broiler chickens. In the study, eight animals were treated orally by gastric tube with a carbofuran-containing insecticide at a single dose of 2.5 mg/kg b.w. One animal served as untreated control specimen. Forage and drinking water were provided ad libitum. After the treatment, the possible clinical signs were observed carefully, blood samples were obtained from each bird and after exsanguinations liver, breast and leg muscle samples and stomach content were taken. The carbofuran concentration in blood, tissues and stomach content was determined by gas chromatographic method. Thirty minutes after poisoning, the average carbofuran concentration in breast muscle of chickens exceeded the maximum level of 0.1 mg/kg permitted in edible tissues, whereas ninety minutes after poisoning the concentration of one sample was still above the limit value. In the liver, leg-muscle and blood samples, the measured carbofuran concentration was lower than the permitted maximum value, except in the blood of two animals. The carbofuran concentration of the stomach content markedly exceeded the limit value. The sublethal concentration of the pesticides can reduce the capable of living of wild animals. Due to the sub toxic dose the poisoned birds can survive; however, the residue of insecticides can lead to secondary toxicosis of other animals.

Eye irritation study of some pesticides on chorioallantoic membrane of the egg.

The chorioallantoic membrane of chick embryo has been used extensively for many years in various fields of biological research, including virology, bacteriology and toxicology. The chorioallantoic membrane (CAM) is a complete tissue that responds to injury with a complete inflammatory reaction, this process similar to that induced by chemicals in the conjunctival tissue of the rabbit eye. A possible model for assessing the irritation potential of a chemical or product to such a vascularised tissue is the choriallantoic membrane of the embryonated hen's egg, as this is a highly vascular, thin membrane with relatively easy access for both treatment and assessment. In recent years various in vitro methods have been developed to replace the heavily criticized Draize rabbit eye test for irritation testing. One of the most studied alternative methods is the Hen's Egg Test - Chorioallantoic membrane (HET-CAM). In our studies a comparative screening was done with a set of pesticides to establish parallel data on in vitro (HET-CAM) and in vivo (Draize) results. In most cases good correlation was found between the HET-CAM assessment and results from the Draize rabbit eye test. The actual form of the HET-CAM test is a valuable pre-screen for predicting ocular irritation potential of chemicals, and can be used to reduce the number of experimental animals. The HET-CAM test is useful as a part of a battery of tests to replace the Draize rabbit eye test.

Pharmacokinetics and bioavailability of doxycycline in fasted and nonfasted broiler chickens.

The pharmacokinetics and the influence of food on the kinetic profile and bioavailability of doxycycline was studied after a single intravenous (i.v.) and oral dose of 10.0 mg/kg body weight in 7-week-old broiler chickens. Following i.v. administration the drug was rapidly distributed in the body with a distribution half-life of 0.21 +/- 0.01 h. The elimination half-life of 6.78 +/- 0.06 h was relatively long and resulted from both a low total body clearance of 0.139 +/- 0.007 L/h.kg and a large volume of distribution of 1.36 +/- 0.06 L/kg. After oral administration to fasted chickens, the absorption of doxycycline was quite fast and substantial as shown by the absorption half-life of 0.39 +/- 0.03 h, the maximal plasma concentration of 4.47 +/- 0.16 micrograms/mL and the time to reach the Cmax of 1.73 +/- 0.06 h. The distribution and the final elimination of the drug were slower than after i.v. administration. The absolute bioavailability was 73.4 +/- 2.5%. The presence of food in the intestinal tract reduced and extended the absorption (t1/2a = 1.23 +/- 0.21 h; Cmax = 3.07 +/- 0.23 micrograms/mL; tmax = 3.34 +/- 0.21 h). The absolute bioavailability was reduced to 61.1% +/- 4.4%.

Toxicological studies on potentiated ionophores in chickens. I. Tolerance study.

The tolerance of chickens to monensin (12.5 mg/kg of feed) and maduramicin (3.0 mg/kg of feed) fed at a reduced dose in the presence of the antioxidant duokvin was studied in two experiments including 2 x 200 Tetra-82 broiler chickens. Tolerance was assessed by the appearance of clinical signs indicative of a toxic effect, the number of deaths, the groups' body weight gain, feed and drinking water intake, the aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) activities, calcium ion, inorganic phosphate and total protein content of the blood plasma, the haematocrit value, and haemoglobin concentration. When applied at a dose that had proved to be optimum in the efficacy studies, neither the monensin-duokvin combination (12.5 mg monensin per kg of feed + 120 mg duokvin per kg of feed) nor the maduramicin-duokvin combination (3.0 mg maduramicin per kg of feed + 120 mg duokvin per kg of feed) exerted a statistically significant influence on the parameters tested.

Toxicological studies on potentiated ionophores in chickens. II. Compatibility study.

Two trials were carried out on a total of 2 x 360 Tetra-82 broiler chickens to study how the presence of the antioxidant duokvin as potentiating agent influenced the compatibility of reduced doses of monensin (12.5 mg/kg of feed) or maduramicin (3.0 mg/kg of feed) with other chemotherapeutic agents (tiamulin, erythromycin, sulfaquinoxaline, sulfachlorpyrazine, flumequine, tylosin, kitasamycin) widely used in broiler rearing. Compatibility was assessed on the basis of the appearance of clinical signs suggestive of toxic interaction, the mortality rate, body mass gain, feed consumption and drinking water intake, and changes in AST and LDH activities of the blood plasma. The monensin-duokvin combination (12.5 mg monensin/kg of feed + 120 mg duokvin/kg of feed) was found to be compatible with erythromycin, sulfaquinoxaline, sulfachlorpyrazine, flumequine, tylosin and kitasamycin. For tiamulin, a slight incompatibility was observed; however, this was much less severe than that found for monensin administered at a dose of 100 mg/kg of feed. The maduramicin-duokvin combination (3.0 mg maduramicin/kg of feed + 120 mg duokvin/kg of feed) was compatible with all the compounds tested; thus, it can be safely applied also in combination with tiamulin.

Toxicological studies on potentiated ionophores in chickens. III. Electrotoxicological investigations.

The effect exerted by the simultaneous administration of tiamulin (40 mg/kg body mass) and a lowered dose (12 mg/kg of feed) of monensin combined with the antioxidant duokvin on the conductivity of peripheral nerve and the electrophysiological function of the heart was studied in two trials comprising 3 x 6 broiler chickens. For the study of peripheral nerve function, chickens were anaesthetized with pentobarbital sodium, the sciatic nerve (n. ischiadicus) was exposed in the sciatic arc, and bipolar stimulating and efferent electrodes were inserted directly beside the nerve, at an average distance of 20 mm from each other. For the recordings, the nerve was stimulated with 16 supramaximal impulses of 0.02 ms duration, applied at 15-s intervals. The responses given to the stimulus were recorded with the help of a polygraph and evaluated with a computer in online mode. For the study of cardiac function, the chickens were anaesthetized with methomidate. The electrocardiograms were taken in Einthoven's lead II with the help of needle electrodes, and then evaluated with computer in online mode. In control chickens, maximum conduction velocity was 30.70 +/- 0.52 m/s on the average. That value did not change after a single treatment with monensin-duokvin + tiamulin, while it underwent a slight but statistically significant decrease after two treatments. Studies on the electrical function of the heart revealed no signs of dysfunction in either of the treated groups as compared to the control.(ABSTRACT TRUNCATED AT 250 WORDS)

Potentiation of ionophorous anticoccidials with dihydroquinolines: battery trials against Eimeria tenella in chickens.

Seven battery tests, employing a total of 1344 Hybro cockerels, were conducted in order to study the interaction between ionophorous anticoccidials and a new dihydroquinoline-type antioxidant known as duokvin. A significant, non-selective, toxic interaction was established, resulting in growth depression and improved anticoccidal efficacy against Eimeria tenella and E. mitis in these chickens. The duokvin itself showed no appreciable activity against the coccidia. The adverse effects of the interaction were eliminated, and the anticoccidial efficacy of the approved levels of ionophores was maintained, when the dietary levels of monovalent cation ionophorous monensin, salinomycin and narasin were reduced to approximately 12% in the presence of 120 p.p.m. duokvin. No adverse effects on the growth of chickens appeared in the combination with maduramicin, yet the enhancement of anticoccidial activity allowed an approx. 50% reduction of this ionophore as well.

Potentiation of ionophorous anticoccidials with dihydroquinolines: reduction of adverse interactions with antimicrobials.

In two experiments, the compatibility of the anticoccidial combinations of monensin and duokvin, as well as that of maduramicin and duokvin, with some antimicrobials widely used in the broiler production was studied in cockerels. The monensin-duokvin combination was found to be fully compatible with erythromycin, sulphachlorpyrazine, and sulphaquinoxaline. With tiamulin, a slight interaction was observed, but it was far less severe than the toxic interaction between monensin and the diterpene antibiotic. The maduramicin-duokvin combination proved to be compatible with all of the chemotherapeutics tested, including tiamulin. The results of the studies indicate that the adverse interactions of monensin and maduramicin with certain antimicrobials can be considerably diminished or even abolished by using them in reduced doses in combination with the dihydroquinoline compound duokvin.

[Effect of equilateral administration of monensin and tiamulin on the peripheral nerve function and the electrocardiogram of broilers]. / Einfluss der gleichzeitigen Applikation von Monensin und Tiamulin auf die periphere Nerventätigkeit und das Elektrokardiogramm bei Broilern.

The influence of the simultaneous administration of monensin and tiamulin on the motor nerve conduction velocity and the refractory periods of the N. ischiadicus was studied on 4-week old cockerels. The combined administration of the two substances resulted in significant decrease of the conduction velocities of the peripheral nerve, while the relative and absolute refractory periods were seen to increase significantly as a result of the medications. The observed alteration of the peripheral nerve function in the early phase can already attribute to the development of the serious clinical signs of the toxic interaction. The simultaneous application of monensin and tiamulin caused also changes in the electrocardiogram which was expressed in a prolongation of the RS interval and the ST segment. At the same time the heart rate was reduced.