Resorcylic Acid Lactones in Urine Samples of Croatian Farm Animals.

Metabolic transformation of zearalenone (ZEA), a mycotoxin which can contaminate both food and feed, results in the formation of five metabolites, one of them being zeranol (α-ZAL), which can be abused in farm animals as a growth promoter. To the best of our knowledge, there is no analytical method that can distinguish whether α-ZAL is present in an animal urine sample as a result of ZEA biotransformation or as a result of anabolic abuse. This study aimed at monitoring resorcylic acid lactones (RALs) concentration in urine of farm animals over several years. Six hundred and three cattle and pig urine samples were collected on farms in different Croatian regions and analyzed for RAL presence. Based on the testing results, all RAL-positive samples were considered to be consequential to feed contamination. The difference in primary ZEA metabolites' ratio (α-zearalenol/ß-zearalenol) was observed between cattle (0.03-0.41) and pig (2.05-17.39) urine samples. If the animals are treated with α-ZAL and fed on ZEA-contaminated feed, α-ZAL and taleranol found in their organisms could come from two sources, so that the reliability of the statistical model might be questionable. Based on these findings, there exists the need for improving the approach to the distinction between α-ZAL abuse and ZEA feed contamination.

Preparation of fowl testes as recipient organs to germ-line chimeras by means of gamma-radiation.

1. The aim of the study was to investigate the possibility of preparing adult fowl testes for the production of exogenous germ-lines by eradication of recipient spermatogenesis using gamma-radiation. 2. A comparison between several radiation therapy treatments (based on 60Co isotope) of male testes was conducted using gamma-rays of 18, 22 and 26 Gy in a single dose or repeated doses of 5 x 8 Gy over a 15-d period. Sperm concentration and motility were determined after each treatment. 3. Altered spermatogenesis was observed after a single treatment dose of 18 Gy, while single doses of 26 Gy were followed by reduced sperm numbers (from 22 x 10(9) to 31 x 10(6) sperm/ml) within 60 to 100 d after treatment. After a single treatment of 26 Gy sperm motility was reduced by 50%. In contrast, a fractionated treatment (5 x 8 Gy) with gamma-rays halted spermatogenesis 39 d after the distribution of the first 8 Gy dose. 4. Observations of the seminiferous tubules by electron microscopy performed 12 months after this treatment confirmed that moderate doses of gamma-rays (8 Gy) distributed repeatedly (5 x) over a limited period (15 d) sterilise adult fowl testes but maintain morphologically normal somatic (Leydig and Sertoli) cell populations.

Expression of human erythropoietin gene in the mammary gland of a transgenic mouse.

WAP is being recognized as the principal milk protein expressed in pregnant or lactating females of several mammalian species. Recently, it has been shown that the 6.3-kb 5' untranslated region of the rWAP gene is able to control, and almost completely restrict, the expression of the transgene into the mammary gland of the transgenic animal. We cloned the genomic fragment carrying the rWAP gene locus from the rabbit phage genomic library and used the 8.5-kb long 5' untranslated part of the rWAP gene to target the expression of hEPO, cloned from the human phage genomic library, into the mammary gland of the mouse. The vectors, carrying either the hEPO gene or the rWAP-hEPO hybrid gene, were injected into the mouse ova, and 12 transgenic animals were identified by PCR and Southern blot from the progeny of 168 tested littermates. Transgenic mice were viable, fertile and displayed a normal development. Recombinant human erythropoietin was produced in the milk of a transgenic mouse female at a secretion level of 5.3 mIU/ml, as detected by ELISA. Despite the low production of the transgenic glycoprotein in the milk we demonstrate that the hybrid gene can be expressed in the mammary gland of the host animal. Thus, WAP-based recombinant vectors, with additional optimizing modifications, can be useful for production of therapeutic proteins in the transgenic mammals.

Sexing of chicken feather follicle, blastodermal and blood cells.

The production of chicken chimeras using donor and acceptor cells which can be of opposite sex has necessitated the utilization of methods developed to distinguish the sex of chickens. We demonstrate one of these methods, based on the polymerase chain reaction which amplifies the EcoRI repeat unit of the fowl W chromosome, and how this technique may be used to sex various cell types in chickens as well as small numbers of blastodermal cells. Our results demonstrate the ability to sex chickens using EcoRI primers, specific for the W chromosome, from as little as 2 ng of female genomic DNA isolated from blood and feathers--the latter being the result of DNA extraction from only one feather. Also evident in this study is the detection of the W chromosome by PCR from approximately 50 blastodermal cells originating from the developing blastodisc at stage X.