In vivo screening of five phytochemicals/extracts and a fungal immunomodulatory protein against colibacillosis in broilers.

Five phytochemicals/extracts (an extract from Echinacea purpurea, a ß-glucan-rich extract from Shiitake, betaine [Betain™], curcumin from Curcuma longa [turmeric] powder, carvacrol and also a recombinant fungal immunomodulatory protein [FIP] from Ganoderma lucidum) cloned and expressed in Escherichia coli were investigated for their anticolibacillosis potential in three chicken experiments, which were conducted in floor pens. Birds that were inoculated with E. coli intratracheally were treated with the phytochemicals/extracts or the FIP and compared with doxycycline-medicated and non-medicated infected broilers. Non-medicated and non-infected birds were used as negative controls. Mortality, colibacillosis lesions and body weight gains were used as parameters. Considering the sum of dead birds and chickens with generalized colibacillosis per group, there was no significant difference between the positive control groups and birds treated with phytochemicals/extracts or the FIP. In contrast, doxycycline-treated birds showed significantly lower mortality and generalized colibacillosis. Moreover, none of the phytochemicals/extracts and the FIP improved recovery from colibacillosis lesions, while all doxycycline-treated broilers recovered completely. The negative control birds and doxycycline-treated groups consistently showed the highest weight gains. Pulsed-field gel electrophoresis of reisolates showed that they were genetically indistinguishable from the inoculation strain. In conclusion, none of the tested phytochemicals/extracts and the FIP significantly reduced the E. coli-induced mortality and generalized colibacillosis, and nor did they improve recovery from colibacillosis lesions.

Collaborative study of a microbiological screening method (three-plate) for the banned antimicrobial growth promotors tylosin, virginiamycin, spiramycin, zinc bacitracin and avoparcin in animal feed.

A microbiological screening method (three-plate) for the detection of the antimicrobial growth promoters tylosin, spiramycin, virginiamycin, zinc bacitracin, and avoparcin in animal feed has been developed and validated successfully. A collaborative study involving 18 laboratories receiving 172 samples was carried out to verify the performance characteristics. The detection level for tylosin/virginiamycin/spiramycin, expressed in microbiological activity, was 1 mg kg(-1) (false-positives, 2%; false-negatives, 3, 0, and 6%, respectively). Avoparcin could be detected at 1 mg kg(-1) in feed in general (false-positives, 2%; false-negatives, 0%). However, in calf feed the sensitivity was lower. The percentages of false-negatives were found to be 12%, 7%, and 0% at 1, 3, and 5 mg kg(-1), respectively (false-positives, 4%). The limit of detection for zinc bacitracin was 3-5 mg kg(-1) (false-positives, 5-10%; false-negatives, 77% at 1 mg kg(-1), 45% at 2 mg kg(-1), 12% at 3 mg kg(-1), and 4% at 5 mg kg(-1)). The method allowed for a distinction to be made between the groups of antibiotics: avoparcin/zinc bacitracin versus tylosin/virginiamycin/spiramycin. This definitely gives added value to the method in the framework of a follow-up of positive screening results by post-screening and confirmatory analysis.

Flubendazole residues in eggs after oral administration to laying hens: determination with reversed phase liquid chromatography.

Flubendazole residues in eggs were experimentally induced by providing groups of 8 laying hens feed with approximately 3, 10 and 30 mg kg-1 flubendazole for 21 days. Eggs were sampled during this period and one week after the administration. Samples of both whole egg and egg white/yolk were analysed separately. Flubendazole analysis was performed by reversed phase HPLC and UV detection at 250 nm (eggs) or 320 nm (feed). The limit of detection (LOD) for flubendazole in feed was 0.3 mg kg-1 and in whole egg 0.012 mg kg-1. Both the hydrolysed and reduced metabolites of flubendazole were also determined quantitatively. The eggs of control hens housed in the same room during the study period did not contain any detectable flubendazole or metabolite residue. The eggs from the lowest dosed group (3 mg kg-1 feed) did contain residues, but most of them were only slightly higher than the LOD. Residues in eggs collected from the laying hens that obtained feed with 10 and 30 mg kg-1 flubendazole reached a plateau level after some 10 days and there was a good dose response relation between levels in feed and those in eggs. The residues of parent compound and metabolites almost exclusively occurred in yolk, the metabolites accounting for some 60-65% of the total residue. The residues of the parent compound and its metabolites declined below 100 micrograms kg-1 5 days after the administration of dosed feed had ended.

Determination of sulphonamides in pork meat extracts by capillary zone electrophoresis.

For sixteen sulphonamides the effective mobility was measured as a function of pH and from the effective mobilities determined in two different electrolyte systems the pK value and mobility at infinite dilution were calculated. A pH of 7.0 was found to be the optimum pH for the separation for both standard mixtures and mixtures of sulphonamides dissolved in pork meat extracts. For the determination of the sulphonamides in pork meat only a very simple pretreatment consisting of extraction with acetonitrile and centrifugation is suitable, as the matrix effects at pH 7.0 do not affect the separation. Calibration graphs for five sulphonamides were constructed, and regression coefficients of at least 0.999 were obtained. The limit of detection for the method varies from 2 to 9 ppm for a pressure injection time of 10 s (injection volume ca. 18 nl) using a Polymicro Technology capillary of length 116.45 cm, distance between injection and detection 109.75 cm and I.D. 50 microns.