Three trials involving 184 male Sprague Dawley rats were conducted to identify the biologically active fraction of endophyte-infected tall fescue seed. In trial 1, seed infected (>95%) with the endophytic fungus Acremonium coenophialum and shown to be toxic to cattle, was serially extracted with hexane, ethylacetate, and methanol, respectively. A second sample of this seed was extracted with chloroform only. Controls included a solvent treatment (spraying the various solvents on endophyte-free seed and carrying the mixture to dryness), the seed post-extraction, endophyte-infected seed and endophyte-free seed. Laboratory chow was mixed with all diets to provide 50% of the mix. Trial 2 examined the same treatments as trial 1 except that the chloroform treatment was omitted. Dosages 3 times (equivalent to-22.5 g seed), those used in trial 1 were applied to endophyte-free seed (7.5 g) in an attempt to accentuate any differences. Trial 3 examined the methanol fraction extracted serially, a methanol fraction extracted batchwise, and a water fraction extracted batchwise. Controls included seed residues post-extraction, solvent residue from methanol sprayed on endophyte-free seed, and endophyte-infected seed. Lowered feed intake and weight gains, and depressed serum prolactin concentrations were used as indicators of toxicity. Feed intake was lower in all three trials for rats fed the methanol fraction. In all three trials mean serum prolactin concentrations in rats fed the methanol extracts were not different from those fed endophyte-infected fescue seed. Rats fed the water extract in trial 3 had lower feed intakes than rats fed endophyte-free seed. However, rats fed the seed, post-water extraction, had serum prolactin concentrations similar to that of the negative control. Results suggest that the toxic factor(s) may be concentrated in the methanol extract.
To compare the recovery rate of morphologically normal and chromatin condensed spermatozoa from native semen samples using the SpermPrep filtration columns and Percoll gradient centrifugation and to determine the influence of the two processing techniques on fertilization and pregnancy rates in an IVF-ET program.Sixteen semen samples obtained from patient's husband were included in this study. Each was divided into two aliquots. The first aliquot was processed with SpermPrep filtration columns and the second, Percoll gradient centrifugation. Smears were made before and after semen processing with both methods for the evaluation of chromatin condensation (chromomycine CMA3) as well as morphology (strict criteria) of spermatozoa. One hundred and seventy oocytes were retrieved from the patients and the oocytes from each patient were subdivided into two sets: one set was inseminated using spermatozoa processed with SpermPrep and the other inseminated after semen processing with Percoll gradient centrifugation.The Percoll method yielded a significantly higher percentage of chromatin condensed (90.8 +/- 6.5% vs 82.3 +/- 8.8%, P = 0.017) and morphologically normal spermatozoa (12.9 +/- 7.4% vs 6.9 +/- 4.8%, P = 0.001) in comparison to SpermPrep. Whereas, sperm count recovery rate was significantly higher after the use of SpermPrep than after the Percoll gradient centrifugation. The fertilization rate was similar between the two methods.Semen processing with Percoll should be recommended for intracytoplasmic sperm injection as the natural selection is bypassed and the SpermPrep technique could be recommended for IVF and IUI programs as the sperm concentration plays a more significant role in these procedures.
