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Milk production and lamb growth were characterized in 118 multiparous, 3- to 7-yr-old Rambouillet, Columbia, Polypay, and Suffolk ewes under spring sage range and high mountain meadow grazing from 28 to 98 d postpartum. Daily milk yield as measured by the lamb suckling weight differential technique did not differ (P greater than .05) among breeds, although milk production of Suffolk ewes tended to be higher than that of the other three breeds. Within the Rambouillet, Columbia, and Polypay breeds, total estimated yield of ewes with twins was 13 to 17% higher than that of ewes with singles, whereas in the Suffolk breed, suckling twins increased total milk yield 61% over that of ewes with singles. Twin lambs induced a larger differential in dam milk production in late lactation (70 to 98 d) than in earlier lactation (28 to 70 d). Number of lambs did not influence milk protein, Ca, or P content (P greater than .05). Fat levels in colostrum and 4-d milk were elevated 14 and 20%, respectively, in ewes suckling twins compared with ewes suckling singles. Under range conditions, Suffolk ewes suckling single or twin lambs lost more BW (12 and 21% of 4-d postpartum body weight, respectively) than Rambouillet (4 and 7%), Columbia (5 and 8%), or Polypay (8 and 8%) ewes. Correlation coefficients of milk production and lamb growth rate were positive and significant (P less than .05) up to 56 d of age. Growth rate was less closely associated with milk production for twin than for single lambs.

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Summary. Background: Large animal models that accurately mimic human hemophilia A (HA) are in great demand for developing and testing novel therapies to treat HA. Objectives: To re-establish a line of sheep exhibiting a spontaneous bleeding disorder closely mimicking severe human HA, fully characterize their clinical presentation, and define the molecular basis for disease. Patients/methods: Sequential reproductive manipulations were performed with cryopreserved semen from a deceased affected ram. The resultant animals were examined for hematologic parameters, clinical symptoms, and responsiveness to human FVIII (hFVIII). The full coding region of sheep FVIII mRNA was sequenced to identify the genetic lesion. Results and conclusions: The combined reproductive technologies yielded 36 carriers and 8 affected animals. The latter had almost non-existent levels of FVIII:C and extremely prolonged aPTT, with otherwise normal hematologic parameters. These animals exhibited bleeding from the umbilical cord, prolonged tail and nail cuticle bleeding time, and multiple episodes of severe spontaneous bleeding, including hemarthroses, muscle hematomas and hematuria, all of which responded to hFVIII. Inhibitors of hFVIII were detected in four treated animals, further establishing the preclinical value of this model. Sequencing identified a premature stop codon and frame-shift in exon 14, providing a molecular explanation for HA. Given the decades of experience using sheep to study both normal physiology and a wide array of diseases and the high homology between human and sheep FVIII, this new model will enable a better understanding of HA and facilitate the development and testing of novel treatments that can directly translate to HA patients.

The recent observation of vector sequences in the semen of men undergoing clinical gene therapy for hemophilia has highlighted the need to evaluate the risk of inadvertent germ-line transduction in a clinically relevant animal model. In the present study, we used three different approaches to investigate whether the germ line is at risk of inadvertent alteration following in utero retroviral gene transfer in the clinically relevant, random-bred sheep model. First, we conducted breeding studies. All organs from the 10 resultant offspring were devoid of proviral DNA, suggesting that the germ line had not been altered. As a second approach, we performed PCR on gradient-enriched, forensically purified sperm cells from in utero-transduced rams. The purified sperm cells from 6 of 19 of these rams were PCR positive for provirus, providing compelling evidence that the germ line had been transduced. As a third approach, we performed immunohistochemistry on sections of the testis from in utero-transduced sheep. Numerous somatic cells and very low levels of germ cells within the male reproductive tissues were transduced. In conclusion, our analysis on over 3 x 10(9) sperm cells suggests that the direct-injection approach employed in these studies may result in the inadvertent transduction of very low numbers of male germ cells.