Truncated gamma-glutamyl carboxylase in rambouillet sheep.

A flock of Rambouillet sheep was examined because of increased lamb mortality due to ineffective hemostasis at parturition. Decreased activities of coagulation factors II, VII, IX, and X, and severely reduced hepatic gamma-glutamyl carboxylase activity with adequate vitamin K 2,3 epoxide reductase activity was determined.(1,)(21) Parenteral vitamin K(1) supplementation did not improve vitamin K-dependent coagulation factor activities in 3 affected lambs. Affected lamb gamma-glutamyl carboxylase deoxyribonucleic acid was sequenced, and 4 single nucleotide polymorphisms (SNPs 2-5) of the gamma-glutamyl carboxylase gene were identified. Single nucleotide polymorphism-4 results in an arginine to stop codon (UGA) substitution, which prematurely terminates the peptide at residue 686 (R686Stop). This genotype (GATT/GATT) has a strong association with the coagulopathy observed in clinically affected lambs, P < 0.001. The frequency of SNP-3 in exon 11 (R486H) within the MARC 1.1 database is high in the US sheep population overall. Gamma-glutamyl carboxylase activity in hepatic microsomes from a SNP-3 homozygous lamb lacking the SNP-4 mutation (GACC/GACC) was similar to control sheep homozygous for arginine at 486 and also lacking SNP-4 (TGCC/TGCC), indicating that the R486H does not measurably impact gamma-glutamyl carboxylase activity. The remaining two SNPs (2 and 5) are located within non-coding intron sequences. These 4 SNPs allowed for determining the genotype associated with the observed fatal coagulopathy. Screening for the premature truncation (SNP-4) based on the presence of a Bbv I restriction site in clinically normal lambs but not in the homozygous affected lambs allows for detection of the heterozygous state (GATT/GACC), because carrier animals are clinically normal.

Dietary retinol inhibits inflammatory responses of rats treated with monocrotaline.

This study was designed to test the effectiveness of dietary retinol in protecting the heart and lung parenchyma in a monocrotaline model for lung injury and pulmonary hypertension in rats. Male rats were assigned to three groups. Two groups were injected subcutaneously with monocrotaline (17 mg/kg body weight) and fed either the control AIN-93G diet (MC) or the control diet supplemented with retinol (17 mg retinyl palmitate/kg diet)(MR). The third group was fed the control diet and injected with the vehicle only (VC). Four weeks after monocrotaline treatment, the MR group had less thickening of the alveolar septal wall, less myocardial inflammation and degeneration of the right ventricle, and less vascular inflammation in the lung compared with the MC group. The supplemented dietary retinol, however, did not prevent development of right ventricular hypertrophy and did not affect the synthesis and secretion of surfactant phospholipids in type II pneumocytes. The results indicate that dietary retinol suppresses the inflammatory responses in the heart and lungs of rats treated with monocrotaline.

Toxicosis in cattle from concurrent feeding of monensin and dried distiller's grains contaminated with macrolide antibiotics.

Consumption of monensin-containing feed contaminated with macrolide antibiotic residues resulted in the death of cattle from multiple feedlots in south-central Kansas. Cattle were fed milo dried distiller's grains (DDG) with solubles from a common source in conjunction with the ionophore antibiotic, monensin. Deaths occurred as early as 72-96 hours after feeding and were preceded by either no premonitory signs or 1 or more of the following: anorexia, depression, dyspnea, locomotor deficits, and recumbency. Significant gross lesions were pulmonary and mesenteric edema, hepatomegaly, and generalized myocardial and skeletal muscle pallor that was confirmed histologically as acute myodegeneration and necrosis. Other significant histologic lesions included centrolobular hepatocellular necrosis, congestion, and pulmonary interstitial and alveolar edema with fibrin exudation. Animals that survived beyond 6 weeks had poor weight gain and coalescing foci of myocardial fibrosis with residual myocardial degeneration. Analysis of trace mineral supplements for monensin were within the manufacturer's label range. The DDG samples from affected feedlots had 50-1,500 ppm of erythromycin, clarithromycin, and related macrolide antibiotic analogues, which originated in the alcohol residue. In a preliminary feeding trial, cattle fed this contaminated DDG in combination with monensin had clinical signs and died with gross and histologic findings comparable to those of the field cases. Even though rations supplemented with the contaminated DDG contained approved levels of monensin, the clinical and postmortem findings were consistent with those expected for monensin toxicosis. The presence of macrolide antibiotic residues in the contaminated feed appeared to affect the biotransformation of otherwise nontoxic levels of monensin, leading to clinical ionophore toxicosis.

Norepinephrine stimulates in vitro growth but does not increase pathogenicity of Salmonella choleraesuis in an in vivo model.

Norepinephrine stimulates growth of Escherichia coli, Yersinia enterocolitica, and Pseudomonas aeruginosa in serum-supplemented media, and in vivo increases in norepinephrine may be important in the pathogenesis of sepsis by gram-negative bacteria. Because salmonellosis often is associated with stress, the effects of norepinephrine on in vitro growth, and in vivo pathogenicity of the swine pathogen Salmonella choleraesuis were investigated. When RPMI 1640 with and without pig serum was inoculated with fewer than 100 S. choleraesuis/ml and incubated overnight, bacterial numbers were 10(4) to 10(6) lower in RPMI containing serum. Norepinephrine restored bacterial growth in RPMI with serum to normal levels, but it did not increase growth in serum-free RPMI. Similar results were obtained with SAPI, a nutrient-poor medium previously used to study the effect of norepinephrine on growth of gram-negative bacteria. Conditioned media were produced by growing S. choleraesuis in RPMI containing serum with and without norepinephrine and filter sterilizing. Conditioned medium produced with norepinephrine stimulated growth of S. choleraesuis but not E. coli, whereas conditioned medium produced without norepinephrine stimulated growth of both bacteria. To determine the in vivo effects of norepinephrine, rats were implanted with tablets that secrete norepinephrine for 20 to 24 hours or with identical tablets without norepinephrine and infected intraperitoneally with graded doses of S. choleraesuis. The LD-50 of S. choleraesuis was the same in both groups, and norepinephrine did not affect the carrier rate at 30 days after infection. We concluded that although norepinephrine stimulates in vitro growth of S. choleraesuis in serum-based media, the increase in norepinephrine levels in the present in vivo system was probably not sufficient to influence the pathogenesis of S. choleraesuis infection.

Cell-to-cell contact not soluble factors mediate suppression of lymphocyte proliferation by bovine parainfluenza virus type 3.

We have previously characterized the ability of parainfluenza virus type 3-infected (PIV-3) and noninfected bovine alveolar macrophages (BAM) to support lymphocyte proliferation. While uninfected macrophages support proliferation of lymphocytes stimulated with concanavalin A (Con A), ovalbumin, and interleukin 2 (IL-2), lymphocyte [3H]thymidine incorporation was suppressed in the presence of PIV-3-infected BAM. Since viral infection of macrophages has been shown to alter arachidonic acid metabolism and cytokine secretion, we have determined if arachidonate metabolism or the lack of IL-1 and IL-2 mediated the suppression of lymphocyte proliferation by PIV-3. Inhibition of arachidonic acid metabolism failed to reverse the suppressive effect of viral infection as did supplementation of cultures with bovine recombinant IL-1 beta, IL-2, or lymphocyte-conditioned medium. Further, lymphocytes proliferated normally when physically separated from virus infected BAM by a semipermeable membrane. Stimulation of lymphocytes in contact with infected BAM resulted in marked suppression of lymphocyte [3H]thymidine incorporation. Interactions between stimulated lymphocytes and PIV-3-infected BAM resulted in PIV-3 infection of lymphocytes. Virus infection of lymphocytes was confirmed ultrastructurally by the presence of characteristic parainfluenza virus inclusions and virus budding from lymphocyte plasma membranes. It was concluded that suppression of lymphocyte proliferation by PIV-3 is mediated in part by infection of stimulated lymphocytes during cell-to-cell contact with BAM.