Diagnostic specificity of a real-time RT-PCR in cattle for foot-and-mouth disease and swine for foot-and-mouth disease and classical swine fever based on non-invasive specimen collection.

Foot-and-mouth disease virus (FMDV) and classical swine fever virus (CSFV) are highly contagious and can cause great economic losses when introduced into disease-free regions. Accurate estimates of diagnostic specificity (Sp) are important when considering the implementation of surveillance for these agents. The purpose of this study was to estimate diagnostic Sp of a real-time reverse-transcriptase PCR assay developed for detection of FMDV in cattle and domestic swine and CSFV in domestic swine based on non-invasive specimen collection. One thousand and eighty-eight range beef cattle were sampled from thirteen geographic locations throughout Texas. One thousand and one hundred market hogs and cull sows were sampled. Results for both FMDV and CSFV were considered positive if amplification occurred at or before 40 PCR cycles, inconclusive between 40 and 45 cycles and negative otherwise. Ten cattle had nonspecific PCR amplifications for FMDV, but none were classified as positive and only one as inconclusive. Specificity (95% confidence interval) was estimated as 100% (99.7, 100). There were 19 nonspecific PCR amplifications for FMDV in sampled swine with 1 classified as positive, 6 as inconclusive, and 12 as negative. Specificity (95% confidence interval) was estimated as 99.9% (99.5, 100). There were 21 nonspecific PCR amplifications for CSFV, and 1 was classified as positive. Specificity (95% confidence interval) was estimated as 99.9% (99.5, 100). These assays have high Sp, but nonspecific PCR amplifications can occur.

Effect of recombinant porcine somatotropin on fetal and placental growth in gilts with reduced uterine capacity.

Crowded uterine conditions were induced by unilateral hysterectomy-ovariectomy (UHO) in 42 gilts to determine the effect of recombinant porcine somatotropin on fetal and placental growth. Gilts were randomly assigned across three replicates to one of three treatments: Control (C; n = 14), daily injections of 1 mL saline from d 0 to 64 of gestation, Early (E; n = 12), 5 mg of rpST/d from d 0 to 30, followed by 1 mL saline from d 31 to 64, and Late (L; n = 16), 1 mL saline/d from d 0 to 29, followed by 5 mg of rpST/d from d 30 to 64 of gestation. Blood was collected from each gilt via jugular venipuncture at d 0 and every 15 d thereafter. Gilts were hysterectomized on d 65 of gestation. Length of placental attachment and fetal crown-rump length were measured. Placentas and fetuses were weighed. Placental length, wet weight, and dry weight were recorded. Treatment with rpST (either E or L) increased (P < 0.0001) maternal plasma IGF-I concentrations relative to controls. Treatment with rpST did not affect placental wet weight or placental DNA content. However, E and L treatments increased the percentage of placental protein (P = 0.01) and placental dry matter (P = 0.10) and increased contact area of uterine-placental interface (P = 0.01). Despite changes in placental composition and morphology, weights of fetuses collected from L-treated gilts did not differ from controls, whereas weights of fetuses collected from E-treated gilts tended to be less than controls (P < 0.06). Administration of rpST increased maternal IGF-I concentrations and placental surface area but failed to increase fetal growth in the UHO model. Therefore, mechanisms that are independent of maternal IGF-I or placental contact area may control early fetal growth under crowded uterine conditions.

Birth by caesarian section alters postnatal function of the hypothalamic-pituitary-adrenal axis in young pigs.

Eight crossbred sows were selected for the present study (n = 4 vaginal delivery and n = 4 Caesarian section [C-section]). Gestation length did not differ between vaginal delivery and C-section pigs (113.6 +/- .1 and 113.2 +/- .3 d, respectively; P > .16). Blood and tissue samples from 38 pigs were collected at birth. All remaining pigs were sustained with vaginal-delivery sows until 2 wk of age (n = 39). At 2 wk of age, remaining pigs were catheterized for blood sample collection to assess pituitary-adrenal responsiveness to an injection of corticotropin-releasing hormone (CRH; 10 microg/kg). Blood samples were collected at -30, -15, 0, 5, 10, 20, 40, 60, and 90 min; pigs received CRH or saline at time 0. Pigs were killed and tissue samples were collected immediately following the last blood sample. Serum concentrations of ACTH and cortisol (CS) were measured. Total RNA was isolated from the pituitary and adrenal glands to evaluate gene expression for mRNA specific for pro-opiomelanocortin (POMC) and for the ACTH receptor. Centrifuged clot:blood ratio was reduced in the C-section pigs at birth (P < .001) and at 2 wk of age (P < .043), compared with the vaginally delivered pigs. Basal serum concentration of ACTH was greater in C-section than in vaginally delivered pigs at birth (P = .01) but did not differ at 2 wk of age (P = .42). Basal serum concentration of CS was not different at birth (P = .86) but was greater in C-section pigs than in vaginally delivered pigs at 2 wk of age (P < .04). Serum concentration of ACTH was not different (P > .99) between the two groups of pigs following the CRH challenge. However, serum concentration of CS was greater (P < .05) in C-section pigs following the CRH challenge. Expression of ACTH receptor mRNA tended to be greater in C-section pigs at birth (P < .063) and at 2 wk of age (P < .016). There was no difference in POMC mRNA between treatments (P > .73); however, there was a developmental increase (P < .001) from birth to 2 wk of age. These data indicate that the birth process plays an important role in postnatal function of the hypothalamic-pituitary-adrenal axis in young pigs.

Insulin-like growth factor (IGF)-I, IGF-II, IGF-binding protein-2 and pregnancy-associated glycoprotein mRNA in pigs with somatotropin-enhanced fetal growth.

Fetal growth is increased when pregnant gilts are treated with recombinant porcine somatotropin. The mechanism for increased fetal growth was examined by measuring the expression of IGF-I and -II and IGF-binding protein-2 (IGFBP-2) mRNA in liver and reproductive tissues of somatotropin- and saline-treated pregnant gilts. Twenty-four pregnant gilts received daily injections of either saline (control; n=12) or 5 mg recombinant porcine somatotropin (n=12) from day 30 to day 43 of gestation. Gilts were slaughtered on day 44 of gestation and liver, ovary, placenta, placental uterus (uterus with adjacent placental tissue) and non-placental uterus (region of the necrotic tip) were collected. The mRNAs for somatotropin receptor, IGFs -I and -II, IGFBP-2 and pregnancy-associated glycoprotein (a marker of trophoblast tissue) were analyzed by Northern blotting or ribonuclease protection assay. Gilts treated with somatotropin had heavier fetuses and placentas. The concentration of mRNA for the components of the IGF system was tissue-dependent. The uterine IGF-I mRNA concentration was greater in non-placental than in placental uterus. The greatest IGF-II mRNA concentration was observed in placenta, and adjacent uterine tissue expressed IGFBP-2 mRNA intensely. In non-placental uterus, IGFBP-2 mRNA was nearly undetectable. Somatotropin-dependent regulation of IGF-I was only observed in liver, where the greatest somatotropin receptor mRNA concentration was found. In the pregnant uterus, somatotropin failed to change the concentration of IGF or IGFBP-2 mRNA. Pregnancy-associated glycoprotein mRNA concentration was decreased by somatotropin. In summary, increased fetal growth in somatotropin-treated pregnant pigs was not associated with changes in IGF or IGFBP-2 mRNA concentration in reproductive tissues. Other mechanisms, therefore, lead to enhanced fetal growth in somatotropin-treated pregnant pigs.

Relationships of serum insulin-like growth factor II concentrations to growth, compositional, and reproductive traits of swine.

Insulin-like growth factors I and II (IGF-I and -II) are peptide hormones involved in metabolic regulation of growth. The objective of this study was to determine whether IGF-II concentration was predictive of growth, compositional, and reproductive traits of pigs. Forty male and sixty female pigs, divided equally between two locations, were weighed at 3-wk intervals from birth to 21 wk and bled at 9 and 21 wk of age. At each sampling, two blood samples were collected via jugular venipuncture at an interval of at least 1 h. Serum was separated and IGF-I, IGF-II, and growth hormone (GH) concentrations were determined via RIA. Traits measured included age at puberty and first parity litter size for gilts and backfat and longissimus muscle area. Blood was collected from a random sample of 52 progeny from 13 litters at 9 wk of age and serum was assayed for IGF-II concentrations. Effects of age, sex, location, and pig within sex x location on square-root transformed IGF-II concentrations were determined by analyzing data as a splitplot. Performance traits were fitted to a model including the effects of IGF-II concentration and combinations with IGF-I concentration, sex, location, and interactions. Concentrations of IGF-II were greater at 9 than at 21 wk of age (226.7 vs 159.3 ng/mL, respectively; P < .001) but did not differ between sexes. The correlation between serum IGF-II concentrations assayed from samples collected at 9 and 21 wk was .08. The partial correlations between IGF-I and IGF-II concentrations were .33 and .14 at 9 and 21 wk, respectively. The heritability of IGF-II concentration estimated from offspring-midparent regression was .08 +/- .20. Nine-week IGF-II concentration was positively associated with increased weight from weaning to 12 wk (P < .001). However, the sum of 9-wk IGF-I and IGF-II concentrations had a greater relationship to weight and gain in the growing phase than the concentration of either hormone alone. Concentration of IGF-II at 9 or 21 wk alone did not affect backfat thickness, longissimus muscle area, percentage lean, days to 100 kg, weight at 21 wk, age at puberty, or litter size. The sum of IGF-I and IGF-II concentrations was, however, associated with increased backfat and decreased days to 100 kg.

Responses of porcine corpora lutea to somatotropin administration during pregnancy.

The effects of somatotropin (ST) on functions of porcine corpora lutea (CL) during pregnancy were investigated. Twenty-four crossbred (Yorkshire/Landrace) gilts from d 30 to 43 of pregnancy were injected daily with 5 mg of recombinant porcine somatotropin (rpST; n = 12) or 1 mL of saline (control, n = 12). Blood was collected on d 30, 37, and 43 for analyses of plasma progesterone. Gilts were killed on d 44 of pregnancy, and mRNA were isolated from CL, ovary, and liver. Messenger RNA expression for LH receptor, FSH receptor, ST receptor, 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), and cytochrome P450 side-chain cleavage enzyme (SCC) were measured. Liver, CL, and ovary contained a 4.7-kb mRNA of ST receptor, but the liver contained more mRNA for ST receptor than did CL or ovary (.97 +/- .18, .47 +/- .04, and .25 +/- .04 units, respectively). There were two variants of LH receptor mRNA in CL (6.8 and 4.4 kb). The CL also contained a 1.8-kb mRNA of SCC and a 1.7-kb mRNA of 3 beta-HSD. No FSH receptor mRNA was detected in CL of the pig. The rpST treatment did not affect the mRNA level of ST receptor, 3 beta-HSD, SCC, or 4.4-kb mRNA of the LH receptor. The 6.8-kb mRNA for the LH receptor was decreased (P < .05) by rpST (.56 +/- .04 vs .78 +/- .05 units). Furthermore, concentrations of plasma progesterone decreased (P < .001) in gilts treated with rpST. Decreased luteal function was associated with decreased expression of LH receptor in rpST-treated gilts. The luteotropic effects of ST observed in vitro do not necessarily occur in vivo when gilts are administered rpST during pregnancy.

Effects of exposure to an estrual female on the attainment of puberty in gilts.

A total of 304 prepubertal gilts were randomly allocated to 4 treatment groups across 10 replications for a 50 d treatment period beginning at 170 d of age. The 4 treatment groups consisted of: 1) Gilts that were continuously exposed to one of a group of older, ovariectomized females that had been treated with 2 mg/ml estradiol benzoate to stimulate estrus (SE); 2) Gilts that were continuously exposed to an older, anestrous, ovariectomized female (OVX); 3) Gilts that were exposed to a mature boar for 15 min/d (BE); 4) Gilts that were isolated from any direct physical contact with other pigs (C). A gilt was considered to have attained puberty when she exhibited a standing reflex when mounted by the boar (BE group only) or to pressure applied manually to the back or had plasma progesterone concentrations > 2 ng/ml for 2 consecutive weeks. Data were analyzed as a randomized complete block design with treatment and replication in the model. A higher percentage of gilts attained puberty in the BE group than in the 3 other groups (52 vs 26, 30 and 32%, BE vs SE, OVX and C, respectively; P = 0.002). Gilts exposed to an estrual female or a mature boar attained puberty sooner after treatment was initiated than gilts in other treatment groups (12.6 and 17.8 vs 26.7 and 24.1 d, SE and BE vs OVX and C, respectively; P = 0.0003). Of the gilts attaining puberty during the experimental period, the highest percentage of gilts exhibited estrus within 10 d of treatment in the SE group (55.0 vs 26.1, 37.8 and 16.7%, BE vs SE, OVX and C, respectively; P = 0.05). Age at puberty was also lower SE or BE than OVX or C groups (176.3 and 181.0 vs 189.4 and 188.1 d, respectively; P = 0.0001). Weight at puberty was unaffected by treatment. These results suggest that exposure to an estrual female was effective in stimulating peripubertal females to express estrus, thus reducing the age at puberty. Boar exposure had a stimulatory effect not only at the initiation of exposure but throughout the experimental period, resulting in a higher percentage of gilts attaining puberty.

Effects of recombinant porcine somatotropin on placental size, fetal growth, and IGF-I and IGF-II concentrations in pigs.

The objective of this study was to determine the effects of recombinant porcine somatotropin (rpST) on placental size, fetal growth, and maternal and fetal IGF-I and IGF-II concentrations. Twenty-four pregnant gilts received daily injections of either 1 mL of saline (control) (n = 12) or 5 mg of rpST (n = 12) from d 30 to 43 of gestation. Gilts were slaughtered on d 44 of gestation, reproductive tracts were removed, and fetal weight and length, placental weight, and implantation length were recorded. There was no effect of rpST on fetal or implantation length. Placental weight increased with rpST administration (71.20 +/- 3.52 vs 58.35 +/- 3.41 g; P < .02), as did fetal weight (18.06 +/- .55 vs 16.44 +/- .53 g; rpST vs control, respectively; P < .05). Implantation lengths were partitioned into quartiles to determine the effect of rpST on fetuses with different implantation lengths. The effect of rpST of fetal weight was greater in the first quartile ( < 137.5 mm) than in the fourth quartile ( > 240 mm) (16.04 vs 13.86 g compared with 19.47 vs 18.21 g, respectively). Analysis using a modified Brody curve suggests that the effect of rpST treatment on fetal weight is equivalent to the effect of increasing implantation length by 58.8 mm. Administration of rpST numerically raised IGF-I (P = .07) and IGF-II (P = .12) concentrations in fetal serum. Although maternal serum IGF-I concentrations were similar at d 30, treatment with rpST increased these concentrations over time (77.76, 247.75, 267.85 vs 82.59, 79.59, 77.97 ng/mL on d 30, 37, 43, respectively; P < .001, SE = 14.09). Maternal IGF-II concentrations were also similar at d 30 but decreased over time with rpST treatment (265.78, 219.61, 191.05 vs 285.44, 284.72, 283.05 ng/mL; P < .03, SE = 14.03). Increased maternal IGF-I concentrations may exhibit negative feedback on maternal IGF-II concentrations. The more pronounced effect of rpST on growth in fetuses with shorter implantation lengths suggests that rpST may increase uptake or utilization of nutrients by fetuses. In addition, nutrient transfer across placental membranes may be enhanced by rpST.