Inducing ovulation with hCG improves the fertility of dairy cows during the warm season

This study was designed to assess the effects of human chorionic gonadotrophin (hCG), given within a timed artificial insemination program, on plasma progesterone concentrations and subsequent fertility in lactating dairy cows during the warm and cold seasons of the year. Cows were treated intramuscularly with GnRH-agonist (Day 0) and PGF₂ (Day 7) followed by either GnRH-agonist (GPG treatment; 60 animals) or hCG (GPH treatment; 60 animals) on Day 9. All cows were fixed-time inseminated (TAI) 16–22 h after the end of treatment. To determine plasma progesterone levels, blood was withdrawn from all animals on Days 3, 6, 9, 12 and 15 after TAI. During the warm period, the pregnancy rate recorded at TAI was similar for the GPG and GPH groups (20% vs. 23%) while the cumulative pregnancy rate within 30 days of TAI was lower (P < 0.05) for the GPG than the GPH group (36% vs. 63%). No differences were observed during the cold period. During the warm period, embryo losses between Days 28 and 45 after TAI were greater (P < 0.05) for the GPG group compared to the GPH group (36% vs. 5%) while again no differences emerged during the cold period. Mean plasma progesterone levels were higher (P < 0.05) in the GPH group than GPG group on Days 3, 6 and 9 post-insemination. Our findings indicate that the use of hCG to induce ovulation in a timed artificial insemination protocol increases plasma progesterone levels and improves fertility in dairy cows during the warmer period of the year.     

Parity of female goats does not influence their estrous and ovulatory responses to the male effect

The objective of the present study was to determine whether parity is a factor that influences the estrous and ovulatory responses of female goats when they are stimulated by males that show increased sexual activity. To stimulate sexual activity, four adult male goats were subjected to photoperiodic treatment for 2.5 months comprising long days, with the treatment commencing on 1 November. On 14 April at 1900 h, a group of multiparous females (n = 21) and a group of 16 months-old nulliparous females (n = 19) were exposed to four bucks (two per group) for 15 days. Throughout the study period, the estrous behavior of these female goats was detected twice on a daily basis. Ovulations of the female goats were determined by ecography on days 7 and 18 after exposure to males. The sexual behavior of males was recorded twice every day from 0800 to 0900 h and from 1730 to 1830 h during the first 4 days after introduction in the pen of females. The total cumulative proportion of multiparous females that had ovulations (100%) and displayed estrous behavior (100%) during the 15 days of exposure to males did not differ (P > 0.05) from that of nulliparous females (100% and 95%, respectively). The interval between introduction of males and onset of estrous behavior did not differ (P > 0.05) between multiparous (1.9 ± 0.1 days) and nulliparous (1.7 ± 0.2 days) females. The proportion of females displaying a short estrous cycle was greater (P < 0.05) in multiparous (13/21, 62%) than in nulliparous (5/19, 26%) females. Duration of these shorter than typical estrous cycles did not differ (P > 0.05) between groups (multiparous: 5.2 ± 0.3 days, nulliparous: 4.5 ± 0.1 days). The number of anogenital sniffings was greater (P < 0.001) in males exposed to nulliparous than in those exposed to multiparous females. In contrast, the number of mounting attempts was greater (P < 0.01) in males that were introduced to multiparous than in those that were introduced to nulliparous does. The number of flehmen, nudging, self-marking with urine, and mounts was not different (P > 0.05) between males that were in contact with multiparous and nulliparous females. These results indicate that regardless of parity, female goats respond to male introduction if they are stimulated by males that were previously exposed to artificial long days to increase their sexual behavior.     

Effect of strategic supplementation with multi-nutrient urea molasses blocks on body weight and body condition score of Lohi sheep owned by tenants of Pakistan

The effect of strategic supplementation with multi-nutrient urea molasses blocks (MNUMB) on BW and body condition score (BCS) in Lohi ewes (treated, n = 514) during late gestation and lactation was compared with those (control, n = 391) grazing on only post-harvest crop residues and road side in the irrigated district of Okara in central Punjab (Pakistan). Analysis of variance revealed highly significant (P < 0.01) differences in body weight (BW) and body condition score (BCS) of ewes of various ages with different reproductive status and seasons under both flocks. Analysis of variance also revealed a significant interaction (P < 0.05) between reproductive status and seasons in favor of BCS. Ewes aged 48 months in average constituted the highest (34.5% and 35.6%), whereas those aged 60 months had the smallest (10% and 4%) proportion in the control as well as in the treated flock. Mean BW and BCS in ewes of control flock was 33.5 and 2.08 kg, and lower (P < 0.05) than 35.0 and 2.31 in ewes in the treated flock, respectively. Ewes aging 12, 24 and 36 months treated with strategic supplementation of MNUMBs were not only heavier (P < 0.01) but also had highest BCS of 2.34. Lactating ewes constituted highest proportion (39%, 51%) followed by pregnant (35%, 32%) ewes in both flocks, respectively. Proportion of dry (16%) and freshly conceived (9.5%) ewes tended to be higher in the control than in the treated flock. BW was 8–11% higher (P < 0.01) in pregnant than in lactating or dry ewes in both flocks with similar BCS. Seasons of autumn and summer were found to affect BW more (P < 0.01) than BCS. Pregnant ewes in treated flocks had gained highest BW, 10–12% higher than ewes under control (P < 0.01) or than non-pregnant ewes (P < 0.05) but lost at a rate of 5–6% at lambing. BW in lactating ewes in treated flock was higher (P < 0.01) than ewes in control. Lambs suckling ewes with strategic supplementation of MNUMBs grew at a faster rate (122 g/day) with 10–15% higher survival rate than those (97 g/day) in the control flock during lactation of 16 weeks but non-significantly. Based on this improvement it can be concluded that supplementation with appropriate sources of energy and N exerts desirable effects on the traits of economic importance in sheep.     

Development to the blastocyst stage of immature pig oocytes arrested before the metaphase-II stage and fertilized in vitro

In vitro fertilization (IVF) and embryonic development of mature and meiotically arrested porcine oocytes were compared in the present study. After in vitro maturation (IVM) of cumulus-oocyte complexes for 48 h, 75.4% of them extruded a visible polar body (PB). Most of the oocytes with a first polar body (PB+ group) were at the metaphase-II (M-II) stage (91.4%). Most of the oocytes without a visible polar body (PB− group) appeared to be arrested at the germinal vesicle (GV) (41.6%) and metaphase-I (M-I) (34.0%) stages. After IVF of oocytes (day of IVF = Day 0), there was no difference between PB⁺ and PB⁻ groups in rates of sperm penetration, mono-spermy, however oocyte activation rate after penetration was greater in the PB+ than in the PB− group (P < 0.05). On Day 2, there was no difference between rates of embryos cleaved at the 2–4 cell stages in PB+ and PB− groups (42.1 ± 48.8% and 33.6 ± 2.1%, respectively). On Day 4, the rate of PB+ embryos developing beyond the 4-cell stage was greater than that of PB− embryos (P < 0.05, 31.7 ± 3.9% and 14.1 ± 1.5%, respectively), and PB+ embryos had more cells than the PB− embryos (P < 0.05, 8.3 ± 0.4 and 6.0 ± 0.8 cells, respectively). On Day 6, a greater proportion of PB+ embryos developed to the blastocyst stage than did PB− embryos (P < 0.05, 34.6 ± 2.4% and 20.7 ± 2.8%, respectively). However, when the GV oocytes of the PB− group were not included in recalculations, there was no difference in blastocyst rates between M-I arrested and M-II oocytes (35.3 and 34.6%, respectively). The number of blastomere nuclei in embryos obtained from the PB+ group (52.0 ± 2.5) was greater than that from the PB− group (P < 0.05, 29.1 ± 2.8). The proportion of degenerated parts in the blastocysts, as determined by morphological appearance, was the same in the PB+ and PB− groups. Although the quality of PB+ embryos was enhanced as compared with that of the PB− group, the proportion of inner cell mass and trophectoderm cells in PB+ and PB− blastocysts did not differ (1:1.9 and 1:2.2, respectively). Chromosome analysis revealed that PB+ blastocysts had more diploidy (P < 0.05, 69.7%) than did PB− blastocysts (44.0%), whereas PB− blastocysts had more triploid cells (P < 0.05, 34.0%) than did PB+ oocytes (8.4%). These results indicate that pig oocytes arrested before the M-II stage (M-I oocytes) undergo cytoplasmic maturation during maturation culture and have the same ability to develop to blastocysts after IVF as M-II oocytes, but some of them resulted in degeneration or delayed development with poor embryo quality.     

Evaluation of sperm functional attributes in relation to in vitro sperm-zona pellucida binding ability and cleavage rate in assessing frozen thawed buffalo (Bubalus bubalis) semen quality

The objective of this study was to evaluate sperm functional attributes in relation to in vitro sperm-zona binding ability and cleavage rate in assessing frozen thawed buffalo (Bubalus bubalis) semen quality. Frozen-thawed forty-eight ejaculates from eight Surti buffalo bulls (six ejaculates/bull) obtained by artificial vagina were used. Frozen semen from each bull was thawed, pooled, and subjected for sperm functional (six replicates) and in vitro fertilization (four replicates) tests. The progressive forward motility, plasmalemma functional integrity assessed by fluorogenic [6-carboxyfluorescein diacetate (CFDA), and propidium iodide (PI)], hypoosmotic swelling (HOS), and hypoosmotic swelling-Giemsa (HOS-G) test, mitochondrial membrane potential, sperm nuclear morphology, the number of sperm bound to zona and cleavage rate differed significantly (P < 0.05) between bulls. When the animals were grouped based on cleavage rate (group I, >40% cleavage rate, n = 5, and group II, <40% cleavage rate, n = 3), in vitro fertility parameters and all the sperm functional attributes except sperm nuclear morphology differed significantly (P < 0.05). The proportions of sperm with functional plasmalemma in the tail and intact acrosome assessed by HOS-G test (25.33, range: 17.48–40.27) were significantly (P < 0.001) lower than the functional plasmalemma in the tail assessed by HOS test (39.80, range: 27.85–54.67). The number of sperm bound to zona had significant correlations with the mitochondrial membrane potential (r = 0.90, P < 0.01) and plasmalemma integrity (fluorogenic, r = 0.74 and HOS, r = 0.79, P < 0.05) and HOS-G, r = 0.87, P < 0.01). The cleavage rate had significant (P < 0.05) correlations with the mitochondrial membrane potential (r = 0.70) and plasmalemma integrity measured by HOS-G test (r = 0.68). The present study indicates that these attributes could represent important determinants of buffalo sperm quality influencing cleavage rate.     

Coincident increases in oxytocin receptor expression and EMG responsiveness to oxytocin in the ovine cervix at oestrus

This study has localised oxytocin receptor (OTR) mRNA expression within the cervix of non-pregnant ewes and related this to changes in the sensitivity of the cervical musculature to administered oxytocin (OT) during the oestrous cycle by recording electromyographic (EMG) activity. Cervices were collected from 34 ewes at specified time points throughout the cycle. OTR mRNA expression was localised by in situ hybridisation and results were expressed as optical density measurements from autoradiographs in each of four different cervical regions. EMG recordings were made for up to 8 h per day from four non-pregnant ewes undergoing seasonal oestrous cycles between Days −3 and +3 relative to oestrus (Day 0). The highest concentrations of OTR mRNA were detectable within the luminal epithelium (LE) of the cervix, with values increasing from Day 15 of the cycle, peaking during the follicular phase (P<0.001, compared to the mid-luteal phase) and returning to basal by Day 2. There was a small but significant increase in OTR mRNA hybridisation (above basal/luteal phase values) within the stromal cells (STR) adjacent to the lumen (P<0.05) during the same time period, but no differences from basal values were detectable in the dense collagenous annular ring or in tissue superficial to this. Analysis of pooled EMG activity recorded daily from the cervix indicated that endogenous contractile activity was higher on Day 0 than on the Days +1 (P<0.05), −2, +2 and +3 (P<0.001). The response to bolus intravenous (i.v.) injections of 25 mU OT (25 mU) varied with day of the cycle. This dose produced a measurable and significant response on Days 0 (P<0.001) and +1 (P<0.001), but not on any of the other days, indicating that the sensitivity of the cervical musculature to OT peaked on these days. These data show that the cervix is highly responsive to OT at oestrus. This coincides with an increase in OTR mRNA expression in the luminal epithelial cells, suggesting the likely production of an intermediary messenger between the epithelial and smooth muscle cells.     

Behaviour of Merinos divergently selected for multiple rearing ability in response to external stimuli

Data of Merino sheep descended from two divergent selection lines for (H line) and against (L line) multiple rearing ability were collected in three different trials to compare response to stressful situations between lines. Trial 1: the behaviour of 138, 20-month-old sheep born in 2001 and 124, 8-month-old sheep born in 2002 were observed in an arena test. Experimental animals amounted to 111 and 101 for the H line in the respective years. Corresponding numbers for the L line were 27 and 23, respectively. Progeny born in 2001 were accustomed to handling, as the tests were performed shortly after routine performance recording. Lambs born in 2002 were unaccustomed to handling when tested. Progeny from the H line born in 2002 allowed a smaller (P < 0.01) minimum distance between them and the human operator seated between them and a group of similar aged sheep than L line contemporaries. Progeny born in the L line during 2002 defecated more (P < 0.05) frequently than H line contemporaries. No line differences were found in progeny born during 2001. Trials 2 and 3 were designed to test whether the putative line difference in stress response would impact on physical meat quality. Trial 2: a total of 103, 18–20-month-old progeny from the two selection lines were slaughtered during 2001, 2002–2003. Mean slaughter weight, dressing percentage and carcass weight of H line animals were between 5 and 13% higher (P < 0.05) than that of L line animals. There were no differences in cooking loss, drip loss and tenderness between the two lines. As pertaining to the meat colour, L line progeny had less vivid red meat, as seen by their lower (P < 0.05) a* colour reading and chroma values. The L line animals had lower (P < 0.01) initial pH values as measured in the Musculus longissimus dorsi. These results are indicative of susceptibility to acute, short-term stress. Trial 3: 20 progeny of the two selection lines were slaughtered and their muscle pH measured at 11 intervals from 45 min to 48 h post slaughter. As in Trial 1, the initial pH of the L line animals were lower (P < 0.05) than that of H line animals. An opposite tendency was observed by 48 h post slaughter. The obtained results support a hypothesis that divergent selection for ewe multiple ability resulted in adaptations to cope with unfamiliar conditions.     

Interaction of calf suckling, use of oxytocin and milk yield with reproductive performance of dairy buffaloes

Calf suckling and oxytocin injections are commonly used for pre-milking stimulus in dairy buffaloes under field conditions. A study was conducted to investigate effect of these treatments on reproductive performance. Fifty one Nili-Ravi buffaloes were monitored from parturition up to 150 days postpartum through rectal examination. Data on milk yield, body condition score (BCS) and reproductive parameters were recorded weekly. Postpartum ovulation interval (POI) was determined by presence of an ovulation depression or a very soft corpus luteum haemorrhagicum and was confirmed through milk progesterone levels (MPL). Suckling was used to stimulate milk let down, and where the calf had died, injection of oxytocin was resorted to. Milk samples were analyzed for MPL using radioimmunoassay (RIA) and fat; and milk yield was converted to 4% fat corrected milk (FCM). The mean postpartum uterine involution length (PUI) was 34.30 ± 1.33 days. Mean POI was 59.37 ± 4.76 days and mean postpartum estrus interval (PEI) was 69.03 ± 6.03 days. Suckling period averaged 26.40 ± 5.57 days and correlated with POI (r = 0.19, P < 0.01) and PEI (r = 0.23, P < 0.01). POI was shortest in buffaloes suckled for one month (P < 0.05). Oxytocin was used with a mean dosage of 7.50 IU, delaying placental expulsion time (PET) and POI but shortening PEI. BCS shortened PET, POI and PEI (P < 0.01). Mean FCM was 14.50 ± 0.20, ranging from 2 to 35 kg/d; and was higher in estrus group; correlating positively with POI (r = 0.31, P < 0.01). MPL were 1.37 ± 0.17 ng/ml and increased after ovulation, remaining greater than 1.5 ng/ml from Day 4 to 14 of the estrus cycle, followed by a rapid decline up to next estrus. BCS in buffaloes resuming oestrus was constantly higher than those failing to resume ovarian cyclicity. Live weight, prepartum was 510.0 ± 5.9 kg with a loss of 3.7 ± 2.12 kg, 30 days postpartum. The present study suggests a lower reproductive efficiency of dairy buffaloes under the peri-urban farming system reflected by ovarian cyclicity in 68.63% buffaloes within 150 days postpartum and silent estrus in 51.5% of the cases. Increasing suckling duration and use of oxytocin delayed POI, however, POI was shortest in buffaloes suckled for one month. The high yielding buffaloes also manifested better reproductive cyclicity; while moderate yielder showed shorter ovulation intervals and higher conception rate.     

Effects of Tasco (a seaweed extract) and heat stress on N metabolism and meat fatty acids in wether lambs fed hays containing endophyte-infected fescue

Wether lambs (n = 27, average BW = 40 kg) were used to test response to forage treated with Tasco-Forage (an extract of the brown kelp Ascophyllum nodosum) prior to conserving, or to direct feeding of the extract (Tasco-EX). Hays made from endophyte (Neoyphodium coenophialum)-infested tall fescue (Festuca arundinacea)-based pasture received 0 or 3 kg of Tasco/ha prior to harvest. Lambs, blocked by weight, were randomly allotted to three diets: (1) control hay, (2) treated hay, and (3) #1 + Tasco-EX fed at 1% of the diet. Hays were low in CP (<7%) so all lambs were fed soybean meal (12% of the diet) in addition to trace mineralized salt. Diets were fed at 1.5% BW to prevent refusals. Total collections (7 d) were made during periods without or with applied heat stress. After each period, rumen contents were obtained to determine pH, NH₃ and VFA. Lambs were sacrificed post-trial. A subset was used to evaluate sensory traits and muscle fatty acids. Lambs were in negative N balance during the study and Tasco treatments did not affect N metabolism. Fecal N tended (P < 0.10) to increase with short duration heat stress causing a concomitant decrease (P < 0.05) in apparent N digestibility (58.6 versus 56.1%; S.E. = 0.7). Urinary N loss decreased (P < 0.001) with heat stress (8.0 versus 5.9 g/d; S.E. = 0.2), resulting in increased (P < 0.001) N retention (−2.1 versus −0.3 g/d; S.E. = 0.2). Apparent OM digestibility was not affected by heat stress but was greater (P < 0.05) for lambs fed Tasco-EX treatment than those fed treated hay. Treatment diets decreased (P < 0.05) ruminal butyrate. Heat stress increased (P < 0.05) acetate and total VFA and decreased (P < 0.01) ruminal pH. A tendency (P < 0.11) of increased 14:1ω5, decreased (P < 0.05) 18:0 and total saturated fatty acids in muscle was observed with Tasco diets. Meat sensory characteristics were not affected by treatment. Tasco may alter some aspects of rumen or lipid metabolism but has no effect on N metabolism or meat sensory characteristics of sheep fed restricted, low-quality diets.     

Prolactin regulation of testicular development and sexual behavior in yearling Suffolk rams

The aim of this study was to determine how the yearly prolactin rhythm might affect the sexual development of Suffolk rams (latitude 50°N). Five rams were injected daily with bromocriptine (35–45 μg kg⁻¹ body weight) for 1 year, beginning in January (early winter) when rams were 11 months of age. Five control rams each received daily injections of the vehicle. In the controls, blood prolactin was <7.5 ng ml⁻¹ in winter, increased (P < 0.01) to a peak of 172.6 ± 11.9 ng ml⁻¹ after the spring equinox, and remained high during summer before declining (P < 0.01) to 29.6 ± 6.6 ng ml⁻¹ at the autumn equinox. Suppression of the seasonal rise in prolactin secretion with bromocriptine slowed testicular growth (50%; P < 0.05) in April and May (spring), thus delaying the time of peak testis size and sperm production by 1 month. Serum testosterone level was lower (50%; P < 0.01) in the treated rams than the controls in June and July (early summer), due mainly to reduced stimulation of the testes by smaller (P < 0.01) LH pulse releases or to smaller (P < 0.01) testosterone responses to LH releases, respectively. Suppression of prolactin also seemed to disrupt the central activation of gonadotropin secretion in that seasonal increases in serum FSH level and LH pulse amplitude and frequency were unusually slow (P < 0.05). These anomalies did not affect testis growth, which was normal from June until development was complete. Rams were sexually inexperienced when libido was first tested in July (non-breeding season). Both groups were equally capable of learning and expressing sexual behavior (i.e. normal mounting and ejaculation frequencies), which was more intense in September (breeding season; P < 0.05). Results support the hypothesis (based on the location of prolactin receptors) that the spring increase in prolactin secretion could target both the testes and the hypothalamic–pituitary system and be involved in the seasonal regulation of sexual function in the young adult Suffolk ram.     

Control of postpartum anestrous with an intra-vaginal progesterone device plus eCG or calf removal for 120 h in suckled crossbred cows managed in a pasture-based system

To control postpartum anestrus and reduce calving to conception interval, 167 crossbred non-pregnant cows that were 90–130 days postpartum were allotted randomly to one of the following treatments: PH (n = 59), intra-vaginal sponge with 250 mg of medroxyprogesterone acetate (MAP) for 7 days plus 50 mg of MAP and 5 mg 17-β estradiol (17β-E) in the first day of treatment (day −8), 500 UI eCG (day −3) and 1.5 mg 17β-E in 24 h after sponge removal (day 0); CR (n = 57), temporary calf removal for 120 h; CG (n = 51), control group without treatment. Estrus rate differed among treatments (P < 0.01) being greater in PH (78.2%), followed by CR (52.0%) and CG (22.9%). A greater proportion of cows in the PH (80.0%) and CR (54%) groups had ovulations when compared to CG (35.4%). Intervals to first estrus were 13.5 ± 6.3 days, 26.1 ± 6.4 days and 52.5 ± 7.5 days for the PH, CR and CG groups, respectively. First insemination conception was similar in the three groups. Postpartum intervals to first breeding (PFS) and to conception (PCI) were longer in CG than PH and CR groups (P < 0.05; P < 0.01). The PH and CR groups had a similar PFS but PCI was different (P < 0.02). Accumulated pregnancy rate at 30 and 60 but not at 90 days were different (30 days: P < 0.09; P < 0.01; P < 0.09; 60 days: P < 0.06; P < 0.01; P < 0.03) among treatments. After 90 days post-treatment, 9%, 18% and 33% of cows from the PH, CR and CG groups had not conceived. Similarly, 5.4%, 6.0% and 12.5% of cows from the PH, CR and CG groups, respectively, were culled from the herd because of lack of pregnancy after 180 days post treatment. In the group of cows evaluated by ultrasonography, only those cows having larger ovaries and dominant follicles had ovulations. It was concluded that the hormonal treatment was more efficient in inducing a fertile estrus and reducing calving to conception interval followed by the calf removal for 120 h. Each method can be considered as an important tool to reduce the postpartum anestrous period in dual purpose herds when AI is conduct in the tropics.     

Effects of ileal endogenous nitrogen losses and dietary amino acid supplementation on nitrogen retention in growing pigs

The relationship between ileal endogenous nitrogen (N) losses and N retention was studied in two experiments with growing pigs of 40 to 60 kg. In Experiment 1, 13 ileal cannulated castrated males were fed diets based on maize starch, containing either soyabean meal (SBM) with a low trypsin inhibitor activity (TIA), a mixture of toasted and untoasted soyabean meal with a high TIA (mSBM), a commercial batch of peas, or rapeseed expeller cake (RC). Ileal endogenous N recovery was measured using the ¹⁵N-isotope dilution technique. Apparent ileal digestibility of crude protein (CP) for the SBM, mSBM, pea and RC diets were 82.8, 72.0, 76.7 and 68.7% (P < 0.05). True ileal CP digestibility for the diets was 96.5, 93.0, 94.0 and 87.5% (P < 0.05), and the recovery of ileal endogenous N was 3.08, 6.01, 4.55 and 5.36 g/kg DMI (P < 0.05). In Experiment 2, sixteen castrated males were used to determine N retention, using almost similar diets as in Experiment 1. The diets contained either SBM, mSBM or peas and were balanced for the contents of apparent ileal digestible (ID) CP (96 g/kg) and ID essential amino acids (EAA; at least 85% of requirement values). The fourth treatment was a diet with mSBM as protein source, but supplemented with EAA to the level of 95% of the requirement values (diet mSBMs). Apparent faecal CP digestibility for the SBM, mSBM, pea and mSBMs diets was 88.6, 87.2, 86.1 and 86.0% (P < 0.05). Urinary N excretion and N retention for these treatments were 0.39, 0.59, 0.40, 0.53 (P < 0.05) and 0.87, 0.80, 0.85, 0.84 g/kg^0.75/day (P < 0.05), respectively. Utilization of dietary ID N for N retention were 79.8, 73.3, 78.2 and 77.6% (P < 0.05), respectively. The study showed that increased ileal endogenous N losses are associated with higher losses of urinary N and with a lower N retention. Supplementation of extra essential AA to a diet causing a relatively high flow of ileal endogenous N, may compensate for the lower N utilization under these conditions, and thus limit effects on N retention.     

Surgical castration of pigs affects the behavioural response to a low-dose lipopolysaccharide (LPS) challenge after weaning

The objective of the present study was to evaluate the effect of an acute stressor in the early postnatal life of pigs, surgical castration, on post-weaning behaviour, and on the behavioural, endocrine and immune responses elicited by a low-dose lipopolysaccharide (LPS) challenge after weaning. At 5-days-of-age, 64 male piglets were randomly assigned to undergo surgical castration or were left untreated (treatment). Pigs were weaned at 28 days-of-age. Behaviour post-weaning and mixing was assessed during a 1-h period, during which agonistic interactions were recorded. One day post-weaning, pigs were injected intraperitoneally with a single dose of 0 or 5 μg/kg of BW of LPS from Escherichia coli (challenge). Sickness behaviour was studied by scan sampling every 5 min for 45 min at 0, 1, 2, 3, 4, 6 and 8 h after the challenge. Blood samples were taken at 0, 2, 12 or 24 h after injection and were analysed for plasma concentrations of tumor necrosis factor-alpha (TNF-), interleukin-1beta (IL-1β), C-reactive protein (CRP), serum amyloid A (SAA) and cortisol. Results showed that non-castrated pigs were more aggressive than castrated pigs immediately after weaning (P < 0.05). Administration of LPS provoked behaviours characteristic of sickness including a reduction in general activity, as well as decreased eating and exploratory behaviours (P < 0.05). These altered behaviours occurred predominantly 3-h post injection (P < 0.05). Significant treatment by challenge interactions showed that castration reduced the occurrence of sickness behaviours induced by LPS, such as depressed general activity (P < 0.01), anorexia (P < 0.01) and reduced exploratory behaviours (P < 0.05). LPS administration increased TNF- levels (P < 0.05), with peak concentrations 2 h after injection (P < 0.01). CRP levels of LPS-treated pigs were higher than saline-treated animals at 12 h (P < 0.05). LPS administration tended to increase plasma SAA levels (P < 0.1), but did not increase cortisol levels (P > 0.1). However, castration did not affect the response of pro-inflammatory cytokines, acute phase proteins and cortisol to the challenge. These results show that surgical castration reduces aggressiveness at weaning and affects specific sickness behaviours but not the endocrine and immune responses elicited by low-dose endotoxin challenge in weaned pigs.     

Association between numbers of ovarian follicles in the first follicle wave and superovulatory response in ewes

The aim of this study was to examine the variability in the number of ovarian follicles in sheep and to determine if the average number of follicles per day influences the response to superovulation and resulting embryo quality. Ewes (n = 83) were synchronized and the number of follicles (≥2 mm diameter) in the ovaries were counted daily between Days 0 and 4 of the oestrous cycle using transrectal ultrasonography. Fourteen to 21 days later, 47 ewes were randomly chosen from the group and were treated with an intravaginal progestagen pessary for 12 days and superovulated with 1500 IU eCG administered as a single injection 10 days after sponge insertion. Ewes were mated and reproductive tracts were recovered after slaughter on Day 6 of pregnancy. The number of corpora lutea was counted, uterine horns were flushed and the morphology and developmental stage of the recovered oocytes/embryos was assessed. The mean daily number (±S.D.) (≥2 mm diameter) of follicles per ewe was 8.5 ± 2.8 (ranging between 3 and 16). After superovulation animals with few follicles (Low group: <8 follicles/day; n = 21) had fewer (P < 0.005) corpora lutea, total structures (unfertilized oocytes and embryos), good quality and total embryos compared to animals with many follicles (High group: ≥8 follicles/day; n = 23). No difference was found in the proportion of good quality embryos (relative to the total number; Low 0.68 ± 0.11 versus High 0.79 ± 0.08; P = 0.21) between the two groups, or the recovery rate, the number of unfertilized oocytes or the number of poor quality embryos per animal. We conclude that ewes with a higher number of follicles (≥8) during the first follicular wave had a better superovulatory response (in terms of corpora lutea and high quality embryos) 2–3 weeks later; however, there was no relationship between the number of follicles and the proportion of good quality embryos per animal.     

Fattening Pelibuey lambs with sugar cane tops and corn complemented with or without slow intake urea supplement

One hundred and eighty Pelibuey lambs were fattened for 120 days with three treatments evaluating in situ DM disappearance, voluntary DM or OM intake, rumen degradation, rate of passage, NH₃ and VFA concentration, apparent digestibility, ruminal pH, total fermentable carbohydrates and weight gains. The first diet (T1), 60 lambs (25.7 ± 0.7 kg BW) plus two cannulated sheep were fed 100% sugar cane tops (SCT) per day; treatment 2 (T2), 60 lambs (25.4 ± 0.4 kg BW) plus two cannulated sheep diet was SCT supplemented with 200 g DM of slow intake urea supplement (SIUS) (a 4% urea mixture) per day. Treatment 3 (T3), 60 sheep (25.5 + 0.5 kg BW) plus two cannulated animals, fed SCT and corn crop chopped (C) mixture (40:60%) were supplemented with 200 g SIUS per day. Ruminal kinetics were determined thereafter in four adult ruminal cannulated sheep in metabolic cages. A 100% corn diet was added to the second trial (T4). At all times, fibrous forages were available and exceeded VDMI. Daily BW gain by SCT animals was 70 g/d compared to 135 g/d for SCT/SIUS and 218 g/d for SCT/C/SIUS diets (P < 0.05). Total forage intake was similar for T1 and T2 treatments and higher for T3 (P < 0.05). Kinetics observation showed a total DMI variation increasing from 474 g/d for T1; 597 g/d for T4 to 797 g/d for T2, and 917 g/d for T3, being different for the last two treatments compared with the first two (P < 0.001) and similar among SIUS diets. Rumen pH diminished from 6.8 to 6.7 at 2 h, when SIUS was offered, and stayed above 6.5 during the 12 h of sampling when SIUS was the only supplement and 6.3 when corn was added, while in SCT lambs, ruminal pH decreased to 6.2 by 6 h and rose again to 6.4 after 12 h favoring bacterial multiplication. Ammonia concentration and digestion of potential digestible and indigestible fractions were significantly augmented (P < 0.05) by SIUS. In vivo nitrogen digestibility was different (P < 0.05) among diets: T1, T2, T3, and T4 (36.5%, 74.1%, 76.5%, and 39.4%, respectively). In vivo digestibility of DM, OM, cellulose and hemicellulose was similar among T2 and T3 groups but different when T1 or T4 diets were offered (P < 0.05). NDF digestibility was higher for T3 (77.1%) and T2 (75.1%) but different from T1 (57.1%) and T4 (59.1%) (P < 0.01). In situ DM disappearance did not show differences among T2 and T3 diets at 9, 12, 24 and 48 h of incubation but differed in percentage for T1 and T4. Digestion rate constant (k_d) was similar among SIUS diets 0.038 and 0.039, but different from T1 and T4 diets 0.023; 0.021 (P < 0.05). True digestibility of NDF was higher (P < 0.05) in SCT/SIUS/C (32.5%) and SCT/SIUS (32.7%) compared with SCT (26.2%) and C (27.4%). Passage of NDF was different (P < 0.05) between T1 (0.059 h⁻¹) and T4 (0.061 h⁻¹) from 0.079 (T2) and 0.081 k_p/h (T3). The half-time (t_1/2) disappearance for hemicellulose was higher (P < 0.05) for SCT/SIUS (22.1 h) and SCT/SIUS/C (29.2 h) as compared to SCT (18.1 h). Kinetic performance among SCT and C diet did not have a statistical difference. Utilization of fibrous forages by lambs was higher (P < 0.05) for SIUS diets. VDMI and apparent digestibility showed better performance for DM and OM, lower acetate molar proportions and greater feed intake by both SIUS diets which was reflected in body weight gain.     

The relationship between heart rate and energy expenditure in Alpine, Angora, Boer and Spanish goat wethers consuming different quality diets at level of intake near maintenance or fasting

Six Alpine (AL; 38.4 ± 3.0 kg), Angora (AN; 23.1 ± 2.7 kg), Boer (BO; 40.8 ± 4.5 kg) and Spanish (SP; 33.6 ± 2.2 kg) wethers (1.5 yr of age) were used to determine the effects of time of the day and potential interactions between time, genotype and diet quality on energy expenditure (EE), heart rate (HR) and EE:HR when fed near maintenance and fasting. The experiment consisted of four simultaneous crossovers, with 21 d for adaptation before measures. Diets were 60% concentrate (CON: 15% CP) and ground alfalfa hay (FOR: 23% CP), offered in two meals at 8:00 and 16:00 h. Energy expenditure was determined from O₂ consumption and production of CO₂ and CH₄ over 2-day periods in fed and fasting states (total 4-day fasting period). Fasting EE was higher during the day than night, with values generally highest at 16:00–17:00 h. Animal within breed affected EE, HR and EE:HR (P < 0.05). The diurnal pattern in EE varied with diet (P < 0.05), although total daily EE was not different between diets. Before the morning meal, there were a number of hours during which EE was greater for CON than for FOR. However, at both meals the rise in EE was considerably greater for FOR versus CON, lasting for 3–4 h. The same general pattern in HR was observed, although the period of time when there was a dietary difference after the afternoon meal was shorter. For both fed and fasted goats, EE:HR differed among hours of the day (P < 0.05). EE:HR tended (P < 0.09) to differ between diets (5.99 and 6.21 for CON and FOR, respectively) and to be affected (P < 0.09) by an interaction between breed and diet (AL: 5.84 and 6.38; AN: 5.91 and 5.73; BO: 6.05 and 6.58; and SP: 6.17 and 6.15 kJ/(kg BW^0.75 × day):heart beats/min) for CON and FOR, respectively. In conclusion, for use of HR to predict EE by goats, it appears desirable to determine the ratio of EE:HR with a diet similar to that consumed during prediction and over an extended period of time.     

Dynamics of indirect symptoms of skeletal muscle damage after stretch-shortening exercise

Healthy untrained men (age 20.4 ± 1.7 years, n = 20) volunteered to participate in an experiment in order to establish dynamics of indirect symptoms of skeletal muscle damage (ISMD) (decrease in maximal isometric voluntary contraction torque (MVCT) and torque evoked by electrostimulation at different frequencies and at different quadriceps muscle length, height (H) of drop jump (DJ), muscle soreness and creatine kinase (CK) activity in the blood) after 100 DJs from 0.75 m height performed with maximal intensity with an interval of 20 s between the jumps (stretch-shortening exercise, SSE). All ISMDs remained even 72 h after SSE (P < 0.01–0.001). The muscle experienced greater decrease (P < 0.01) in torque evoked by electrostimulation (at low stimulation frequencies and at short muscle length in particular) after SSE than neuromuscular performance (MVCT and H of DJ) which demonstrated secondary decrease (P < 0.01) in neuromuscular performance during the first 48 h after SSE. Within 24–72 h after the SSE the subjects felt an acute muscle pain (5–7 points approximately) and the CK activity in the blood was significantly increased up to 1200 IU/L (P < 0.001). A significant correlation between decrease in MVCT and H of DJ 24–48 h after SSE on the one hand and muscle soreness registered within 24–48 h after SSE on the other was observed, whereas correlation between the other indirect symptoms of skeletal muscle damage was not significant.     

Endothelial nitric oxide synthase gene haplotypes and circulating nitric oxide levels significantly associate with risk of essential hypertension

Nitric oxide (NO), a potent vasodilator, plays a pivotal role in blood pressure regulation. Endothelial NO synthase gene (NOS3) polymorphisms influence NO levels. Here, we investigated the role of the – 922A/G, – 786T/C, 4b/4a, and 894G/T polymorphisms of the NOS3 and NO_x levels in 800 consecutive unrelated subjects comprising 455 patients of essential hypertension and 345 controls. The polymorphisms were investigated independently and as haplotypes. Plasma NO_x levels (nitrate and nitrite) were estimated by the Griess method. Genotype frequencies for the –786T/C, 4b/4a, and 894G/T polymorphisms differed significantly (P < 0.001) between patients and controls and were associated with an increased risk of hypertension (OR = 2.0, OR = 3.8, OR = 1.6, respectively). The 4-locus haplotypes ATaG (H1), ATaT (H2), and GCaG (H3) were significantly associated with essential hypertension and served as susceptible haplotypes (P ≤ 0.0001). On the other hand, haplotypes ATbG (H4) and GTbG (H5) were negatively associated with hypertension and served as protective haplotypes (P < 0.0001). NO_x levels were significantly lower in patients than controls (P < 0.0001). The individual polymorphisms showed marginal association with NO_x level; however, the susceptible haplotype H2 associated significantly with lower NO_x levels in patients (P < 0.001) and conversely the haplotype H4 with higher NO_x levels in controls (P < 0.001). In conclusion, the 4b/4a and likely – 786T/C polymorphisms were identified as the determinants modifying the risk of hypertension. This study identifies the NOS3 variants and haplotypes as genetic risk factors and as useful markers of increased susceptibility to the risk of essential hypertension.     

A note on the effects of co-mingling piglet litters on pre-weaning growth, injuries and responses to behavioural tests

The purpose of this study was to determine how co-mingling litters affected piglets’ pre-weaning growth, ear injuries, suckling behaviour and responses to behavioural tests used to measure coping abilities. Thirty sows and their respective litters were housed in standard farrowing crates until day 13 after birth. On day13, the partition between two neighbouring pens was removed for 20 litters allowing piglets to interact (forming 10 co-mingled litters). The remaining 10 control litters were kept in standard farrowing crates throughout the experiment. Three focal piglets from each litter were used for data collection. Focal piglets were weighed and ear injuries recorded on days 2, 4, 9, 12, 15 and 18 after birth. There were no differences in piglets’ weight gain before or after co-mingling. Ear injuries were more abundant in co-mingled litters on day 15 (P < 0.05) but these differences disappeared by day 18. Suckling behaviour was recorded on days 5, 8, 10, 14, 16 and 18 after birth. There were no differences in teat fidelity, suckling frequency and mother fidelity between treatments. Three behavioural tests, social challenge, isolation, and backtest, were performed before and after co-mingling. There were no treatment effects on piglets’ response to the isolation test and backtest. Co-mingled piglets showed longer latency for the first aggressive interaction (P < 0.05), spent more time in proximity to one another (P < 0.05) and performed less single bites (P < 0.05) than control piglets during the social challenge. In addition, the duration and frequency of aggressive interactions (P < 0.05) were lower in co-mingled piglets than control piglets. Co-mingling did not affect the frequency of single head thrusts or oral–nasal contact, but did tend to increase the frequency of escape attempts (P < 0.10). Our results suggest that co-mingling litters during lactation affects piglets’ social behaviour, by primarily decreasing aggressive interactions during social challenges.     

PAH-DNA adducts in environmentally exposed population in relation to metabolic and DNA repair gene polymorphisms

Epidemiologic studies indicate that prolonged exposure to particulate air pollution may be associated with increased risk of cardiovascular diseases and cancer in general population. These effects may be attributable to polycyclic aromatic hydrocarbons (PAHs) adsorbed to respirable air particles. It is expected that metabolic and DNA repair gene polymorphisms may modulate individual susceptibility to PAH exposure. This study investigates relationships between exposure to PAHs, polymorphisms of these genes and DNA adducts in group of occupationally exposed policemen (EXP, N = 53, males, aged 22–50 years) working outdoors in the downtown area of Prague and in matched “unexposed” controls (CON, N = 52). Personal exposure to eight carcinogenic PAHs (c-PAHs) was evaluated by personal samplers during working shift prior to collection of biological samples. Bulky-aromatic DNA adducts were analyzed in lymphocytes by ³²P-postlabeling assay. Polymorphisms of metabolizing (GSTM1, GSTP1, GSTT1, EPHX1, CYP1A1-MspI) and DNA repair (XRCC1, XPD) genes were determined by PCR-based RFLP assays. As potential modifiers and/or cofounders, urinary cotinine levels were analyzed by radioimmunoassay, plasma levels of vitamins A, C, E and folates by HPLC, cholesterol and triglycerides using commercial kits. During the sampling period ambient particulate air pollution was as follows: PM10 32–55 μg/m³, PM2.5 27–38 μg/m³, c-PAHs 18–22 ng/m³; personal exposure to c-PAHs: 9.7 ng/m³ versus 5.8 ng/m³ (P < 0.01) for EXP and CON groups, respectively. The total DNA adduct levels did not significantly differ between EXP and CON groups (0.92 ± 0.28 adducts/10⁸ nucleotides versus 0.82 ± 0.23 adducts/10⁸ nucleotides, P = 0.065), whereas the level of the B[a]P-“like” adduct was significantly higher in exposed group (0.122 ± 0.036 adducts/10⁸ nucleotides versus 0.099 ± 0.035 adducts/10⁸ nucleotides, P = 0.003). A significant difference in both the total (P < 0.05) and the B[a]P-“like” DNA adducts (P < 0.01) between smokers and nonsmokers within both groups was observed. A significant positive association between DNA adduct and cotinine levels (r = 0.368, P < 0.001) and negative association between DNA adduct and vitamin C levels (r = −0.290, P = 0.004) was found. The results of multivariate regression analysis showed smoking, vitamin C, polymorphisms of XPD repair gene in exon 23 and GSTM1 gene as significant predictors for total DNA adduct levels. Exposure to ambient air pollution, smoking, and polymorphisms of XPD repair gene in exon 6 were significant predictors for B[a]P-“like” DNA adduct. To sum up, this study suggests that polymorphisms of DNA repair genes involved in nucleotide excision repair may modify aromatic DNA adduct levels and may be useful biomarkers to identify individuals susceptible to DNA damage resulting from c-PAHs exposure.