Jean-Paul Laforest

The importance of porcine sperm parameters on fertility in vivo

It would be desirable to use semen parameters to predict the in vivo fertilizing capacity of a particular ejaculate. In animal production, an ejaculate is divided into multiple doses for artificial insemination (AI); therefore, it would be economically beneficial to know the functional quality (i.e., fertility) of the semen before it is inseminated. To identify a predictive assay of the fertilizing capacity of a porcine ejaculate, we performed 4 rapid assays of sperm quality (motility, viability, physiological status as assessed by chlortetracycline fluorescence, and ATP content) on samples from 9 ejaculates, before and after a thermal stress test (42.5 degrees C, 45 min). These parameters were subsequently correlated with in vivo fertility resulting from AI with 2 sperm doses, 3 x 10(9) or 0.3 x 10(9) motile cells in 70 mL (optimal or suboptimal sperm number per insemination, respectively) from these same ejaculates. No parameter was correlated to the fertility rates obtained after inseminating with the optimal semen doses, either before or after the thermal stress test (P > 0.05). However, with respect to the animals inseminated with the suboptimal semen dose, sperm motility (the percentage of motile spermatozoa as assessed visually by microscopy) prior to thermal stress was well-correlated to fertility rates (r = 0.783, P = 0.01). The percentage of spermatozoa displaying the chlortetracycline Pattern AR (acrosome reaction) was also statistically related to fertility (r = 0.05, P = 0.04), but the biological importance of this relationship is questionable given the small variation among ejaculates (range: 0 to 2%). No other sperm parameter was significantly related to fertility rates in this group (P > 0.05). These data, therefore, indicate that sperm motility is a useful indicator of sperm fertilizing capacity in vivo. Moreover, to identify a predictor of semen fertility it is critical that the number of spermatozoa used during insemination is sufficiently low to detect differences in sperm fertilizing efficiency.

Effect of dietary n-3 fatty acids (fish oils) on boar reproduction and semen quality

The aim of this study was to evaluate the effects of dietary supplementation with different fish oils (rich in PUFA) vs. hydrogenated animal fat (SFA) on semen production and quality, fatty acid composition, and preservation properties in boars under controlled and commercial conditions. In Exp. 1 (in a research station), 44 boars, allocated to 4 dietary treatments, received daily 2.5 kg of basal diet with a supplement of 1) 62 g of hydrogenated animal fat (AF, n = 12); 2) 60 g of menhaden oil containing 18% docosahexanoic acid (DHA) and 15% eicosapentanoic acid (EPA; MO, n = 11); 3) 60 g of tuna oil containing 33% DHA and 6.5% EPA (TO, n = 11); and 4) 60 g of menhaden oil and 2 mg/kg of biotin (MO+B, n = 10). Biotin is a critical factor in the elongation of PUFA. Semen was collected according to 3 successive phases: phase 1 (twice per week for 4 wk); phase 2 (daily collection for 2 wk); and phase 3 (twice per week for 10 wk). Experiment 2 was conducted in commercial conditions; 222 boars were randomly allocated to AF, MO, and TO treatments. Semen was collected twice weekly over a 6-mo period. All diets were balanced to be iso-energetic and provided an equivalent of 989 mg of vitamin E per day. Classical measurements of sperm quantity and quality were done for both experiments. Experiment 1 showed, after 28 wk of supplementation, a massive transfer of n-3 PUFA into sperm from boars fed fish oil diets (MO and TO). No differences were observed among dietary treatments for libido (P > 0.30), sperm production (P > 0.20), or percentage of motile cell (P > 0.20). Unexpectedly, MO+B diet reduced the percentage of normal sperm compared with the other treatments (P < 0.03). In conclusion, although it modified the fatty acid composition of sperm, supplementation of boars with dietary fish oils, rich in long chain n-3 fatty acids, did not influence semen production or quality postejaculation.

Dietary omega-3 fatty acids (fish oils) have limited effects on boar semen stored at 17 °C or cryopreserved

To evaluate the influence of dietary supplementation of omega-3 polyunsaturated fatty acids (n-3 PUFA) on storage of boar semen, three experiments were conducted: two involved long-term, fresh semen storage (Exp. 1 and Exp. 2), whereas the other involved cryopreservation (Exp. 3). Boars were allocated randomly to three dietary treatments (for 6-7 mo). In addition to a daily allowance of 2.5 kg of a basal diet, they received: 1) 62 g of hydrogenated animal fat (AF); 2) 60 g of menhaden oil (MO), containing 18% docosahexanoic acid (DHA) and 15% eicosapentanoic acid (EPA); or 3) 60 g of tuna oil (TO), containing 33% DHA and 6.5% EPA. In Experiment 1 (n = 26) and Experiment 2 (n = 18), semen was cooled and stored in vitro for several days at 17 °C before assessment, whereas in Experiment 3 (n = 18), viability, motility, acrosomal integrity, susceptibility to peroxidation (LPO), and DNA fragmentation were determined in fresh and frozen-thawed sperm. In Experiment 1, sperm from boars fed TO had better resistance to fresh storage; even after 7 or 9 d of storage at 17 °C, there were more (P = 0.03) motile sperm in boars fed TO (>60%) than in those fed AF or MO. In Experiment 2, fish oil supplementation did not influence any aspect of sperm quality during semen storage (P > 0.10). In Experiment 3, cryopreservation decreased the proportion of motile and viable frozen-thawed sperm as well as acrosomal integrity and increased DNA fragmentation and LPO (P < 0.01) relative to fresh semen, although sperm quality was unaffected by treatments (P > 0.09). In conclusion, although adding fish oil to the diet failed to significantly improve the quality of cryopreserved boar sperm, inconsistent responses of long-term storage of cooled sperm to dietary n-3 PUFA supplementation warrant further investigation.

Effects of Breed, Parity, and Folic Acid Supplement on the Expression of Leptin and Its Receptors' Genes in Embryonic and Endometrial Tissues from Pigs at Day 25 of Gestation

Abstract Recent evidence has pointed toward a possible role of leptin (Lep) and its receptor (Lepr) in early gestation materno-fetal cross-talk. However, in gestating sows, exhaustive characterization of leptin mRNA expression in backfat and leptin-receptor mRNA expression in endometrial and embryonic tissues is still pending. The objectives of this study were to characterize the Lep, Lepr, and long Lepr-L isoform mRNA expression according to the breed and parity of gestating sows or to specific folic acid (B9) + glycine dietary treatments. To this end, nulliparous (GT) and multiparous occidental Yorkshire-Landrace (YL) sows as well as multiparous Chinese Meishan-Landrace (ML) sows were used. These sows were randomly assigned to two different dietary treatments: 0 or 15 ppm of B9 + 0.6% glycine, given from the estrous preceding mating until slaughter on Day 25 of gestation. Jugular blood samples were collected at mating and on Day 25 of gestation and assayed for circulating leptin concentrations. Expression levels of Lep in backfat and of Lepr and Lepr-L in endometrial and embryonic tissues were performed using semiquantitative reverse transcription-polymerase chain reaction. Results demonstrated that on Day 25 of pregnancy, the ML sows showed higher concentrations of circulating leptin along with higher backfat thickness and higher expression of Lep in backfat tissue. Moreover, in embryonic tissues, the mRNA expression levels of Lepr and Lepr-L genes were higher in ML than in YL sows. Parity effects were observed for mRNA expression of Lepr in both endometrial and embryonic tissues, whereas mRNA levels were higher in YL than in GT sows. In addition, embryonic Lepr-L mRNA levels were higher in GT than in YL sows, and B9 + glycine dietary supplement decreased the mRNA expression levels of Lep in backfat and of Lepr in embryonic tissues. These decreases were independent of breed or parity of the sows. The effect of B9 + glycine on Lepr-L mRNA expression levels was only seen in YL sows, whereas the treatment lowered Lepr-L expression levels in both endometrial and embryonic tissues. These results indicate that leptin and its receptor may play a role during early stages of development of the pig embryo-fetus, and that these roles could be modulated according to the breed and parity of the sows. Moreover, the effects of B9 + glycine on expression levels of embryonic and endometrial Lepr-L mRNA in YL sows may explain the previously reported effects of B9 on embryo survival rate and litter size observed in occidental multiparous sows.

Fish oil diets alter the phospholipid balance, fatty acid composition, and steroid hormone concentrations in testes of adult pigs

The objective was to determine the effect of long-term dietary supplementation of two types of fish oil on lipid composition and steroidogenesis in adult pig testis. Twenty-four Duroc boars, aged 204.5 ± 9.4 d (body weight 128.1 ± 16.7 kg) received daily 2.5 kg of an iso-caloric basal diet supplemented with: 1) 62 g of hydrogenated animal fat (AF); 2) 60 g of menhaden oil (MO) containing 16% of eicosapentaenoic acid (EPA) and 18% of docosahexaenoic acid (DHA); or 3) 60 g of tuna oil (TO) containing 7% of EPA and 33% of DHA. After these diets were consumed for 7 mo, testicular hormones, phospholipid content, and fatty acid composition of individual phospholipids in testis were determined. Body and reproductive organ weights were not significantly affected by dietary treatments. Testicular tissue from boars fed a TO diet, followed by those receiving MO and AF diets, had the lowest level of phosphatidylethanolamine (TO < MO < AF; P < 0.01) but the highest sphingomyelin (TO > MO > AF; P < 0.01). For each phospholipid, boars fed either the MO or TO diet had increased total omega-3 fatty acids, particularly DHA (P < 0.01), by reciprocal replacement of total omega-6 fatty acids (20:4n-6, 22:5n-6). The MO diet increased EPA more than the other diets. Testicular concentrations of testosterone and estradiol were lower in boars fed a TO diet than a MO diet (P < 0.02). In conclusion, long-term dietary supplementation of fish oil, regardless of the EPA/DHA ratio, modified the fatty acid compositions in testis and affected steroid production of healthy adult boars, which may represent a promising models for future studies on fertility.

Effects of folic acid and vitamin B12 supplements on folate and homocysteine metabolism in pigs during early pregnancy

The present experiment aimed to determine the effects of supplements of folic acid (FA) alone or in combination with vitamin B12 on folate and homocysteine metabolism in gestating nulliparous Yorkshire-Landrace (YL) and multiparous Landrace (LD) occidental sows and multiparous Chinese Meishan-Landrace (ML) sows. LD sows were randomly assigned to two treatments: 0 or 15 mg FA/kg diet while YL and ML sows were assigned to three treatments: 0 mg FA/kg diet, 15 mg FA/kg or 15 mg vitamin B12/kg diet. Supplements were given from the oestrus preceding insemination up to slaughter on day 15 of gestation. At slaughter, a uterine flush was collected to determine uterine contents of homocysteine, methionine, tetrahydrofolate (THF), 5-methyl-THF, pyridoxal 5-phosphate (P5P) and vitamin B12. Blood samples were taken at first oestrus, at insemination and on days 5, 10 and 15 of gestation to determine plasma concentrations of homocysteine, methionine, THF, 5-methyl-THF, P5P, vitamin B12 and relative total folate-binding capacity. In occidental sows (YL and LD), the FA supplement tended to decrease uterine flush content of homocysteine (P=0.06) and concentrations of plasma homocysteine (P=0.09). Nulliparous YL sows had lower concentrations of plasma homocysteine, methionine, THF and 5-methyl-THF (P<0.05) than multiparous LD sows. Multiparous ML and LD sows had similar concentrations of plasma THF, 5-methyl-THF, methionine and vitamin B12, but ML sows had lower concentrations of plasma homocysteine (P<0.05). The vitamin B12 supplement increased concentrations of plasma vitamin B12 (P<0.05) both in multiparous ML and nulliparous YL sows, but had no effect on the composition of either uterine flush or plasma. The present results showed also that sows had a low vitamin B12 status (<200 pg/ml) and high circulating homocysteine levels (>15 microm) during the first 15 d of gestation. Furthermore, the vitamin B12 content in uterine secretions represented between 180 and 300 % of the total content in plasma. The low plasma concentrations of homocysteine in multiparous ML sows suggest a more efficient remethylation pathway which may not be dependent upon dietary supply of FA or vitamin B12. In nulliparous YL sows, low concentrations of both homocysteine and methionine suggest that the methionine requirement for protein deposition might have reduced the amount of methionine available for the methylation pathway. The results of the present experiment suggest that the reduction of uterine homocysteine may be an important aspect of the role of FA supplement on the uterine environment in occidental sows. The presence of high levels of vitamin B12 in uterine secretions merits further investigation in relation to embryonic development.

Reproductive performance and uterine prostaglandin secretion in gilts conditioned with dead semen and receiving dietary supplements of folic acid

The effects of dietary supplements of folic acid at 0 (F−) or 15 ppm (F+) and of conditioning the uterus (C+) or not (C−) with an intrauterine infusion of dead semen on embryonic survival and prostaglandin E2 (PGE2) concentrations in the allantoic fluid on day 30 of gestation were studied. Sixty-four F1 crossbred gilts (Yorkshire × Landrace) were used in a 2 × 2 factorial design. The initiation of folate supplementation and the intrauterine infusion of dead semen were done at the estrus preceding the fertile mating. Folate status in serum was lower in F+ gilts than in F− gilts during the estrus cycle (folic acid × period interaction, P ≤ 0.007). Conditioning increased ovulation rate in F− gilts while it tended to decrease it in F+ gilts (interaction folic acid × conditioning, P ≤ 0.03). A similar interaction (P ≤ 0.02) was observed on the number of presumably live embryos at 30 d of gestation. Mean PGE2 concentration in the allantoic fluid was not affected by treatments (P ≥ 0.44), but the frequency of so...

Effects of dietary vitamin supplementation and semen collection frequency on reproductive performance and semen quality in boars.

The present study was undertaken to assess the relevance of increasing the daily provision of dietary vitamins on vitamin metabolic status and semen characteristics of boars under controlled and commercial conditions as well as to evaluate the efficiency of this vitamin supplement to allow boars to cope with intensive semen collection frequency. In the first experiment, 39 boars were allocated to 2 dietary treatments, a basal diet (control) and the basal diet supplemented with extra fat- and water-soluble vitamins (Vit). Within each treatment, boars were submitted to 2 regimens of semen collection frequency: 3 times per 2 wk (3/2) and 3 times per week (3/1) over a 12-wk period. Afterwards, all boars were intensively collected (daily) for 2 wk. A resting period of 4 wk followed, and all boars were collected 2 times per week. Thereafter, collection frequencies were reversed, and the same procedure was followed until the end of the intensive collection period. A second experiment was conducted in commercial conditions at a commercial stud, and 252 boars were randomly allocated to the control and Vit dietary treatments. All boars were collected 2 times per week over a 6-mo period. Classical measurements of ejaculate and sperm quality were assessed, and blood samples were collected throughout both experiments to quantify vitamin concentrations. In the first experiment, vitamin concentrations in blood and seminal plasma increased in Vit boars (P < 0.05); however, vitamin concentrations were not affected by collection frequency (P > 0.14). The Vit supplement did not affect sperm production or sperm quality (P > 0.28), although semen volume increased during the 12-wk periods for Vit boars (P < 0.05). The 3/1 boars produced fewer doses per ejaculate than 3/2 boars (P < 0.01); however, the cumulative sperm production for the 12-wk periods increased by 19% in 3/1 boars compared with 3/2 boars. In the second experiment, blood plasma concentrations of vitamin B(9) were greater (P < 0.01) in Vit than control boars. The vitamin supplement did not increase sperm production of boars (P > 0.61). In conclusion, dietary supplements of fat- and water-soluble vitamins increase the amount of vitamins available for the animal, and the collection frequencies had no effect on vitamin status. Moreover, in spite of an effect on the ejaculate volume, the dietary supplement of extra vitamins had no effect on sperm production or quality.

The Way Humans Behave Modulates the Emotional State of Piglets.

The emotional state can influence decision-making under ambiguity. Cognitive bias tests (CBT) proved to be a promising indicator of the affective valence of animals in a context of farm animal welfare. Although it is well-known that humans can influence the intensity of fear and reactions of animals, research on cognitive bias often focusses on housing and management conditions and neglects the role of humans on emotional states of animals. The present study aimed at investigating whether humans can modulate the emotional state of weaned piglets. Fifty-four piglets received a chronic experience with humans: gentle (GEN), rough (ROU) or minimal contact (MIN). Simultaneously, they were individually trained on a go/no-go task to discriminate a positive auditory cue, associated with food reward in a trough, from a negative one, associated with punishments (e.g. water spray). Independently of the treatment (P = 0.82), 59% of piglets completed the training. Successfully trained piglets were then subjected to CBT, including ambiguous cues in presence or absence of a human observer. As hypothesized, GEN piglets showed a positive judgement bias, as shown by their higher percentage of go responses following an ambiguous cue compared to ROU (P = 0.03) and MIN (P = 0.02) piglets, whereas ROU and MIN piglets did not differ (P > 0.10). The presence of an observer during CBT did not modulate the percentage of go responses following an ambiguous cue (P > 0.10). However, regardless of the treatment, piglets spent less time in contact with the trough following positive cues during CBT in which the observer was present than absent (P < 0.0001). This study originally demonstrates that the nature of a chronic experience with humans can induce a judgement bias indicating that the emotional state of farm animals such as piglets can be affected by the way humans interact with them.

Induction of estrus with intramuscular injections of GnRH or PMSG in lactating goats (Capra hircus) primed with a progestagen during seasonal anestrus

In Experiment 1, goats in seasonal anestrus (n=154) were treated with sponges impregnated with 1 of 2 types of progestagen (MAP or FGA) followed by PMSG (400 IU im) 48 h before sponge removal. The type of progestagen used had no effect on kidding, abortion, pseudogestation, multiple births, stillbirths, number of live births per doe or gestation length. In Experiment 2, lactating goats (n=24) in seasonal anestrus were treated with progestagen sponges (MAP). At sponge removal they received one of the following treatments: 1 injection of PMSG (400 IU im), 1 injection of GnRH (125 mug im; GnRH-1), or 2 injections of GnRH (125 mug/injection im; GnRH-2) at a 48 h interval. Serum samples were taken at 6-h intervals for 96 h, starting 12 h after sponge removal. Heterologous radioimmunoassays were validated for the measurements of goat FSH, LH, E(2) and P(4). The onset of estrus (P=0.004), mean doe receptivity (P=0.0006), maximum preovulatory E(2) concentrations (P=0.0001) and LH peak concentrations (P=0.08) occurred significantly later for GnRH-1 and GnRH-2 than for PMSG treatment. The PMSG treatment induced a preovulatory LH peak in a greater number of goats (P=0.05) and gave a higher gestation rate than GnRH-1 and GnRH-2 treatments (57 vs 0 vs 12%; P=0.03). It is likely that the GnRH treatments administered did not reactivate the hypothalamo-pituitary-gonadal axis. Thus, intramuscular injections of GnRH in lactating goats primed with a progestagen were not as effective in regulating reproductive performance during seasonal anestrus as were injections of PMSG.