Ovsynch Plus protocol improves ovarian response in anovular Murrah buffaloes in low‐breeding season

The objective of the study was to evaluate the efficacy of ovarian response and pregnancy rate in anovular buffaloes following Ovsynch and Ovsynch Plus protocols. Buffaloes (n = 55) were divided into two groups: Ovsynch group (n = 26): GnRH (10 μg, GnRH1) on Day 0, PGF₂α (25 mg) on Day 7, GnRH (10 μg, GnRH2) on Day 9; Ovsynch Plus group (n = 29): 500 IU equine chorionic gonadotropin (eCG) 72 hr (day ?3) prior to Ovsynch protocol, followed by fixed timed artificial insemination (FTAI) 6 and 24 hr after GnRH2 injection in bot groups. Transrectal ultrasonography was performed daily, that is, from day 0 and ?3 in Ovsynch and Ovsynch Plus group, respectively for ovarian response and pregnancy diagnosis at day 30 post‐insemination. In Ovsynch Plus group, administration of eCG prior to GnRH1 increased (p < .001) the diameter (mm) of dominant follicle (DF) from 10.15 ± 0.26 to 12.23 ± 0.34 within 72 hr of treatment resulting higher ovulatory response to GnRH1. Ovulation after GnRH1 was higher (p < .01) in Ovsynch Plus group (96.6%) than Ovsynch group (61.5%). However, ovulation rate to GnRH2 was similar (p > .05) between groups (Ovsynch group: 76.9% vs. Ovsynch Plus group: 70.0%). Mean DF diameter (mm) that ovulated to both GnRHs was higher (p < .01) than non‐ovulated counterparts in both groups (Ovsynch group: 10.80 ± 0.27 vs. 8.47 ± 0.53; Ovsynch Plus group: 11.99 ± 0.24 vs. 9.5 ± 0.63). Pregnancy was established in buffaloes which responded to both GnRHs, irrespective of groups, being higher (p = .52) in Ovsynch Plus group (34.5%) than Ovsynch group (23.1%), though non‐significant. In summary, this study showed that eCG inclusion prior to Ovsynch regimen improves ovulatory response in anovular buffaloes during low‐breeding season.     

Cryopreservation of Nili‐Ravi buffalo (Bubalus bubalis) semen in AndroMed® extender; in vitro and in vivo evaluation

The study was designed to evaluate AndroMed^® for the freezability and fertility of Nili‐Ravi buffalo semen. Semen was collected from four adult Nili‐Ravi buffalo (Bubalus bubalis) bulls for 3 weeks (replicate). Semen ejaculates from each buffalo bull were divided into three aliquots. One aliquot was used for evaluation of motility, plasma membrane integrity, livability, viability, DNA integrity and normal apical ridge. Remaining two aliquots were diluted (37°C; 50 × 10⁶ spermatozoa/ml) in tris‐citric egg yolk or AndroMed^® extender and cryopreserved in 0.5 ml French straws. After thawing, per cent post‐thaw motility (47.9 ± 0.8, 49.2 ± 1.7), plasma membrane integrity (44.4 ± 1.2, 46.8 ± 1.8) and normal apical ridge (81.4 ± 0.3, 83.2 ± 0.3) were recorded similar (p > .05) in tris‐citric egg yolk and AndroMed^® extender. Higher (p < .05) percentage of sperm livability (70.5 ± 1.4 and 64.4 ± 1.0), viability (67.5 ± 1.5 and 61.5 ± 0.6) and DNA integrity (97.0 ± 0.3 and 93.4 ± 0.21) were recorded in AndroMed^® compared to tris‐citric egg yolk post‐thaw. Values for all the aforementioned spermatozoal quality parameters were observed lower (p < .05) in frozen‐thawed compared to fresh semen irrespective of the experimental extenders. Fertility rates of buffalo semen did not differ (p > .05) either cryopreserved in tris‐citric egg yolk or AndroMed^® extender (45.5% vs. 49%). It is concluded that AndroMed^® is capable in protecting the buffalo bull sperm during freeze‐thawing process and can be adopted safely for routine use replacing the tris‐citric egg yolk extender in artificial insemination programme.     

Effects of two freezing methods and two cryopreservation media on post‐thaw quality of stallion spermatozoa

Glycerol‐based extenders are widely utilized for freezing equine semen, but media combining methylformamide may better preserve sperm motility and mitochondrial function. Semen is cryopreserved utilizing either a Styrofoam box filled with liquid nitrogen or an automatic freezer. The objective of this experiment was to compare the post‐thaw characteristics of the same ejaculates cryopreserved in a Styrofoam box or in an automatic freezer, utilizing a glycerol‐based extender (Gent) and an extender that combines methylformamide and glycerol (BotuCrio^®). For that, one ejaculate from 30 stallions collected in two different centres was used. For data analysis, a mixed linear model with laboratory, medium and freezing method and respective interactions as fixed effects was used. Stallion was taken into account as a random effect. There was no influence (p > .05) of laboratory, while stallion effect was marked. Semen frozen in BotuCrio^® in the automatic freezer had higher (p < .001) VCL than semen cryopreserved in Gent using the Styrofoam box. VCL was also higher (p = .068) for semen frozen in BotuCrio^® in the Styrofoam box than for semen cryopreserved in Gent using the same method. The difference between percentage of sperm with intact plasma membrane frozen in Gent using the Styrofoam box (44.43% ± 2.44%) compared to spermatozoa cryopreserved in BotuCrio^® using the same method (40.78% ± 2.42%) approached significance (p = .0507). The percentage of sperm with intact acrosome membrane was higher (p < .05) in semen frozen in BotuCrio^® (79.08% ± 1.79%) than semen frozen in Gent (75.15% ± 1.80%). A higher (p = .0125) percentage (32.24% ± 2.18%) of semen extended in Gent and cryopreserved in the Styrofoam box had high mitochondrial membrane potential than semen frozen in BotuCrio^® using the same method (26.02% ± 2.15%). Fertility studies are warranted to assess whether differences found have any effect on the fertility of inseminated mares.     

Early luteal development in Santa Inês ewes superovulated with reduced doses of porcine follicle‐stimulating hormone

The aim was to compare the early luteal development in ewes superovulated with different doses of pFSH. Twenty‐nine Santa Inês ewes received a progesterone device (CIDR®) for 8 days. Gonadotrophic treatment started on Day 6: G200 (control, n = 9, 200 mg); G133 (n = 10, 133 mg); and G100 (n = 10, 100 mg of pFSH). On Day 6, all females received eCG (300 IU). B‐mode and spectral Doppler ultrasonography were performed daily during the early luteal phase (Days 11–15) to monitor the development of corpora lutea (CLs; dimensions) and ovarian arteries indices. CLs were also classified as normal or prematurely regressed (PRCL) on Day 15 by videolaparoscopy. Ewes from G100 and G133 showed gradual increase in luteal diameter during the early luteal phase (p < 0.001), whereas G200 animals presented increase from Day 11 to Day 13, and then decrease on Days 14 and 15 (p < 0.001). The G200 females showed greater percentage of PRCL (45.20%) than those of the other groups (p < 0.001). The normal CLs number was greater in G100 than in G133 (p = 0.04), while the PRCL number was greater in G200 than in the other groups (p = 0.03). Resistive index (RI) was greater in G200 than in G100 (p = 0.02). RI was lower in Day 12 than Day 15 (p = 0.02). Pulsatility index (PI) was greater on Days 14 and 15 (p < 0.01). In conclusion, the lowest dose of pFSH (100 mg) can be considered sufficient for an efficient superovulatory response in sheep, producing better CLs development dynamic in early luteal phase and ovarian blood perfusion and smaller number of PRCL than the traditional (200 mg) pFSH dose.     

Pharmacokinetics of intravenous gentamicin in healthy young‐adult compared to aged alpacas

The study objective was to evaluate the effects of age on aminoglycoside pharmacokinetics in eight young‐adult (<4 years) and eight aged (≥14 years) healthy alpacas, receiving a single 6.6 mg/kg intravenous gentamicin injection. Heparinized plasma samples were obtained at designated time points following drug administration and frozen at ?80°C until assayed by a validated immunoassay (QMS ^®). Compartmental and noncompartmental analyses of gentamicin plasma concentrations versus time were performed using WinNonlin (v6.4) software. Baseline physical and hematological parameters were not significantly different between young and old animals with the exception of sex. Data were best fitted to a two‐compartment pharmacokinetic model. The peak drug concentration at 30 min after dosing (23.8 ± 2.1 vs. 26.1 ± 2 μg/ml, p = .043 ) and area under the curve (70.4 ± 10.5 vs. 90.4 ± 17.6 μg hr/ml, p = .015 ) were significantly lower in young‐adult compared to aged alpacas. Accordingly, young alpacas had a significantly greater systemic clearance than older animals (95.5 ± 14.4 and 75.6 ± 16.1 ml hr^?1 kg^?1; p = .018 ), respectively). In conclusion, a single 6.6 mg/kg intravenous gentamicin injection achieves target blood concentrations of >10 times the MIC of gentamicin‐susceptible pathogens with MIC levels ≤2 μg/ml, in both young‐adult and geriatric alpacas. However, the observed reduction in gentamicin clearance in aged alpacas may increase their risk for gentamicin‐related adverse drug reactions.     

Assessment of the usage of biodegradable polymeric matrix in vaginal devices to sustain progesterone release in cows

The usage of timed artificial insemination (TAI) at a low cost leading to better reproductive rates has been the aim of several research groups in the field. Usually during TAI protocols, sustained progesterone (P₄) release devices are employed. Most devices are constituted of a nylon skeleton covered with a silicon layer with P₄. A device based on biopolymers was developed in order to reduce costs and decrease its environmental impact. In this study, we compared the kinetics of sustained progesterone release among devices manufactured with a polymeric blend made of polyhydroxybutyrate‐valerate (PHBV) and poly‐ε‐caprolactone (PCL) (DISP) which were compared with DIB® (Internal Bovine Device) used as the control. In the in vitro and in vivo progesterone release tests, two types of biopolymer‐based devices with a superficial area of 147 cm² were used: DISP8 (46% PHBV, 46% PCL and 8% P₄; n  = 4), DISP10 (45% PHBV, 45% PCL, 10% P₄; n  = 4) and DIB® (1 g P₄, 120 cm² area; n  = 3). The in vitro tests were carried out according to USP XXIII specifications and were performed in a dissolutor sink using an alcohol/water mixture (60/40 v/v) as a release media and samples were collected at 2 min, 2, 4, 8, 12, 24, 48, 60, 72, 84 and 96 h. P₄ concentrations were measured through spectrophotometry in a 244 nm long wave. Three to 3 comparisons of angular coefficients of the straight lines obtained by regression analysis of accumulated P₄ concentrations as a function of square root of time were carried out. Furthermore, the diffusion coefficient values of P₄ were also determined for DISP8 and DISP10. The results showed that the concentrations of P₄ were higher in the DISP10 (774.63 ± 45.26 μg/cm²/t^1/2) compared to DISP8 (566.17 ± 3.68 μg/cm²/t^1/2) (P  < 0.05). However, both DISP10 and DISP8 P₄ concentrations did not differ from DIB® (677.39 ± 16.13 μg/cm²/t^1/2). For the analysis of released quantities per day of the in vitro test, four periods were considered: 0–24, 24–48, 48–72 and 72–96 h. In the first 24 h, DISP8 released significantly less P₄ than DISP10 or DIB®, which did not differ among them. Between 24 and 48 h, DISP10 released significantly more P₄ than DIB®. DISP8 released an intermediate P₄ amount and did not differ significantly from DIB® or DISP10. Between 48 and 72 h, P₄ quantity released by DISP10 was significant higher (P  < 0.01) than that of DIB® and DISP8, which did not differ among themselves. Between 72 and 96 h, DISP10 released significantly more P₄ than DIB®, and DISP8 released an intermediate amount which did not differ from DIB® or DISP10 (P  < 0.01). There was interaction between treatment and time (P  = 0.0024). The diffusion coefficient values were: 1.36 × 10^?8 (cm²/s) for DISP10 and 1.12 × 10^?8 (cm²/s) for DISP8. For the in vivo test, ovariectomized crossbred cows received DIB® (n  = 4) or DISP8 (n  = 8) in an alternate design with a non‐balanced sequence (cross‐over) added of measures repeated in time referring to 16 days of blood samples collection. Samples were analyzed through radioimmunoassay in solid phase using the commercial kit of DPC (Diagnostics Products Corporation). Plasma concentrations of P₄ peaked at 4 h after the placement of the device, this being the only time in which plasma P₄ concentrations differed between DIB® (11.45 ± 1.96) compared with DISP8 (9.23 ± 1.15 ng/mL) (P  = 0.027). On day 8, plasma P₄ concentrations were similar for DIB® (2.44 ± 0.09) and DISP8 (1.89 ± 0.13 ng/mL) (P  = 0.58) showing that both devices were able to keep P₄ concentrations above 1 ng/mL in the plasma of the cow during the 16 day in vivo test. In conclusion, devices manufactured with the blend of PHBV/PCL biopolymers can sustain the release of P₄ in a similar manner as silicon.     

Maturation ability after transfer of freeze‐dried germinal vesicles from porcine oocytes

The purpose of this study was to examine whether freeze‐dried germinal vesicles (GV) can be matured in vitro after being injected into enucleated fresh oocytes in pigs as an alternative method for conservation of genetic resources. Although no reduction of the size of GV (p = .094), resveratrol treatment significantly enhanced the survival rates following GV transfer (GVT) (p < .001). Supplementation with 100 or 200 mmol/L trehalose in freeze‐drying medium significantly increased the proportions of GVs with intact nuclear membrane and DNA integrity compared with the control group. Following transfer of freeze‐dried GVs into enucleated fresh oocytes, the proportion of reconstructed oocytes reached the metaphase‐II stage (2.4% ± 1.4%) was significantly lower (p < .05) than that of the in vitro matured control group (83.2% ± 2.5%), it was comparable with the GVT control group (7.4% ± 2.7%). The rates of freeze‐dried GVs with intact nuclear membrane and DNA stored at ?20°C for 5 days were significantly higher (p < .05) than those at 4°C and room temperature. The rates of intact nuclear membrane and DNA in the freeze‐dried GV stored for 15 or 30 days at ?20, 4°C and RT were not significantly different. In conclusion, matured oocytes were produced derived from freeze‐dried GVs.     

Comparison of in vitro and in vivo fertilizing potential of buffalo bull semen frozen in egg yolk‐, soya bean lecithin‐ and liposome‐based extenders

The objective of this study was to compare different extenders for post‐thaw in vitro sperm function and in vivo fertility of buffalo semen. Accordingly, sperm of 30 ejaculates extended in egg yolk (TRIS with 20% egg yolk; EY), two soya lecithin‐based (SL‐1; AndroMed^® and SL‐2; Bioxcell^®) and a liposome‐based extender (LS; OptiXcell^®) were tested. The post‐thaw semen was evaluated for computer‐assisted sperm analysis (CASA), sperm viability, membrane and acrosome integrity, DNA integrity and acrosome reaction and first service pregnancy rate (FSPR) in a fixed‐time artificial insemination programme. Total motility and VCL were the only CASA‐based parameters that exhibited significantly higher (p < .05) percentage in LS among these extenders. Post‐thaw percentage of acrosome integrity (55.9 ± 1.4, 58.1 ± 2.0, 55.8 ± 2.0, 56.6 ± 2.3) and DNA integrity (68.8 ± 2.0, 69.2 ± 2.3, 71.3 ± 2.1, 69.1 ± 2.1) did not differ (p > .05) in EY, SL‐1, SL‐2 and LS extender, respectively. However, a variable response in terms of efficacy of different extenders for sperm viability and plasma membrane integrity was observed. Assessment of inducibility of acrosome reaction showed significant differences between extenders (51.9 ± 2.1, 44.3 ± 2.4, 46.1 ± 2.3 and 58.1 ± 3.1%, respectively, for EY, SL‐1, SL‐2 and LS). Furthermore, field trials revealed significantly higher (p < .05) FSPR of LS‐extended semen as compared to that for EY, SL‐1 and SL‐2 extender (46.3%, 41.2%, 31.2% and 29.7%, respectively). It is concluded that the liposome‐based extender is more effective than egg yolk‐ and soya lecithin‐based extenders and may be used for cryopreservation of buffalo semen in the future.     

Pre‐Selection Test to Identify High Responder Donor Goats

The aim of the study was to evaluate the feasibility of pre‐selection of high or low responder does prior to the superovulatory protocols. Twenty Saanen does received 800 IU of equine chorionic gonadotropin (eCG) at the end of long‐term progestogen treatment. Fourteen days later, a second progestogen protocol associated with a multiple‐dose follicle stimulation hormone (FSH) treatment (5 IU/kg of FSH, in six decreasing doses between days 4 to 6 of the protocol) was administered. Transrectal ultrasound was used to assess the follicular status at the beginning of superovulatory treatments, at the oestrous onset and on the seventh day of the oestrous cycle for counting corpora lutea (CL). A significant lower number of CL was obtained in eCG‐treated in comparision with FSH‐treated does (p < 0.05). A quartic regression was able to explain the relationship between the number of CL in response to both treatments (r²=0.50; p < 0.05). Seventy per cent (14 of 20) of does maintained the same ovulatory response (high or low) after treatments. The Kappa (κ = 0.40; p < 0.05) and Spearman (rs = 0.39; p = 0.08) coefficients were able to show a relationship between treatments. Regarding the follicular status, there is a significant relationship between the number of small follicles (r = 0.71; r²=0.47; p < 0.01) and total follicles (r = 0.60; p < 0.01) at eCG and first FSH dose with the number of CL. Moreover, it was found a negative relationship between the presence of large follicles and the number of CL in response to eCG treatment (r = ?0.44; p < 0.05), but not from FSH (p > 0.05). In conclusion, the screening test with eCG has the potential to identify Saanen does that will better respond to the superovulatory protocol with FSH. In addition, it highlighted the importance of an ultrasound evaluation prior to the beginning of superovulatory treatments with FSH to characterize the follicular status and identify the potential donors of high ovulatory response in MOET programmes in goats.     

Ophthalmic examination findings in a colony of Screech owls (Megascops asio)

Objective To report ophthalmic findings in the Screech owl (Megascops asio). Sample population Twenty‐three, apparently healthy adult captive Screech owls in Maryland. Procedures OU of all owls underwent complete ophthalmic examination. One randomly assigned eye of each bird was measured by phenol red thread tear test (PRT), and the other eye by Schirmer tear test (STT). TonoVet^® rebound tonometry and TonoPen‐XL^® applanation tonometry were performed in each eye to measure IOP. Conjunctival swabs were cultured from one eye of 10 birds, corneal diameter was measured in OU of eight birds, and streak retinoscopy was performed on OU of seven birds. Ten birds were anesthetized, and A‐scan ultrasonography using a 15‐MHz probe was performed to obtain axial intraocular measurements. Results Ophthalmic abnormalities were noted in 24/46 (52%) of eyes. Median STT result was ≤ 2 mm/min, ranging ≤ 2–6 mm/min, and mean ± SD PRT was 15 ± 4.3 mm/15 s. Mean ± SD IOP were 9 ± 1.8 mmHg TonoVet^®‐P, 14 ± 2.4 mmHg TonoVet^®‐D, and 11 ± 1.9 mmHg TonoPen‐XL^®. Coagulase negative staphylococcal organisms were cultured from all conjunctival swabs. Mean ± SD corneal dimensions were 14.5 ± 0.5 mm vertically and 15.25 ± 0.5 mm horizontally. All refracted birds were within one diopter of emmetropia. Mean ± SD axial distance from the cornea to the anterior lens capsule was 4.03 ± 0.3 mm, from cornea to the posterior lens capsule was 10.8 ± 0.5 mm, and from cornea to sclera was 20.33 ± 0.6 mm. Conclusions This study reports ophthalmic examination findings in Screech owls, and provide means and ranges for various ocular measurements. This is the first report of rebound tonometry and PRT in owls.     

Retracted: Reliable collection of Caspian brown trout (Salmo trutta caspius) sperm using a catheter

The traditional stripping procedure for collecting fish semen is associated with the risk of urine contamination, which may significantly affect semen quality and quantity. The use of a catheter as an alternative method for semen collection may overcome this problem. Therefore, this study compared Caspian brown trout (Salmo trutta caspius) semen parameters (i.e. sperm density, seminal plasma osmolality, motility parameters of spermatozoa analysed using computer‐assisted sperm analysis and fertility) between the traditional stripping method and the use of a catheter. All parameter values of the semen collected with a catheter were significantly higher (p < .05; density = 7.67 ± 1.02 × 10⁹ ml^?1 and osmolality = 279.28 ± 32.84 mOsm kg^?1) than those collected with stripping method (density = 4.85 ± 0.47 × 10⁹ ml^?1 and osmolality = 216.42 ± 20.75 mOsm kg^?1). Semen collected with a catheter was characterized by higher spermatozoa motility compared with sperm collected via stripping. Similarly, the fertilization ability of sperm collected with a catheter was significantly greater (p < .05) than sperm collected with the traditional stripping method. In conclusion, collection of sperm with a catheter was shown to effectively reduce urine contamination and is therefore recommended for the collection of Caspian brown trout sperm.     

Selection of red deer spermatozoa with different cryoresistance using density gradients

The objective of sperm selection media is selecting the best spermatozoa and to remove seminal plasma and diluent for using them in assisted reproductive techniques. It is known that individuals show different cryoresistance in response to the same freezing procedure. Our hypothesis was that the efficacy of selection media could be dissimilar for samples with different sperm quality after thawing. Epididymal sperm samples from mature Iberian red deer were collected and frozen. Males were classified as with high post‐thaw sperm quality when sperm motility (SM) ≥ 70%, or as with low post‐thaw sperm quality when SM ≤ 69%. Samples were centrifuged using the following density gradients (DG): Percoll^®, Puresperm^® and Bovipure^?, and several functional sperm parameters were assessed after sperm selecting and washing. Males classified with high sperm quality had higher post‐thawing values (p > .05) for all parameters evaluated, except for linearity index, than those categorized as low sperm quality. After selection, some sperm characteristics improved (viability, apoptosis and mitochondrial activity) for both groups, showing the males with high sperm quality higher values in all sperm parameters except for kinematic traits and DNA fragmentation index (%DFI), regardless of DG. Bovipure^? yield lower values of sperm motility, viability, apoptosis and mitochondrial activity in relation to Percoll^® and Puresperm^® considering both quality groups. There was an interaction between the type of DG and sperm quality group for sperm viability (p = .040) and apoptosis (p = .003). Thus, Percoll^® selected less live and more apoptotic spermatozoa than Puresperm^® and Bovipure^? for males with low sperm quality. In conclusion, the DG are more efficient selecting spermatozoa from samples with high sperm quality, acting differently depending on initial sperm quality.     

Dynamics of sperm DNA fragmentation in raw boar semen and fertility

The aims were to evaluate sperm DNA fragmentation (SDF) in boars through the dispersion of their chromatin in raw semen samples, quantifying the extent of SDF, and to assess dynamic aspects of sperm DNA damage after incubation to obtain the rate of sperm DNA fragmentation (rSDF) under thermal conditions similar to the uterus (37°C) over a period of up to 24 hr and to correlate the reproductive outcome of the sows with the SDF of the boars at ejaculation. The study was performed on a pig‐breeding farm in southern Uruguay. Sixty‐one ejaculates from five of the most frequently used hybrid boars were evaluated. Semen was collected weekly from each of the boars, using the gloved‐hand technique and discarding the jelly‐like fraction of the ejaculate. Fresh semen was kept in a water bath at 37°C and protected from light, and was thereafter processed with Sperm‐Sus‐Halomax^® to evaluate SDF. The smears for time 0 (T0) were made on farm, and thereafter smears were made at the laboratory at 4 hr of obtaining the semen (T4), then every 2 hr (T6, T8, T10, T12) and a final fixation at 24 hr (T24). Differences in SDF were observed among exposure times for all boars (p < .05), but not between T10 and T12 (p = .7751) nor T4 and T24 (p = .9113). In none of the T24 samples, sperm heads could be seen with chromatin dispersion halos. Furthermore, there were differences among boars when comparing sperm rSDF (p < .05). Farrowing rate was not affected by SDF at T0 (r = .38, p = .75), nor was litter size (r = .16, p = .70). With the present experimental conditions, we have not been able to show a relationship between sperm DNA fragmentation at ejaculation and reproductive performance. However, this could be a result of the low number of ejaculates and boars used.     

Immunohistochemical study of Ki‐67 protein,androgen receptor,and estrogen receptor beta in testicular tissues of male pigs immunocastrated with different times of GnRH vaccination

This study aimed to investigate the immunoexpression of Ki‐67 protein, androgen receptor (AR), and estrogen receptor beta (ERβ) in testicular tissues of male pigs immunocastrated using GnRH vaccine (Improvac?, Zoetis Co., Ltd., Thailand) with different times. Totally, 30 male pigs were classified by castration protocol into three groups: T1 (n = 10) consisted of pigs immunocastrated at 14 and 18 weeks of age, T2 (n = 10) included pigs immunocastrated at 9 and 19 weeks of age, and C (n = 10) contained intact pigs. The results revealed that testicular length of pigs in C was longer than that of both T1 (8.1 ± 0.76 vs 6.5 ± 0.5 cm, p < 0.001) and T2 (8.1 ± 0.76 vs 6.9 ± 1.0, p = 0.007). Spearman correlation coefficients showed negative correlation between testicular length and H‐score of AR (r = ?0.38, p = 0.037), as well as positive correlation between testicular length and Ki‐67 index (r = 0.602, p < 0.001). Generally, mean Ki‐67 index and mean H‐scores of AR and ERβ of pigs in T1 were not different from those in T2 (p > 0.05). However, mean Ki‐67 index and mean AR H‐scores of T1 and T2 were significantly different from C group (p < 0.05). In summary, the immunocastration significantly affected testicular length, including expressions of Ki‐67, AR, and ERβ in pig testes. Moreover, the duration between two shots of GnRH vaccine could be extended from 4 to 10 weeks without difference in Ki‐67 protein, AR, and ERβ immunoexpressions.     

In vitro studies of Norwegian Red bovine semen immobilized and cryopreserved in alginate solid gel network

Development of new semen cryopreservation techniques improving sperm survival and ensuring availability of viable spermatozoa for a prolonged time‐period after AI is promising tools to reduce sensitivity of timing of AI and enhance overall fertility. The SpermVital^® technology utilizes immobilization of bull spermatozoa in a solid network of alginate gel prior to freezing, which will provide a gradual release of spermatozoa after AI. The objective of this study was to compare post‐thaw sperm quality and in vitro sperm survival over time of Norwegian Red bull semen processed by the SpermVital^® (SV) technology, the first commercialized production line of SpermVital^® (C) and by conventional procedure applying Biladyl^® extender (B). Post‐thaw sperm motility was not significantly different between SV, C and B semen (p > .05). However, sperm viability and acrosome intactness were higher for SV than C and B semen (p < .05). Small differences in DNA quality were observed (p < .05). Sperm viability after storage in uterus ex vivo was higher for SV than for C semen (p < .05). Furthermore, sperm survival in vitro over time at physiological temperature was significantly higher for SV semen than C semen as well as B semen during the incubation period of 48 hr (p < .05). In conclusion, the SpermVital^® technology is improved and is more efficient in conserving post‐thaw sperm quality and results in higher sperm viability over time in vitro for SV than for C and B semen.     

Pharmacokinetics and antinociceptive effects of the soluble epoxide hydrolase inhibitor t‐TUCB in horses with experimentally induced radiocarpal synovitis

This study determined the pharmacokinetics, antinociceptive, and anti‐inflammatory effects of the soluble epoxide hydrolase (sEH ) inhibitor t ‐TUCB (trans ‐4‐{4‐[3‐(4‐Trifluoromethoxy‐phenyl)‐ureido]‐cyclohexyloxy}‐benzoic acid) in horses with lipopolysaccharide (LPS )‐induced radiocarpal synovitis. A total of seven adult healthy mares (n  = 4–6/treatment) were administered 3 μg LPS into one radiocarpal joint and t ‐TUCB intravenously (i.v.) at 0 (control), 0.03, 0.1, 0.3, and 1 mg/kg in a blinded, randomized, crossover design with at least 3 weeks washout between. Two investigators independently assigned pain scores (at rest, walk and trot) and lameness scores before and up to 48 hr after t ‐TUCB /LPS . Responses to touching the joint skin to assess tactile allodynia, plasma, and synovial fluid (SF ) t ‐TUCB concentrations were determined before and up to 48 hr after t ‐TUCB /LPS . Blood and SF were collected for clinical laboratory evaluations before and up to 48 hr after t ‐TUCB /LPS . Areas under the curves of pain and lameness scores were calculated and compared between control and treatments. Data were analyzed using repeated measures ANOVA with Dunnett or Bonferroni post‐test. p  < .05 was considered significant. Data are mean ± SEM . Compared to control, pain, lameness, and tactile allodynia were significantly lower with 1 mg/kg t ‐TUCB , but not the other doses. For 0.1, 0.3, and 1 mg/kg t ‐TUCB treatments, plasma terminal half‐lives were 13 ± 3, 13 ± 0.5, and 24 ± 5 hr, and clearances were 68 ± 15, 48 ± 5, and 14 ± 1 ml hr^?1 kg^?1. The 1 mg/kg t ‐TUCB reached the SF at high concentrations. There were no important anti‐inflammatory effects. In conclusion, sEH inhibition with t ‐TUCB may provide analgesia in horses with inflammatory joint pain.     

Growth and physiological responses of growing pigs to wheat–corn distillers dried grains with solubles

Gaining a detailed knowledge on the impact of a feedstuff on pig growth and physiological responses is critical for its effective utilization. Thus, the purpose of this study was to investigate the effect of distillers dried grains with solubles derived from co‐fermentation of wheat and corn (wcDDGS) on performance, carcass and visceral organ weights, whole‐body O₂ consumption and heat production (HP) in growing barrows. The experimental diets were as follows: corn–soybean meal diet (Control), Control + 15% wcDDGS and Control + 30% wcDDGS. In Exp. 1, 48 pair‐housed pigs of average BW 18.6 ± 1.5 kg (mean ± SD) were allotted to the 3 diets (n = 8). Pigs had free access to water and feed for a 28‐day period during which ADG and ADFI were calculated weekly. Thereafter, 1 pig/pen was killed to measure carcass and visceral organ weights. Overall, wcDDGS linearly decreased (p < 0.05) ADFI and ADG but had no effect on G:F (p > 0.10). The ADFI was 1.55, 1.45 and 1.36 kg/day for diets containing 0, 15 and 30% wcDDGS respectively; corresponding values for ADG were 0.79, 0.75 and 0.67 kg/day respectively. A linear decline (p = 0.01) in eviscerated hot carcass weight was observed as dietary wcDDGS increased. In Exp. 2, 18 pigs of average BW 20.4 ± 2.4 kg (mean ± SD) were individually housed in metabolism crates and fed the 3 diets (n = 6) at 550 kcal ME kg BW^?0.60day for a 16‐day period followed by measurement of O₂ consumption using an indirect calorimeter. Diet had no effect (p > 0.10) on whole‐body O₂ consumption and HP. In conclusion, increasing wcDDGS content in growing pig diets linearly reduced ADFI, ADG and eviscerated hot carcass weight but had no effect on G:F, visceral organ weights or HP.     

Collection of peripheral blood CD34+ progenitor cells from healthy dogs and dogs diagnosed with lymphoproliferative diseases using a Baxter-Fenwal CS-3000 Plus blood cell separator

Background

Canine peripheral blood mononuclear cell (PBMC) apheresis using a Baxter‐Fenwal CS‐3000 Plus automated blood cell separator has not been reported.

Objective

To determine the feasibility and safety of using a CS‐3000 Plus blood cell separator with a small volume separation container holder (SVSCH) and small volume collection chamber (SVCC) to harvest canine PBMCs from dogs weighing <50 kg.

Animals

Eight healthy mongrel dogs and 11 client‐owned dogs in clinical remission for lymphoproliferative diseases (LPD).

Methods

In this prospective study, aphereses were performed using a Baxter‐Fenwal CS‐3000 Plus blood cell separator, with or without recombinant human granulocyte colony‐stimulating factor (rhG‐CSF) treatment.

Results

Aphereses from 6 healthy dogs given rhG‐CSF yielded an average of 1.1 × 10⁷ ± 8.2 × 10⁶ CD34+ cells/kg. Aphereses from LPD dogs given rhG‐CSF yielded an average of 5.4 × 10⁶ ± 3.25 × 10⁶ CD34+ cells/kg (P = .17). Higher hematocrit in both groups of dogs receiving rhG‐CSF correlated with an increased number of CD34+ cells/kg harvested (healthy, P = .04; LPD, P = .05). Apheresis was well tolerated by all dogs.

Conclusions and Clinical Importance

Canine PBMC apheresis using the Baxter‐Fenwal CS‐3000 Plus cell separator with an SVSCH and SVCC is a feasible and safe option for harvesting an adequate number of CD34+ peripheral blood progenitor cells from dogs weighing ≥17 kg for hematopoietic cell transplantation.     

Effects of dietary protein levels and 2‐methylbutyrate on ruminal fermentation,nutrient degradability,bacterial populations and urinary purine derivatives in Simmental steers

The objective of this study was to evaluate the effects of dietary crude protein (CP ) levels and 2‐methylbutyrate (MB ) supplementation on ruminal fermentation, bacterial populations, microbial enzyme activity and urinary excretion of purine derivatives (PD ) in Simmental steers. Eight ruminally cannulated Simmental steers, averaging 18 months of age and 465 ± 8.6 kg of body weight (BW ), were used in a replicated 4 × 4 Latin square design by a 2 × 2 factorial arrangement. Low protein (98.5 g CP /kg dry matter [LP ] or high protein (128.7 g CP /kg dry matter [HP ]) diets were fed with MB supplementation (0 g [MB ?] or 16.8 g steer^?1 day^?1 [MB +]). Steers were fed a total mixed ration with dietary corn straw to concentrate ratio of 50:50 (dry matter [DM ] basis). The CP  × MB interaction was observed for ruminal total VFA , molar proportions of acetate and propionate, acetate to propionate ratio, ammonia‐N, effective degradability of neutral detergent fibre (NDF ) and CP , microbial enzyme activity, bacterial populations and total PD excretion (p  < .05). Ruminal pH decreased (p  < .05), but ruminal total VFA concentration increased (p  < .05) with increasing dietary CP level or MB supplementation. Acetate molar proportion increased (p  = .043) with MB supplementation, but was not affected by dietary CP level. Propionate molar proportion decreased (p  < .05) with increasing dietary CP level or MB supplementation. Consequently, acetate‐to‐propionate ratio increased (p  = .001) with MB supplementation, but was not affected by dietary CP level. Ruminal ammonia‐N content increased (p  = .034) with increasing dietary CP level, but decreased (p  = .012) with MB supplementation. The effective degradability of NDF and CP increased (p  < .05) with increasing dietary CP level or MB supplementation. Microbial enzyme activity, bacterial populations and total PD excretion also increased (p  < .05) with increasing dietary CP level or MB supplementation. The results indicated that ruminal fermentation, nutrient degradability, microbial enzyme activity, ruminal bacterial populations and microbial protein synthesis improved with increasing dietary CP level or MB supplementation in steers.     

Exogenous paraoxonase‐1 during oocyte maturation improves bovine embryo development in vitro

Paraoxonase‐1 (PON1) is an enzyme found in serum and follicular fluid that protects cell membrane and circulating lipids against oxidative damage. The aims of this study were to measure the direct effects of recombinant PON1 (rPON1) on bovine oocyte maturation at the molecular level (gene expression) and to measure the carry‐over effects of PON1 on pre‐implantation embryo development in vitro. COCs were submitted to IVM with the addition of 0.0, 0.02, 0.04 and 0.08 mg ml^?1 of rPON1, corresponding to an average PON1 arylesterase enzyme activity of 2.2 ± 0.4, 15.5 ± 1.5, 30.2 ± 3.0 and 57.9 ± 5.0 U ml^?1, respectively. The results indicated that addition of rPON1 during IVM improved embryo development in a dose‐dependent manner as D7 embryo development was 22.2%, 29.4%, 32.2% and 37.0% for the treatment groups, respectively (p = 0.02). In conclusion, addition of PON1 enzyme during IVM exerted dose‐related positive effects on embryo development rates to blastocysts.