101 Assessment of semen traits in servals (Leptailurus serval) and Canada lynx (Lynx canadensis)

2019 ◽  
Vol 31 (1) ◽  
pp. 176
Author(s):  
R. González ◽  
A. Moresco ◽  
A. Miller ◽  
H. Bateman ◽  
L. Vansandt ◽  
...  
Keyword(s):  
Repeated Measures ◽  
Sperm Quality ◽  
Egg Yolk ◽  
Domestic Cat ◽  
Lynx Lynx ◽  
Cleavage Rate ◽  
Urethral Catheterization ◽  
Normal Morphology ◽  
Canada Lynx ◽  
Acrosome Integrity

Servals and Canada lynx are managed by species survival plans in North American zoos, but current populations are not sustainable. Increased knowledge of their reproductive biology would benefit breeding management and development of assisted reproductive techniques. The aims of our study were to (1) evaluate effectiveness of urethral catheterization and electroejaculation (EEJ) for semen collection; (2) characterise basal seminal traits; and (3) compare effectiveness of semen cryopreservation methods. Semen was collected from 6 servals and 9 Canada lynx via a urinary catheter (3.5 Fr×22 cm, inserted 15cm into the urethra), followed by EEJ under dexmedetomidine-ketamine anaesthesia. To assess the effect of seasonality on lynx seminal traits, semen was collected before (late January), during (mid-February to mid-March), and after (early April) the peak breeding season. Serval and lynx semen were frozen by conventional slow freezing (i.e. in 0.25-mL straws cooled to 4°C for 2h and frozen in LN vapor) in a soy lecithin-based (SOY) or egg yolk-based (TEY) extender with 4% glycerol and by ultra-rapid freezing (URF; direct pelleting into LN at ≈104°C/min) in SOY medium with 0.2M sucrose. To evaluate post-thaw sperm function in servals, heterologous IVF of domestic cat oocytes was performed, with cleavage rate assessed at 48h post-insemination. Data were analysed by one-way or repeated-measures ANOVA. Data are mean±standard deviation. Sperm recovery by urethral catheterization was negligible in both species, but EEJ allowed sperm collection in all males. Lynx seminal traits were similar during breeding and nonbreeding seasons. Testicular volume (4.81±1.17cm3) and sperm quality (13±11×106 sperm/ejaculate; 49±14% motility; 29±12% normal morphology; 74±13% acrosome integrity) were consistent with previous findings in the lynx genus. Post-thaw sperm quality in lynx has not yet been evaluated. In servals, testes volume was 6.56±2.11cm3 with good sperm quality for most males (46±36×106 sperm/ejaculate; 75±20% motility; 56±36% normal morphology; 84±7% acrosome integrity). Post-thaw, serval sperm acrosome integrity (31±15, 21±13, 24±13% at 0h for TEY, SOY, and URF, respectively; P>0.05) and motility (40±21% at 0h, 20±11% at 6h for TEY; 24±19% at 0h, 6±4% at 6h for SOY; 21±16% at 0h, 3±2% at 6h for URF; treatment: P>0.05; time: P<0.05; interaction: P>0.05) declined substantially. However, thawed sperm could fertilize domestic cat oocytes with no difference among treatments in cleavage success (53±6, 47±4, or 49±14%; TEY, SOY, and URF, respectively; P>0.05), indicating that standard freezing methods are effective in servals. Our findings provide zoos with valuable information about normative reproductive traits in both species. Supported by IMLS and the Roger & Kathy Gross Post-doctoral Fellowship.

105 Functionality evaluation of two extenders for Leopardus geoffroyi sperm cryopreservation by interspecific IVF with domestic cat oocytes

2019 ◽  
Vol 31 (1) ◽  
pp. 178
Author(s):  
A. J. Sestelo ◽  
M. D. Rodriguez ◽  
N. Gañan ◽  
D. F. Salamone ◽  
L. Barañao ◽  
...  
Keyword(s):  
Sperm Quality ◽  
Blastocyst Stage ◽  
Domestic Cat ◽  
Freezing Process ◽  
Sperm Cryopreservation ◽  
Step Method ◽  
Cleavage Rate ◽  
Sperm Suspension ◽  
Acrosome Integrity

Even though knowledge in sperm cryopreservation of endangered felids advanced in recent years, very little is known about suitable protocols to cryopreserve sperm from Leopardus geoffroyi (LG). In the present study, sperm obtained by electroejaculation from 5 different males were cryopreserved in either a Tes-Tris- or a lactose-based diluent (Gañan et al. 2009 Theriogenology 72, 341-352) with modifications in the freezing process using a one-step method: straws were placed horizontally on a metal rack, 4cm above the surface of liquid nitrogen in a styrofoam box, and kept for 10min before plunging them in LN. The objectives were to (1) compare in vitro motility and acrosome status of LG sperm cryopreserved in both extenders and (2) test functionality of LG sperm cryopreserved in both extenders through their ability to fertilize mature domestic cat oocytes. Straws were thawed by exposing them to air for 10s and then immersing them in a water bath at 37°C for 30s. The contents of the straws were poured into a sterile 1.5-mL microtube prewarmed to 37°C. The sperm suspension was diluted (1:3 vol/vol) by the slow (drop by drop) addition of a modified Tyrode’s solution. Sperm parameters, percentage of motile spermatozoa, and quality of motility were assessed and sperm motility index (SMI) was calculated as follows: [% motile sperm+(quality×20)]/2. Acrosome integrity was assessed by staining with Coomassie brilliant blue. For IVF, in vitro-matured domestic cat oocytes (n=238 Tes-Tris, n=239 lactose) were co-incubated with 0.5×105 motile spermatozoa/mL under 5% CO2 in air at 38.5°C for 18-20h (Pope et al. 2006 Methods Mol. Biol. 254, 227-244). Presumptive zygotes were cultured in vitro in 50-µL drops of modified Tyrode’s medium at 38.5°C in 5% CO2, 5% O2, 90% N2 atmosphere. Cleavage was assessed 48h postfertilization, and 5% FBS was added at Day 5 of in vitro culture. Blastocyst stage was evaluated at Day 8. Data was analysed by Fisher’s exact test using GraphPad Prism 6.0 (GraphPad Inc., San Diego, CA, USA), significant at P<0.05. Results, mean (±standard error of the means), showed that SMI and acrosome integrity (pre- and post-thawing) were similar for both extenders: prethawed (SMI=56±3.3v. 59±5.5; acrosome integrity=88±3.0% v. 90±2.0%), and post-thawed (SMI=46±5.0v. 44±7.0; acrosome integrity=57±7.5% v. 68±2.4%) Tes-Tris v. lactose, respectively. For IVF, results showed a high cleavage rate in both groups (117/238, 49% v. 117/239, 49%), and a high development to morula (96/238, 40% v. 94/239, 39%) and to the blastocyst stage (61/238, 26% v. 51/239, 21%) for all males Tes-Tris v. lactose, respectively. There were no significant differences between groups at any development stage. In conclusion, we found that both extenders can be used to cryopreserve LG sperm maintaining functional conditions and that fertilizing capacity can be tested using in vitro-matured domestic cat oocytes.

56 EFFECTS OF PHYLOGENIC GENERA OF RECIPIENT CYTOPLASTS ON DEVELOPMENT AND VIABILITY OF CANADA LYNX (LYNX CANADENSIS) CLONED EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 186 ◽  
Author(s):  
M. C. Gómez ◽  
J. I. Lyons ◽  
C. E. Pope ◽  
M. Biancardi ◽  
C. Dumas ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Donor Cell ◽  
Domestic Cat ◽  
Developmental Competence ◽  
Lynx Lynx ◽  
Phylogenetic Distance ◽  
Lynx Canadensis ◽  
Gestational Length ◽  
Canada Lynx ◽  
Oocyte Donor

Canada lynx (Lynx canadensis; CL) once occupied 16 states in the Unites States of America, but small populations remain in only 3 states. Interspecies-somatic cell nuclear transfer (Is-SCNT) offers the possibility of preventing their extinction; however, developmental constraints on Is-SCNT embryos are proportional to the phylogenetic distance between the donor cell and the recipient oocyte. Mitochondrial DNA (mtDNA) heteroplasmy may be involved in nuclear-cytoplasmic incompatibilities, thus inhibiting development of cloned embryos at the time of genomic activation. Minimizing the phylogenetic distance between the donor cell and recipient oocyte may enhance development of clone embryos. Caracal (Caracal caracal) may be suitable as an oocyte donor for SCNT and a recipient of CL cloned embryos because caracals hybridize with other felid species and share physical characteristics with the lynx family, marked by being previously classified in the lynx genera and having similar gestational length. To ensure compatibilities between the donor nuclei of the CL and the mitochondria of recipient oocytes, we (1) compared in vitro development of CL cloned embryos reconstructed with domestic cat (Felis catus; DSH) or caracal cytoplasts, (2) examined the mtDNA genotypes in CL cloned embryos, and (3) evaluated in vivo developmental competence of CL cloned embryos after transfer into caracal recipients. A total of 160 and 217 preovulatory oocytes were collected by laparoscopy from gonadotropin-treated caracals (n = 8) and DSH (n = 10) and used as recipient cytoplasts for reconstructing CL embryos. Results indicated that the phylogenetic genera of recipient cytoplasts did not affect embryo cleavage at Day 2 (caracal 50/55, 91% v. DSH 63/65, 97%), but development of CL cloned embryos to the blastocyst stage was higher when caracal oocytes were used as recipient cytoplasts (15/50; 30%) than with DSH cytoplasts (9/63, 14%; P < 0.05). The extent of mtDNA homoplasmy or heteroplasmy in CL cloned embryos was calculated by the number of single nucleotide polymorphisms (SNP) derived from the DSH or caracal oocyte donors and from the somatic cell donor CL. DNA was isolated from 25 and 35 CL cloned embryos reconstructed with caracal or DSH cytoplasts, respectively. All amplified products after PCR were sequenced and SNP analyzed. All CL embryos reconstructed with DSH cytoplasts were homoplasmic, carrying mtDNA only from the DSH oocyte donor (n = 35; SNP DSH = 2-6). Embryos reconstructed with caracal cytoplasts were homoplasmic for CL mtDNA (n = 9; SNPCL = 10-12) or heteroplasmic (caracal × CL, n = 17; SNPCL = 7-9; SNP caracal = 2-3). A total of 69 (mean = 34.5 ± 4.9 per caracal) and 70 (mean = 35.0 ± 9.8 per caracal) CL cloned embryos reconstructed with caracal and DSH cytoplasts, respectively, were transferred into 4 caracal recipients; however, no pregnancies were established. In summary, Is-SCNT between 2 phylogenetically closer species favors retention of the donor’s mitochondria, which might lead to a better nucleo-cytoplasmic interaction for reprogramming of donor nucleus.

97 IMPROVED CRYOPRESERVATION OF DOMESTIC CAT SPERMATOZOA IN A SOY LECITHIN-BASED EXTENDER

2011 ◽  
Vol 23 (1) ◽  
pp. 153 ◽  
Author(s):  
M. M. Vick ◽  
H. L. Bateman ◽  
W. F. Swanson
Keyword(s):  
Sperm Motility ◽  
Room Temperature ◽  
Egg Yolk ◽  
Tris Buffer ◽  
Domestic Cat ◽  
Dna Integrity ◽  
Freezing And Thawing ◽  
Soy Lecithin ◽  
Acridine Orange Staining ◽  
Acrosome Integrity

Development of a chemically defined, plant-based cryopreservation media would reduce extender variability and the potential for transmission of zoonotic pathogens compared with traditional egg-yolk-based extenders. The objective of this study was to compare effects of egg yolk- and soy lecithin-based cryopreservation media and the temperature of glycerol addition on sperm parameters following freezing and thawing of domestic cat spermatozoa. Fresh semen was collected by manual stimulation on 3 separate occasions from 4 adult male cats. Each ejaculate was washed to remove seminal plasma, divided into 4 equal aliquots, and extended at room temperature in one of the following treatments: 1) TEST-egg yolk (Irvine Scientific Inc., Santa Ana, CA, USA) medium with 4% glycerol (EYG); 2) TEST-egg yolk, with 4% glycerol added after cooling to 5°C (EY); 3) TES-Tris buffer with soy-lecithin (1%) and 4% glycerol (SLG); and 4) TES-Tris buffer with 1% soy-lecithin, and 4% glycerol added after cooling to 5°C (SL). Sperm progressive motility (%) and rate of progressive movement (scale of 0–5) were evaluated at 0, 1, 3, 6, and 24 h post-thaw. Sperm capacitation (chlortetracycline staining), acrosome integrity (FITC-PNA staining), and DNA integrity (acridine orange staining) were assessed at 15 min post-thaw. Data were exponentially transformed to achieve normal distribution and then subjected to GLM analysis to determine effects of media and temperature of glycerol addition on sperm traits. At 0 and 1 h post-thaw, acrosome integrity, DNA integrity and % sperm motility did not differ (P > 0.05) among treatments. However, % sperm motility was greater in the soy-based media compared to egg yolk-based media at 3, 6, and 24 h post-thaw (Table 1; P < 0.05). A higher percentage of uncapacitated spermatozoa were present in soy-based compared to egg-yolk based cryopreservation media (63.9 ± 9.3 v. 51.2 ± 11.5, respectively; P < 0.05), regardless of temperature of glycerol addition. Finally, addition of glycerol at 5°C resulted in higher % sperm motility compared to room temperature at 6 and 24 h post-thaw in both medium types (Table 1; P < 0.05). Our results suggest that use of a chemically defined, soy-based medium improves long-term motility and capacitation status of frozen–thawed domestic cat spermatozoa compared with cryopreservation in a traditional egg yolk-based extender. Table 1.Motile spermatazoa and motility score at 3, 6, and 24 h

scholarly journals Effect of whole and clarified egg yolk based extenders on post-thaw semen quality in bulls.

2021 ◽  
Author(s):  
Prosper Kamusasa ◽  
Eddington Gororo ◽  
Fungayi Primrose Chatiza
Keyword(s):  
Semen Quality ◽  
Sperm Quality ◽  
Egg Yolk ◽  
Quality Parameters ◽  
Normal Morphology ◽  
Semen Cryopreservation ◽  
Bull Semen ◽  
Inclusion Level ◽  
Whole Egg

Abstract This study was conducted to evaluate the comparative cryoprotective effects of whole egg yolk and clarified egg yolk on post thaw sperm quality parameters and to determine the optimum clarified egg yolk inclusion level (10-20%) in semen extenders for Mashona bull semen cryopreservation. It was shown that there was a significant decrease in sperm quality variables following cryopreservation. Semen quality increased with the concentration of clarified egg yolk, indicating a positive relationship between egg yolk LDL concentration and maintenance of in vitro sperm quality. The 20% clarified egg yolk (CEY20) extender treatment gave post-thaw motility, viability and normal morphology values which were comparable to the control (20% whole egg yolk, WEY20). The 10% clarified egg yolk concentration gave the least post-thaw quality values and the greatest proportion of defective spermatozoa. This experiment found no advantage of replacing whole egg yolk with up to 15% clarified egg yolk in Mashona bull semen cryopreservation. However, 20% clarified and 20% whole egg yolk performed similarly in the maintenance of post-thaw sperm motility, viability and normal morphology.

scholarly journals Assessment of semen quality, sperm cryopreservation and heterologous IVF in the critically endangered Iberian lynx (Lynx pardinus)

2009 ◽  
Vol 21 (7) ◽  
pp. 848 ◽  
Author(s):  
Natalia Gañán ◽  
Raquel González ◽  
J. Julián Garde ◽  
Fernando Martínez ◽  
Astrid Vargas ◽  
...  
Keyword(s):  
Semen Quality ◽  
Statistical Significance ◽  
Egg Yolk ◽  
Domestic Cat ◽  
Critically Endangered ◽  
Lynx Lynx ◽  
Sperm Cryopreservation ◽  
Iberian Lynx ◽  
Lynx Pardinus

Semen traits and factors affecting sperm cryopreservation were assessed in the Iberian lynx (Lynx pardinus), a species regarded as the most endangered felid in the world. For cryopreservation, semen was washed, resuspended in a Tes–Tris-based diluent (TEST) or a Tris-based diluent (Biladyl), both with 20% egg yolk and 4% glycerol, loaded into straws, cooled to 5°C using an automated programmable system and frozen on nitrogen vapour. Heterologous IVF of in vitro-matured domestic cat oocytes was used to test the fertilising ability of cryopreserved spermatozoa. Electroejaculates from five males were obtained. Characterisation of the electroejaculates revealed mean (± s.e.m.) values of 3.3 ± 0.6 × 106 total spermatozoa, 73.6 ± 4.6% motile spermatozoa, 23.7 ± 4.0% morphologically normal spermatozoa and 40.7 ± 2.3% spermatozoa with intact acrosomes. After thawing a higher percentage of motile spermatozoa was seen in TEST than in Biladyl (34.0 ± 6.2% v. 7.5 ± 4.8%, respectively; P < 0.05); however, there were no differences in the percentage of intact acrosomes between the two diluents. Iberian lynx spermatozoa fertilised domestic cat oocytes in vitro, with higher fertilisation rates observed for spermatozoa cryopreserved in TEST than in Biladyl, although the difference did not reach statistical significance (20.5 ± 4.5% v. 11.5 ± 6.8%, respectively). There were positive significant relations between the fertilisation rates and both the percentage of normal spermatozoa and the percentage of spermatozoa with an intact acrosome before cryopreservation (P = 0.04). This first report of the collection and cryopreservation of Iberian lynx semen and analysis of fertilising ability is an important step in the development of assisted reproductive techniques for this critically endangered felid species.

scholarly journals 97 Social Dominance does not Affect Semen Quality in African Wild Dogs (Lycaon pictus)

2018 ◽  
Vol 30 (1) ◽  
pp. 188
Author(s):  
F. Van den Berghe ◽  
M. C. J. Paris ◽  
Z. Sarnyai ◽  
M. B. Briggs ◽  
R. P. Millar ◽  
...  
Keyword(s):  
Breeding Season ◽  
Prostate Volume ◽  
Sperm Quality ◽  
Dna Integrity ◽  
Normal Morphology ◽  
Progressive Motility ◽  
African Wild Dogs ◽  
Acrosome Integrity ◽  
Urine Contamination ◽  
Wild Dogs

Sperm banking and AI could benefit conservation of endangered African wild dogs (AWD). However, it is not clear whether their strict dominance hierarchy causes subfertility in subdominant males that typically do not breed. Our study investigated the effect of dominance on male reproductive parameters, including faecal glucocorticoids (fGCM) and androgens (fAM), testis and prostate volume, preputial gland size, semen collection success, and the number, motility, morphology, viability, acrosome integrity (PSA-FITC), and DNA integrity (TUNEL) of spermatozoa collected by electroejaculation. Samples were obtained from n = 12 captive AWD (4 US packs) in the pre-breeding season and n = 28 captive AWD (n = 11 from 4 US packs; n = 17 from 3 Namibian packs) in the breeding season. Male hierarchy was clearly determined by behavioural observations in all but 1 Namibian pack. Data were grouped by dominance status and means were compared by ANOVA or t-test; P ≤ 0.05 was significant. In the pre-breeding season, there was no significant difference in body weight, fGCM, fAM, or prostate and testis volume between dominance groups. Semen was successfully collected from all alphas but only half the subdominants; urine contamination was negatively associated with dominance. Sperm quality was low (17.3 ± 10.2% total motility, 12.8 ± 8.5% progressive motility, 27.4 ± 11.5 × 106 ejaculated spermatozoa, 40.6 ± 9.8% normal morphology, 63.1 ± 5.1% viability, 72.6 ± 5.2% acrosome integrity) with no difference observed in any parameter except progressive motility and normal sperm morphology, which were significantly lower in subdominants (27.7 ± 16.8% v. 0.0 ± 0.0% and 59.8 ± 13.0% v. 21.4 ± 5.7%). From pre-breeding to breeding season, testis and prostate volume increased significantly, particularly in beta and gamma males respectively. Prostate volume was higher in alpha than beta males (16.0 ± 6.4 cm3 v. 5.7 ± 1.4 cm3), but testis volume, body weight, fAM, and fGCM did not differ between dominance groups (12.0 ± 0.9 cm3, 28.5 ± 0.8 kg, 0.51 ± 0.07 µg g−1, and 30.6 ± 2.3 ng/g of dry weight). Semen was successfully collected from 75% of males with reduced urine contamination. Collection success, urine contamination, and preputial gland size were not associated with dominance. Sperm quality improved with significantly greater number, viability, and total motility. However, sperm quality did not differ between dominance groups (47.4 ± 6.7% total motility, 30.5 ± 5.8% progressive motility, 32.3 ± 9.2 × 106 ejaculated spermatozoa, 50.9 ± 5.2% normal morphology, 74.4 ± 4.2% viability, 85.6 ± 3.0% acrosome integrity, and 99.7 ± 0.1% DNA integrity). In conclusion, subdominant males are at higher risk of urine contamination and have lower sperm motility and normal morphology when semen is collected in the pre-breeding season. However, their semen is of similar quality to dominant males in the breeding season, indicating that reproductive suppression of subdominant males is only behavioural. Thus, AWD males of all social ranks in the breeding season are suitable candidates for sperm banking.

114 GETTING THE YOLK OUT: THE USE OF A SOY LECITHIN-BASED CRYOMEDIUM FOR SEMEN BANKING IN THE PALLAS’ CAT AND FISHING CAT

2017 ◽  
Vol 29 (1) ◽  
pp. 165
Author(s):  
L. M. Vansandt ◽  
H. L. Bateman ◽  
J. Newsom ◽  
W. F. Swanson
Keyword(s):  
Repeated Measures ◽  
Assisted Reproductive Technologies ◽  
Egg Yolk ◽  
Reproductive Technologies ◽  
Domestic Cat ◽  
Significant Advance ◽  
Sperm Number ◽  
Soy Lecithin ◽  
Embryo Cleavage ◽  
Time Point

Classically, semen from nondomestic felids has been frozen in an egg yolk-based cryomedium (TEY). However, the inclusion of egg yolk is problematic, resulting in possible batch to batch variation, microbial contamination, and regulatory concerns with international semen transport. Our previous research revealed that a soy lecithin-based cryopreservation medium (SOY) was superior to TEY for freezing domestic cat sperm. However, there is limited information on the efficacy of SOY in nondomestic felids. In the present study, our aim was to assess the effect of SOY v. TEY on motility, acrosome status, and fertilizing capacity of frozen-thawed sperm in 2 wild cat species: the Pallas’ cat (Otocolobus manul) and fishing cat (Prionailurus viverrinus). Semen was collected via electroejaculation from male cats (n = 3/species), split into 2 aliquots, extended in either SOY or TEY (containing 4% glycerol), slow-cooled, and frozen in straws over liquid nitrogen vapor. Sperm motility (percent progressively motile, PM; rate of progressive motility on 1 to 5 scale, RPM) was evaluated at 0, 1, 3, 6, and 24 h post-thaw and acrosome status (AS) was assessed via fluorescein isothiocyanate-peanut agglutinin staining at 0 and 6 h post-thaw. Heterologous IVF was performed using oocytes (n = 10–15/experimental unit, 2 replicates) collected laparoscopically from gonadotropin-treated domestic cats. At 48 h post-insemination, Hoechst33342 staining was used to determine oocyte stage, number of blastomeres, and number of accessory sperm bound to the zona pellucidae of embryos and mature oocytes. The PM, RPM, and AS were analysed with repeated-measures ANCOVA, using pre-freeze values as a covariate. Embryo cleavage % and accessory sperm number were analysed with ANOVA. All data are reported as mean ± s.e. In the Pallas’ cat, PM, RPM, and AS of SOY-treated sperm (35.0 ± 7.6% motile, 2.8 ± 0.4 RPM, 41.2 ± 5.4% intact; 0 h) did not differ (P > 0.05) from TEY-treated sperm (36.7 ± 4.4% motile, 2.8 ± 0.4 RPM, 45.3 ± 9.4% intact; 0 h) at any post-thaw time point. Similarly in the fishing cat, post-thaw PM, RPM, and AS of SOY-treated sperm (38.3 ± 7.3% motile, 2.6 ± 0.2 progression, 20.0 ± 2.9% intact; 0 h) did not differ (P > 0.05) from TEY-treated sperm (31.7 ± 8.7% motile, 2.4 ± 0.3 RPM, 18.3 ± 6.4% intact; 0 h) at any time point. In the Pallas’ cat, neither embryo cleavage % (42.6 ± 7.0% SOY; 61.4 ± 14.8% TEY) nor accessory sperm number (12.2 ± 2.8 SOY; 18.1 ± 4.0 TEY) differed (P > 0.05) between treatments. Fishing cat results were similar, with no difference (P > 0.05) between SOY and TEY for cleavage % (60.4 ± 9.4% SOY; 47.9 ± 5.0% TEY) or accessory sperm number (4.6 ± 1.0 SOY; 5.5 ± 1.0 TEY). Collectively, these findings demonstrate that our SOY medium is an effective alternative to TEY for sperm cryopreservation in 2 nondomestic felid species. The replacement of an egg yolk-based cryomedium with a chemically defined, animal protein-free option represents a significant advance in quality control and biosecurity for cat semen banking and may enhance the use of assisted reproductive technologies for population management of imperiled felids. Research was funded by the Institute of Museum and Library Services.

scholarly journals A Comparison of Freezing Methods and Diluents Types on Post-Thaw Sperm Quality of Ram Sperm

2020 ◽  
Vol 7 (2) ◽  
pp. 235-241
Author(s):  
Pankaj Kumar Jha ◽  
M Golam Shahi Alam ◽  
Farida Yeasmin Bari
Keyword(s):  
Plasma Membrane ◽  
Sperm Quality ◽  
Membrane Integrity ◽  
Egg Yolk ◽  
Sperm Parameters ◽  
Plasma Membrane Integrity ◽  
One Step ◽  
Acrosome Integrity ◽  
Ram Sperm

The effect of freezing methods and diluents types on post-thaw sperm quality of Bangladeshi ram semen was studied. Two freezing methods and three diluents was tested as pooling effects (freezing methods or diluents) on post-thaw sperm parameters; sperm motility (SM), viability (SV), plasma membrane integrity (SPMI) and acrosome integrity (SAI), respectively. From selected ten rams, eight ejaculates were used for each freezing group (freezing methods × diluents). Semen samples were diluted by using two-steps for hand-made tris-based diluents (20% egg yolk): D1 (7% glycerol) and D2 (5% glycerol), and one-step dilution for commercial diluents: D3 (Triladyl®) at 35°C. After 4h of equilibration of temperature at 5°C, diluted semen samples was aspirated into 0.25 mL straws, and sealed. Straws were frozen in liquid nitrogen (LN2) vapour using two methods: F1 (manually in Styrofoam box, using three-steps method; +5°C to -80°C at -11.33°C/min, -80°C to -120°C at -26.66°C/min, and -120°C to -140°C at - 13.33°C/min) and F2 (programmable bio-freezer, using two-steps method; +5°C to -100°C at - 20°C/min and -100°C to -140°C at -10°C/min). Two semen straws from each batch were evaluated (37°C for 20 sec) for sperm parameters. In pool effects between freezing methods; SAI differed significantly (P < 0.001). The SM (56%) and SV (72%) were observed competitive. However, SPMI (67.58 ± 2.02%) and SAI (76.13 ± 1.42%) were higher in F1. Among diluents, SM (P < 0.006), SV (P < 0.008), SPMI (P < 0.012) and SAI (P < 0.019) differed significantly. The SM (61.25 ± 1.80%), SV (77.13 ± 1.47%), SPMI (68.31 ± 1.91%) and SAI (74.75 ± 1.64%) were highest in D3. In conclusion, the combination of manual freezing (three-steps) and handmade tris-based diluents (20% egg yolk, 5% glycerol) is suitable and sustainable method for cryopreservation of ram semen. Res. Agric., Livest. Fish.7(2): 235-241,  August 2020

scholarly journals Comparison of the Morphology and Developmental Potential of Oocytes Obtained from Prepubertal and Adult Domestic and Wild Cats

Animals ◽  
2020 ◽  
Vol 11 (1) ◽  
pp. 20
Author(s):  
Joanna Kochan ◽  
Agnieszka Nowak ◽  
Wiesława Młodawska ◽  
Sylwia Prochowska ◽  
Agnieszka Partyka ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Domestic Cat ◽  
Lynx Lynx ◽  
Panthera Tigris ◽  
Domestic Cats ◽  
Cleavage Rate ◽  
Developmental Potential ◽  
Wild Cats

The aim of the study was to compare the morphology and developmental potential of oocytes obtained from adult and prepubertal domestic cats (Felis catus) and wild cats (Lynx lynx, Leptailurus serval, Felis manul, Panthera tigris altaica). The average number of oocytes obtained from an adult domestic cat was 23 ± 11, which was significantly lower than from kittens (43 ± 29). A similar number of oocytes was derived from adult Pallas’s cats (28 ± 8), and serval (30). The lowest number of oocytes was collected from the lynx (5 ± 3). No oocytes were obtained from newborn Amur tiger while in the case of older domestic and Pallas’s cat and lynx kittens (1–3 months) 43, 48 and 41 oocytes were collected, respectively. Significant differences (p < 0.001) were observed between the number of oocytes with dark cytoplasm from adult and prepubertal animals of all analyzed species. The diameter of oocytes from adult and prepubertal animals was similar in all species, and was on average 161 ± 4 µm for oocytes with dark cytoplasm and 150 ± 18 µm for oocytes with light cytoplasm. In all species, oocytes with light cytoplasm were significantly smaller (p < 0.05) than dark ones, and their population was more diverse. Results of in vitro maturation of the domestic and wild cat′s oocytes obtained from adult and prepubertal females were similar (47–52%). The cleavage rate after in vitro fertilization (IVF) was lower for prepubertal than adult domestic cats (42 vs. 51%; p < 0.05%). Moreover, we observed differences in the quantity (28 vs. 39%; p < 0.05) and quality of blastocysts and even greater problems with hatching blastocysts from prepubertal kittens (8 vs. 19%; p < 0.001). More blastomeres were detected in blastocysts of adult cats. They also demonstrated significantly higher number of inner cell mass (ICM) (p < 0.001) and higher number of trophoblast cells (TE) (p < 0.05).

116 EFFECT OF INCUBATION OF BOVINE EPIDIDYMAL SPERMATOZOA IN SEMINAL PLASMA PRIOR TO CRYOPRESERVATION

2007 ◽  
Vol 19 (1) ◽  
pp. 175
Author(s):  
C. A. Guerrero ◽  
J. A. Jenkins ◽  
J. W. Lynn ◽  
K. R. Bondioli ◽  
R. A. Godke
Keyword(s):  
Seminal Plasma ◽  
Sperm Quality ◽  
Egg Yolk ◽  
Quality Parameters ◽  
Water Bath ◽  
Sperm Chromatin ◽  
Mitochondrial Activity ◽  
Dna Integrity ◽  
Epididymal Sperm ◽  
Acrosome Integrity

Studies examining the influence of seminal plasma on sperm function have shown both beneficial and detrimental effects. However, its effect on pre-frozen bovine epididymal sperm (BES) has not been documented. The objective of this study was to determine the effect of a 30-min incubation of BES in bovine seminal plasma (SEMP) prior to freezing on post-thaw sperm parameters. Paired testes were obtained from mature bulls (n = 10) at a local abattoir and transported to the laboratory (25–28°C) within 3–5 h postmortem. BES were harvested by multiple incisions from the caudae epididymides of each bull, pooled, and split into either Treatment A, 3 mL of egg yolk Tris-based medium (EYT) (No SEMP), or Treatment B, 3 mL of SEMP, each for 30 min in a 37°C water bath. SEMP was extracted from ejaculates of mature bulls (n = 6), and then pooled and stored at −20°C until used. Sperm from both treatments were re-suspended in EYT, adjusted to 70 × 106 sperm mL−1 and cooled to 4°C at 0.1°C min−1. Samples were diluted slowly over a 30-min period with 1:1 EYT medium containing 14% glycerol and loaded into 0.5-mL straws. Straws were frozen in liquid nitrogen vapors. For sperm analyses, straws were thawed in a water bath at 37°C for 40 s. Morphology was determined by staining with eosin-nigrosin. Viability and acrosome integrity were measured simultaneously with the combination of SYBR 14, propidium iodide, and PE-PNA. Mitochondrial activity was measured by the combination of MitoTracker Red and SYBR 14. DNA integrity was determined by acridine orange using the sperm chromatin structure assay (SCSA®; SCSA Diagnostics, Inc., Brookings, SD, USA). All assays were performed by multicolor flow cytometry. Differences between treatments were analyzed using one-way ANOVA (P &lt; 0.05). In summary, all sperm quality parameters decreased significantly after thawing in both treatments (Table 1). A significantly higher overall and progressive motility post-thaw was achieved when sperm were incubated in SEMP prior to freezing. However, no difference was detected in sperm viability, acrosome integrity, mitochondrial activity, and DNA integrity between treatments. Also, SEMP reduced the quantity of distal droplets and broken tails post-thaw over that of no SEMP. Results indicate that incubation of BES in bovine seminal plasma prior to freezing improves post-thaw sperm quality. Table 1. Effect of bovine seminal plasma (SEMP) on quality parameters (mean ± SEM) of post-thaw bovine epididymal sperm

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