The effects of rapid cooling (cold shock) of ram semen, photoperiod, and egg yolk in diluents on the survival of spermatozoa before and after freezing

(1) Nine successive ejaculates were collected from each of four Merino rams on three occasions at intervals of three days. The mean time intervals between successive ejaculates on the first, second, and third collecting days were respectively 41, 28, and 30 min. Aliquots from alternate ejaculates (first, third, fifth, seventh, and ninth) were cold-shocked before and after 1 : 4 dilution with an egg yolk–glucose–citrate diluent. Similarly diluted aliquots were stored at +2°C for 16 days and also frozen in pellet form after addition of 6% glycerol to a final 1:8 dilution. (2) Both undiluted and diluted spermatozoa showed increased susceptibility to cold shock with succession of ejaculates. Egg yolk–glucose–citrate diluent ameliorated the deleterious effect. (3) There were no marked differences in reanimation at thawing of pellet-frozen successive ejaculates. During a 6 hr incubation of the thawed semen at 37°, however, there were significant ejaculate differences in viability, the ninth collection having the lowest value. The mean reanimation of pellet-frozen spermatozoa after thawing was 58.7°. During incubation periods of 2,4, and 6 hr at 37°C, 47, 60, and 70% respectively of the original post-thawing motile spermatozoa became immotile. (4) During storage for 16 days at +2°C, the third and fifth ejaculates gave the best viability values. There were, however, no marked differences between ejaculates during the first 6 days of storage. (5) No firm relationships were found between cold shock values and subsequent viability during liquid storage or recovery after pellet-freezing.

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Improved sperm cryopreservation using cold cryoprotectant

Reproduction Fertility and Development ◽

10.1071/rd03007 ◽

2003 ◽

Vol 15 (7) ◽

pp. 377 ◽

Cited By ~ 15

Author(s):

G. N. Clarke ◽

D. Y. Liu ◽

H. W. G. Baker

Keyword(s):

Cold Shock ◽

Freezing Point ◽

Standard Procedure ◽

Egg Yolk ◽

Human Sperm ◽

In Vitro Fertilisation ◽

Donor Insemination ◽

Slow Cooling ◽

Rapid Cooling

It has generally been assumed that very rapid cooling above freezing point would be deleterious to human sperm because it would result in cold shock. Consequently, most routine cryopreservation protocols involve the use of warm (20–30°C) cryoprotectant and slow cooling above the freezing point in order to minimise the risk of cold shock. In order to test this assumption, we added an equal volume of cold (4°C) cryoprotectant in a single aliquot to warm (20, 30 or 37°C) semen to induce rapid cooling. The results of this procedure were compared with those obtained using warm cryoprotectant or with the routine cryopreservation protocol used in this laboratory. The use of cold cryoprotectant resulted in a significant (P = 0.016) improvement (mean 63%, range 42%–79%) in post-thaw motility recovery compared with a standard procedure(mean 47%, range 35%–67%) and a significant (P = 0.016) improvement in post-thaw sperm velocity. A cold glycerol/egg yolk/citrate (GEYC) mixture also gave significantly higher motility recovery than GEYC equilibrated to either room temperature (20°C) or body temperature (37°C). Sperm frozen using the cold cryoprotectant protocol were as efficient at binding to and penetrating the human zona pellucida as sperm frozen with a standard protocol.The modified cryopreservation procedure may lead to improved pregnancy rates in donor insemination and in vitro fertilisation. Further investigation is required to determine how the cold cryoprotectant improves the cryopreservation outcome.

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209FUNCTIONAL ASSESSMENT OF WHITE RHINOCEROS CERATHOTERIUM SIMUM EPIDIDYMAL SPERMATOZOA BEFORE AND AFTER CRYOPRESERVATION

Reproduction Fertility and Development ◽

10.1071/rdv16n1ab209 ◽

2004 ◽

Vol 16 (2) ◽

pp. 226 ◽

Cited By ~ 1

Author(s):

F. Martinez-Pastor ◽

F. Olivier ◽

T. Spies ◽

L. Anel ◽

P. Bartels

Keyword(s):

Plasma Membrane ◽

Assisted Reproduction ◽

Phase Contrast ◽

Egg Yolk ◽

Phase Contrast Microscopy ◽

White Rhinoceros ◽

Contrast Microscopy ◽

Before And After ◽

Epididymal Spermatozoa

Biological Resource Banks represent a potentially valuable tool for species conservation. It is, however, necessary to understand the species-specific cryopreservation process and its consequences for spermatozoa to aid in the development of assisted reproduction as a future conservation tool. The aim of this study was to assess the in vitro functionality of white rhinoceros Cerathoterium simum epididymal spermatozoa both before and after cryopreservation. Testes from a harvested white rhino bull were removed and transported at 5°C to the laboratory within 4h. The cauda epididymis was dissected out and flushed with 2mL of Tris-citrate egg yolk extender (fraction A, Biladyl, Minitüb, Germany). A 0.1mL aliquot was removed for analysis and the balance (9mL; 2mL fraction A+7mL sperm sample) mixed with an additional 27.2mL of Tris-citrate egg yolk with glycerol (fraction B, Bidadyl). The extended sample was allowed to cool to 4°C over a 6-h period before an additional 29.2mL of cooled fraction B were added (final sperm concentration=150×106mL−1). Sperm samples were loaded into 0.25-mL straws and frozen over LN2 vapor (4cm for 20min) for later assessment. Sperm straws were thawed by placing the straws in water at 37°C for 30s. Pre-freeze and post-thaw evaluations were carried out in the same manner. Media used included: HEPES for washing (20mM HEPES, 355mM sucrose, 10mM glucose, 2.5mM KOH) and HEPES saline (197mM NaCl, instead of sucrose). An aliquot was diluted with HEPES (washing) and centrifuged for 5min at 600×g; the pellet was resuspended in HEPES saline. Sperm motility (total motility %, TM;; and progressive motility %, PM) was assessed using phase contrast microscopy (×200; 37°C). Sperm plasma membrane status was assessed using the fluorescent dye, propidium iodide (50ngmL−1 in HEPES saline;; 10min, RT). Percentage of cells with plasma membranes intact (unstained;; PMI) was recorded. Mitochondrial status was assessed with the fluorescent dye, JC-1 (7.5μM in HEPES saline;; 30min, 37°C). The % of cells with an orange-stained midpiece was recorded (active mitochondria;; MIT). Resilience to hypoosmotic shock (HOS test) was assessed by diluting a sample in 100mOsm/kg HEPES saline (1:20; 15min, RT). An aliquot was stained with PI to assess plasma membrane status (HOSPMI), and the rest was fixed with formaldehyde, and % coiled tails (positive endosmosis;; HOST) was estimated using phase contrast microscopy (×400). Evaluations of PMI, MIT and HOSPMI were performed using fluorescence microscopy (×400, 450–490nm excitation filter). The results indicated that quality was good pre-freezing (TM: 60%; PMI: 86%; MIT: 100%), except for a PM value of 15%. After thawing, although there was a drop in TM (30%), there was no decrease in PM (20%). Our in vitro functional assessment indicated a loss of quality between the pre-freeze and post-thaw evaluations, but PMI and MIT maintained their pre-thaw levels (60% and 72%, respectively). The HOS test, which indicates plasma membrane integrity, decreased from the pre-freeze level (91%) to a post-thaw value of 70%. HOSTPMI was 72% pre-freeze, and decreased to 54% post-thaw. In conclusion, epididymal spermatozoa from the white rhino may retain its functionality after cryopreservation in a commerically available cryo-extender (Bidadyl). The use of assisted reproduction techniques could someday play a role in the management and conservation of the white rhinoceros and related species.

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Genetic Recombination in Coprinus

Journal of Cell Science ◽

10.1242/jcs.6.3.669 ◽

1970 ◽

Vol 6 (3) ◽

pp. 669-678

Author(s):

B. C. LU

Keyword(s):

Fruiting Body ◽

Temperature Effects ◽

Cold Shock ◽

Genetic Recombination ◽

Recombination Frequency ◽

Temperature Treatment ◽

Sensitive Period ◽

Crossing Over ◽

Synaptinemal Complex ◽

Before And After

The frequency of genetic recombination in Coprinus lagopus may be modified by heat and cold shock. By removal of samples from a fruiting body before and after temperature treatment, it is possible to study the ultrastructure of chromosomes at the time recombination frequency (between den+ x +me-1) can be modified. The sensitive period for temperature effects and, therefore, probably the time of crossing over, commences with the formation of the synaptinemal complex (S.C.) and ends with its disappearance, i.e. during the entire existence of the S.C. It is concluded that recombination is an event subsequent to the formation of the S.C. and is independent of the process of its formation. It is suggested that the event takes place at the synaptic centre.

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RELEASE OF ULTRAVIOLET-ABSORBING MATERIAL FROM ESCHERICHIA COLI AT SUBZERO TEMPERATURES

Canadian Journal of Microbiology ◽

10.1139/m63-067 ◽

1963 ◽

Vol 9 (4) ◽

pp. 523-530 ◽

Cited By ~ 13

Author(s):

Gösta Lindeberg ◽

Aaslaug Lode

Keyword(s):

Escherichia Coli ◽

Nucleic Acids ◽

Cold Shock ◽

Organic Phosphorus ◽

Lethal Effect ◽

Rapid Cooling ◽

Pronounced Decrease ◽

Subzero Temperatures ◽

Absorbing Material ◽

Freezing Seawater

When cells of Escherichia coli were suspended in dilute artificial seawater and cooled to various subzero temperatures, a maximum lethal effect occurred around −40 °C. In addition, rapid cooling to −26 °C of bacteria, suspended in concentrated, non-freezing seawater caused a pronounced decrease in viability ("cold shock"). The loss in viability was accompanied by a proportional release from the cells of ultraviolet-absorbing material and by an increase in the ribose and organic phosphorus contents of the suspending liquid. It seems possible that the released material, at least partly, consisted of nucleotides or nucleic acids.

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Egg Yolk Antioxidants Profiles: Effect of Diet Supplementation with Linseeds and Tomato-Red Pepper Mixture before and after Storage

Foods ◽

10.3390/foods8080320 ◽

2019 ◽

Vol 8 (8) ◽

pp. 320 ◽

Cited By ~ 3

Author(s):

Besma Omri ◽

Nadir Alloui ◽

Alessandra Durazzo ◽

Massimo Lucarini ◽

Alessandra Aiello ◽

...

Keyword(s):

Antioxidant Activity ◽

Oxidative Stability ◽

Control Diet ◽

Oxidation Stability ◽

Dietary Treatment ◽

Egg Yolk ◽

Red Pepper ◽

Standard Diet ◽

Tomato Paste ◽

Before And After

This study evaluated the effect of dietary incorporation of linseed alone or along with dried tomato paste-pepper powder mix on egg physical characteristics, antioxidant profiles, lipid oxidative status, and yolk coloration before and after storage at 4 °C for one month. Sixty Novogen White laying hens, 27 weeks-old, were divided into three groups and given 100 g/hen/day of a standard diet (C), standard diet containing 4.5% of ground linseed (L), linseed diet containing 1% of dried tomato paste and 1% of sweet red pepper (LTP). Linseeds increased (p < 0.05) egg yolk antioxidant capacity but not lipid oxidative stability (p > 0.05). However, dietary inclusion of LTP did not improve fresh egg yolk antioxidant activity and lipid oxidation stability (p > 0.05). With reference to the stored eggs, only antioxidant activity measured by phosphomolybdenum reduction and lipid oxidative stability were influenced (p < 0.05) by the dietary treatment. Fresh egg yolk of hens fed on linseeds tended to have a slightly more yellow, redder, and less light color than the eggs of hens fed with the control diet. Dietary supplementation of LTP increased (p < 0.05) the Roche yolk color fan (RYCF) score and redness (a*) and decreased (p < 0.05) lightness (L*) without affecting (p > 0.05) saturation (C*). Storage of hens’ eggs fed on the control diet did not influence (p > 0.05) yolk color.

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The Prevention of Temperature Shock of Bull and Ram Semen

Australian Journal of Biological Sciences ◽

10.1071/bi9540573 ◽

1954 ◽

Vol 7 (4) ◽

pp. 573 ◽

Cited By ~ 34

Author(s):

AW Blackshaw

Keyword(s):

Cold Shock ◽

Egg Yolk ◽

Temperature Shock

The decreased vitality of ram and bull spermatozoa caused by sudden cooling (cold shock) may be largely prevented by the presence of egg yolk in the dlluent.

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Freezing buck semen diluted in amine-organic buffers

Animal Science ◽

10.1017/s0003356100006437 ◽

1993 ◽

Vol 56 (3) ◽

pp. 387-391

Author(s):

S. Dunner

Keyword(s):

Cold Shock ◽

Egg Yolk ◽

Freezing And Thawing ◽

Freezing Procedure

AbstractAmine-organic buffers (BESKOH and TESTRIS) were compared for their ability to be used for buck semen dilution in a freezing procedure. BESKOH showed better results (P < 0·05) at the time of dilution and after chilling at +5°C (72% motility v. 54% at dilution and 62% v. 39% after chilling). After freezing and thawing, none of the variables measured (motility, normal acrosome ridge and normal swelling) was significantly different. A washing procedure was necessary when freezing was undertaken and egg yolk addition was necessary to avoid cold shock as the temperature lowered during chilling. There were no significant differences between a 3% and 12% level of egg yolk addition.

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Efektivitas Suplementasi Filtrat Jambu Biji dalam Pengencer AirKelapa-Kuning Telur terhadap Kualitas Semen Cair Sapi Bali (THE EFFECTIVENESS OF GUAVA FILTRATE SUPPLEMENTATION IN COCONUT WATER-EGG YOLK DILUTION ON QUALITY OF LIQUID SEMEN OF BALI CATTLE)

jurnal veteriner ◽

10.19087/jveteriner.2019.20.1.20 ◽

2019 ◽

Vol 20 (1) ◽

pp. 20 ◽

Cited By ~ 1

Author(s):

ALoysius Marawali ◽

Muhammad S. Abdullah ◽

Jalaludin Jalaludin

Keyword(s):

Statistical Analysis ◽

Cold Shock ◽

Egg Yolk ◽

Coconut Water ◽

The Third ◽

Bali Cattle ◽

Artificial Vagina

The aim of this research was to know the effectiveness of guava filtrate supplementation in coconut water- egg yolk dilution on quality of liquid semen stored at 5oC of Bali cattle. Semen collected from a five year old Bali cattle using artificial vagina. Semen of good quality were kept in six tubes based on treatment then stored at 5oC. Treatments of the research were P0 : coconut water 80% + egg yolk 20% without guava filtrate; P1 : coconut water 80% + egg yolk 20% + 0.8% guava filtrate; P2 : coconut water 80% + egg yolk 20% + 0.9% guava filtrate; P3 : coconut water 80% + egg yolk 20% + 1.0 % guava filtrate; P4 : coconut water 80% + egg yolk 20% + 1.1 % guava filtrate and P5 : coconut water 80% + egg yolk 20% + 1.2 % guava filtrate. Each treatment was replicated 8 times making 48 experimental units. Results of the study showed that percentage mean of motility, viability, MPU, and TAU of spermatozoa after three days storage for P0 were : 42.20%, 41.85%, 39.08% and 40.90%; P1 : 50.40%, 53.89%, 52.99% and 54.67%; P2 : 54.67%, 56.97%, 54.51% and 54.36%; P3 : 17.00%, 29.96%, 29.64% and 29.64%; P4 : 23.38%, 24.64%, 21.06% and 24.45%Jurnal Veteriner Maret 2019 Vol. 20 No. 1 : 20 -29 pISSN: 1411-8327; eISSN: 2477-5665 DOI: 10.19087/jveteriner.2019.20.1.20 Terakreditasi Nasional, Dirjen Penguatan Riset dan Pengembangan, online pada http://ojs.unud.ac.id/index.php/jvet Kemenristek Dikti RI S.K. No. 36a/E/KPT/201621PENDAHULUAN Salah satu solusi yang dapat digunakan untuk pengembangan program Inseminasi buatan (IB) secara cepat dan mudah pada sapi bali adalah penggunaan semen cair. Penggunaan semen cair dapat meningkatkan kinerja IB pada sapi bali di Nusa Tenggara Timur (NTT). Keunggulan lain semen cair dapat diproduksi menggunakan bahan pengencer herbal berbasis bahan lokal dan peralatan yang sederhana serta mudah diperoleh dan tidak tergantung dengan persediaan nitrogen cair. Hasil akhir dari metabolisme spermatozoa adalah terbentuknya radikal bebas berupa derivat oksigen di antaranya adalah single1 oksigen (1O2), tripel1 oksigen (3O2), superokside anion (O2-), hidroksil radikal (OH) dan nitrit oxide (NO-) yang semuanya disebut radical oksigen species (ROS). Single1 oksigen dapat merusak ikatan rangkap pada asam lemak sehingga dapat menyebabkan kerusakan Deoxyribo Nuclead Acid (DNA) dan protein. Single1 oksigen bila bereaksi dengan asam amino histidin akan membentuk enzim yang dapat menyebabkan denaturasi protein. Kerusakan spermatozoa pada penyimpanan suhu 5%C akibat radikal bebas dan cold shock inilah merupakan penyebab utama disfungsi semen (Sharma et al., 2000). Oksidasi fosforilasi yang terganggu menyebabkan peningkatan radikal bebas dalam semen. Kadar radikal bebas yang terganggu menyebabkan peningkatan radikal bebas dalam semen. Kadar radikal bebas yang tinggi dalam sel dapat mengoksidasi lipid, protein dan DNA. Lipid membran plasma semen memiliki fosfolipid dengan kadar yang tinggi menyebabkan semen rentan terhadap radikal bebas (Sanoeka dan Kurpisz, 2004). Antioksidan bertindak mengikat asam lemak tak jenuh dan mencegah terjadinyareaksi berantai. Pada proses penyimpanan semen akan terjadi kerusakan membran plasma spermatozoa akibat terbentuknya perioksidasi lipid. Antioksidan-pemutus rantai seperti yang terkandung dalam jambu biji dapat menghambat perioksidasi lipid dalam membran melalui radical peroxyl (RO) dan alkoxyl (ROO) pengurai. Pengunaan jambu biji yang difilter dalam pengencer air kelapa kuning telur dapat menjaga kualitas spermatozoa (motilitas, keutuhan akrosom, viabilitas dan morfologi spermatozoa) semen cair sapi bali selama penyimpanan pada suhu 5%C. Dosis jambu biji yang difilter yang terbaik dalam pengencer air kelapa kuning telur, akan terbaik pula dalam mempertahankan kualitas spermatozoa sampai tujuan IB. Adapun tujuan penelitian ini adalah menguji berbagai level pemberian filtrat jambu biji (FJB) dalam pengencer air kelapa kuning telur terhadap motilitas, viabilitas, membran plasma utuh (MPU) dan tudung akrosom utuh (TAU) spermatozoa sapi bali yang disimpan pada suhu 5%C.METODE PENELITIAN Penelitian ini telah dilakukan di Laboratorium Reproduksi milik Yayasan Wiliams dan Laura yang berlokasi di Tilong, Desa Oelnasi, Kec. Kupang Tengah, Kab. Kupang, Nusa Tenggara Timur, dan berlangsung selama delapan bulan. Materi yang digunakan dalam penelitian ini adalah semen sapi bali yang ditampung dari satu ekor sapi bali jantan berumur lima tahun milik Yayasan Williams dan Laura yang telah dilatih, memiliki performans yang baik, dan organ reproduksi normal. Pakan yang diberikan adalah hijauan berupa rumput dan legum dan pemberian konsentrat secukupnya (dedak padi dan jagung giling).and P5 : 9%, 21.25%, 17.56% and 19.30%. Result of statistical analysis showed that there were a significant effect (P<0.05) between treatment on motility, viability, MPU and TAU of spermatozoa of Bali cattle till the third day of storage. It can be concluded that the supplementation of guava filtrate 0.9% in dilution of coconut water 80% - egg yolk 20% had been able to maintain motility, viability, MPU and TAU of spermatozoa of Bali cattle till the third day of storage at 5oC.

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Effects of Different Molting Procedures on Incidence of Salmonella Infection in Flocks of Naturally Contaminated Laying Hens in a Commercial Egg-Producing Farm by Detection of Yolk Antibodies to Salmonella in Eggs

Journal of Food Protection ◽

10.4315/0362-028x-69.12.2883 ◽

2006 ◽

Vol 69 (12) ◽

pp. 2883-2888 ◽

Cited By ~ 4

Author(s):

TOSHIYUKI MURASE ◽

KAORI CHIBA ◽

TOMOKO SATO ◽

KOICHI OTSUKI ◽

PETER S. HOLT

Keyword(s):

Wheat Bran ◽

Egg Production ◽

Egg Yolk ◽

The Other ◽

Feed Withdrawal ◽

Two Samples ◽

Significant Difference ◽

Before And After ◽

Specific Pathogen ◽

Egg Yolks

Indirect enzyme-linked immunosorbent assays (ELISAs) have been applied to detect immunoglobulin Y antibodies to different serotypes of Salmonella in the yolks of chicken eggs with heat-extracted antigens of Salmonella enterica serotypes Agona (SA), Cerro (SC), Enteritidis (SE), Montevideo (SM), and Putten (SP). The egg yolk samples examined were classified as positive if their ELISA absorbance values exceeded the value for eggs from specific-pathogen-free flocks by more than two standard deviations. Of 30 egg yolk samples from three flocks vaccinated with a killed SE vaccine, 29 were antibody positive by the ELISA assay for the SE antigen. Four to 29 of the 29 yolk samples showed positive results for the other serovars, although the absorbance values for SE were higher than those obtained for the other serotypes in each of the yolk samples. All 30 yolks from three flocks that were not administered any SE vaccines were found to be antibody negative for SE, and two samples were determined to be positive for SC. Thirty-nine or 40 eggs were obtained from each of four layer flocks in a commercial egg production farm where the laying houses were naturally contaminated with SA, SC, SM, SP, Salmonella serovar Infantis (SI), and untypeable strains. The ELISA absorbance values for SM in the egg yolks obtained from the two flocks molted through feed withdrawal when the birds restarted laying were significantly (P < 0.05) higher than those observed in the yolks obtained before the molt. In egg yolks from the two other flocks that were molted through a wheat bran diet, there was no significant difference between the absorbance values before and after the molt. The observations in the present study provide further evidence to suggest that a molt initiated through the administration of a wheat bran diet can reduce the risk for Salmonella problems in a commercial egg-producing setting.

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