scholarly journals Development of a sperm cryopreservation protocol of Gangetic Mystus (Mystus cavasius)

2017 ◽  
Vol 27 (4) ◽  
pp. 517-529
Author(s):  
MM Islam Islam ◽  
MN Noor ◽  
AA Islam ◽  
MRI Sarder ◽  
MZ Islam ◽  
...  
Keyword(s):  
Dimethyl Sulfoxide ◽  
Sperm Motility ◽  
Genetic Material ◽  
Nacl Solution ◽  
Sperm Cryopreservation ◽  
Endangered Fish ◽  
Long Term Preservation ◽  
Cryopreservation Protocol ◽  
Near Threatened

Cryopreservation is considered as one of the most useful techniques for long-term preservation of genetic material specially sperm of fish. This study focused on the development of a sperm cryopreservation protocol for indigenous near threatened gulsha (Mystus cavasius) and a number of experiments were conducted for the purpose. To collect milt, male gulsha were sacrificed and milt was suspended in extenders. Different concentrations of NaCl were used to evaluate the activation of sperm motility and it decreased as the concentration of the extending media increased, therefore, motility was completely inhibited at 0.8% and 1.2% NaCl solution when sperm suspended in Kurokura-2 and Alsever’s solution, respectively. The toxicity of cryoprotectants to sperm were evaluated using two cryoprotectants, dimethyl sulfoxide (DMSO) and methanol along with the extenders, Alsever’s solution and Kurokura-2 solution. DMSO and methanol with 5% and 10% concentrations produced significantly higher motility during 5 and 10 min incubation and their 15% concentration found toxic to sperm. Alsever’s solution with 10% DMSO produced best equilibration (83.75±2.39%) as well as post-thaw motility (67.5±3.23%) while Kurokura-2 solution with DMSO produced similar equilibration motility (81.25±2.39%) but the post-thaw motility (50.0±6.12%) was significantly much lower than that of Alsever’s solution. Sperm preserved with Alsever’s solution plus DMSO produced highest fertilization, 72.5±7.5% and hatching, 56.8±5.6% while fresh sperm yielded 85.0±5.0% and 74.8±3.6% fertilization and hatching, respectively. The protocols that have been developed can be used for conservation of genetic materials of M. cavasius and other endangered fish species and new generations of them can be propagated using the cryopreserved sperm.Progressive Agriculture 27 (4): 517-529, 2016

scholarly journals Acrosomal integrity and capacitation are not influenced by sperm cryopreservation in the giant panda

Reproduction ◽  
2004 ◽  
Vol 127 (5) ◽  
pp. 547-556 ◽  
Author(s):  
R E Spindler ◽  
Y Huang ◽  
J G Howard ◽  
P Wang ◽  
H Zhang ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Giant Panda ◽  
Calcium Ionophore ◽  
Sperm Cryopreservation ◽  
Management Tools ◽  
Giant Pandas ◽  
Before And After ◽  
Cryopreservation Protocol ◽  
Acrosomal Integrity

Sperm cryopreservation and artificial insemination are important management tools for giant panda breeding and the preservation of extant genetic diversity. This study examined the influence of freeze–thawing on sperm function, specifically capacitation. Sperm from nine giant pandas were assessed before and after rapid (− 40 and − 100 °C/min) cryopreservation by incubation in HEPES-buffered Ham’s F10 medium with and without the capacitation accelerators, 3-isobutyl-1-methylxanthine (IBMX) and dibutyryl cyclic AMP (dbcAMP). At 0, 3 and 6 h of exposure, aliquots were assessed for sperm motility traits and capacitation, defined as the proportion of sperm with intact acrosomes following exposure to solubilised zonae pellucidae (ursid or felid) or calcium ionophore subtracted from the proportion of sperm with intact acrosomes before exposure. Although mean±s.e.m. sperm motility post-thaw (56.1 ± 3.9% at 0 h) was less (P < 0.05) than pre-freeze (71.7 ± 6.0%), there was no difference (P > 0.05) in the proportion of acrosome-intact sperm (fresh, 93.0 ± 1.7% versus cryopreserved–thawed, 81.7 ± 4.7% at 0 h). Incidence of capacitation was greater (P < 0.05) in fresh sperm incubated with capacitation accelerators IBMX and dbcAMP (9 h: 50.9 ± 1.1) compared with fresh sperm incubated without accelerators (9 h: 41.2 ± 1.1%). Frozen–thawed sperm preincubated without accelerators underwent capacitation (49.6 ± 1.1%) to a greater extent (P < 0.05) compared with these fresh counterparts. Thawed samples with (9 h: 45.9 ± 1.4%) and without accelerators (9 h: 41.2 ± 1.1%) did not differ (P > 0.05) during the 9-h incubation. We conclude that giant panda spermatozoa (1) undergo capacitation in vitro with or without chemical accelerators and (2) withstand a rapid cryopreservation protocol, including retaining normal acrosomal integrity and functional capacitation ability.

scholarly journals Cryopreservation and transplantation of common carp spermatogonia

2018 ◽  
Author(s):  
Roman Franěk ◽  
Zoran Marinović ◽  
Jelena Lujić ◽  
Béla Urbányi ◽  
Michaela Fučíková ◽  
...  
Keyword(s):  
Common Carp ◽  
Sperm Cryopreservation ◽  
Post Transplantation ◽  
Wide Range ◽  
Cultured Fish ◽  
The World ◽  
Exogenous Origin ◽  
Long Term Preservation ◽  
Surrogate Production

AbstractCommon carp (Cyprinus carpio) is one of the most cultured fish species over the world with many different breeds and plenty of published protocols for sperm cryopreservation, however, data regarding preservation of gonadal tissue and surrogate production is still missing. A protocol for freezing common carp spermatogonia was developed through varying different factors along a set of serial subsequent experiments. Among the six cryoprotectants tested, the best survival was achieved with dimethyl sulfoxide (Me2SO). In the next experiment, a wide range of cooling rates (0.5–10 °C/min) and different concentrations of Me2SO were tested resulting in the highest survival using 2 M Me2SO and cooling rate of –1 Q59 A When testing different tissue sizes and incubation times in the cryomedium, the highest viability was observed when incubating 100 mg tissue fragments for 30 min. Finally, sugar supplementation did not yield significant differences. When testing different equilibration (ES) and vitrification solutions (VS) used for needle-immersed vitrification, no significant differences were observed between the tested groups. Additionally, varied exposure time to VS did not improve the vitrification outcome where the viability was 4-fold lower than that of freezing. The functionality of cryopreserved cells was tested by interspecific transplantation into sterilized goldfish recipients. The exogenous origin of the gonads in goldfish recipients was confirmed by molecular markers and incorporation rate was over 40% in both groups at 3 months post transplantation. Results of this study can serve as an alternative way for long-term preservation of germplasm in carp which can be recovered in a surrogate recipient.

Effect of cooling and cryopreservation on sperm motility and morphology of several species of marsupial

1996 ◽  
Vol 8 (4) ◽  
pp. 673 ◽  
Author(s):  
DA Taggart ◽  
CM Leigh ◽  
VR Steele ◽  
WG Breed ◽  
PD Temple-Smith ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Cold Shock ◽  
Egg Yolk ◽  
The Other ◽  
Brushtail Possum ◽  
Motile Sperm ◽  
Sperm Cryopreservation ◽  
Marsupial Species ◽  
Live Sperm

The effects of long-term cooling and freezing on sperm motility are described for six marsupial species: the fat-tailed dunnart, koala, brushtail possum, long-footed potoroo, northern brown bandicoot and ring-tailed possum. The effects of up to eight days of cooling at 4 degrees C on the motility of dunnart spermatozoa and the effect of cryopreservation on spermatozoa of the other species were determined. The cryoprotectant used was a Tris-citrate-fructose-egg yolk-glycerol diluent. The percentage and rating of sperm motility, and sperm structure, as determined by light microscopy, were investigated. Sperm motility in the fat-tailed dunnart was retained for up to six days when cooled to 4 degrees C, suggesting that sperm from this species have some degree of tolerance to cold shock. After this time, however, the percentage of motile spermatozoa and their motility rating declined. In all species except the fat-tailed dunnart, reinitiation of motility following cryopreservation occurred across a range of glycerol concentrations (4-17%). Cryoprotectant containing 6% and/or 8% glycerol resulted in little change of motility rating or of the percentage of live sperm after thawing, although there was some decline in the percentage of motile sperm. The unusual structural and motility characteristics of dunnart spermatozoa may account for the lack of success of sperm cryopreservation in this species.

scholarly journals Kriopreservasi Tanaman Obat Langka Purwoceng dengan Teknik Enkapsulasi-Vitrifikasi

2016 ◽  
Vol 14 (2) ◽  
pp. 49
Author(s):  
Ika Roostika ◽  
Suci Rahayu ◽  
Novianti Sunarlim
Keyword(s):  
Liquid Nitrogen ◽  
Incubation Period ◽  
Optimization Method ◽  
Endangered Medicinal Plant ◽  
In Vitro Shoots ◽  
Long Term Preservation ◽  
Cryopreservation Protocol ◽  
Cacl2 Solution

<p>Pruatjan (Pimpinella pruatjan Molk.) is an Indonesian endangered medicinal plant, so that it is highly protected. Cryopreservation can be applied to this plant for long-term preservation. The aim of this research was to obtain a method of encapsulation-vitrification by optimizing each step in cryopreservation protocol i.e. preculture, loading, dehydration with and without freezing in liquid nitrogen. The best treatment of each step would be applied in the following step. On preculture experiment, in vitro shoots were planted on the Driver and Kuniyaki (DKW) basal media containing 0.3 M sucrose and incubated for 1, 2, 3, 4, and 5 days. After those incubation period, shoot tips were encapsulated with 2.5% Na-alginate and soaking for 15 minutes in 100 ppm CaCl2 solution before planting. On loading experiment, precultured explants were loaded in DKW basal solution containing 2 M glycerol and 0.4 M sucrose for 0, 30, 60, and 90 minutes. On dehydration experiment, preculturead and loaded explants were dehydrated with PVS2 solution PVS2 (DKW + 30% glycerol + 15% DMSO + 15% ethyleneglicol + 0.4 M sucrose) for 0, 30, 60, 90, and 120 minutes. The parts of them were freezed in liquid nitrogen (-196oC). The result showed that cryopreservation through encapsulation-vitrification technique could be applied on pruatjan. The best preculture treatment was 5 days incubation period. The best loading treatment was 30 minutes. The best dehydration treatment was 90 minutes. The successful level of this research was still low (10%) so that it needs optimization method.</p><p> </p><p><strong>Abstrak</strong></p><p>Purwoceng (Pimpinella pruatjan Molk.) adalah tanaman obat langka asli Indonesia yang hampir punah sehingga harus dilindungi. Kriopreservasi dapat diterapkan pada tanaman ini untuk penyimpanan jangka panjang. Tujuan penelitian adalah untuk memperoleh teknik enkapsulasi-vitrifikasi dengan melakukan optimasi dari tiap-tiap tahapan kriopreservasi yang meliputi perlakuan prakultur, loading, dehidrasi sebelum dan setelah pembekuan dalam nitrogen cair. Perlakuan yang terbaik kemudian diterapkan pada tahapan percobaan berikutnya. Pada perlakuan prakultur, tunas in vitro ditanam pada media Driver dan Kuniyaki (DKW) dengan penambahan sukrosa 0,3 M dengan masa inkubasi 1, 2, 3, 4, dan 5 hari. Setelah itu, pucuk yang berukuran 0,5 cm dienkapsulasi dengan Na-alginat 2,5% (yang mengandung media regenerasi) dalam larutan CaCl2 100 ppm selama 15 menit sebelum penanaman kembali. Pada percobaan loading, terlebih dahulu eksplan diprakultur kemudian direndam dalam larutan DKW + gliserol 2 M + sukrosa 0,4 M dengan durasi rendam selama 0, 30, 60, dan 90 menit. Pada percobaan dehidrasi, eksplan diprakultur dan loading terlebih dahulu, kemudian direndam dalam larutan krioprotektan PVS2 (DKW + gliserol 30% + DMSO 15% + etilen glikol 15% + sukrosa 0,4 M ) selama 0, 30, 60, 90, dan 120 menit. Eksplan tersebut sebagian dibekukan dalam nitrogen cair (-196oC) dan sebagian lainnya tidak dibekukan. Hasil penelitian menunjukkan bahwa kriopreservasi secara enkapsulasi-vitrifikasi berpeluang diterapkan pada tanaman purwoceng. Perlakuan prakultur terbaik adalah 5 hari. Perlakuan loading terbaik adalah 30 menit dan perlakuan dehidrasi terbaik 90 menit. Tingkat keberhasilan ini masih rendah (10%) sehingga diperlukan optimasi metode.</p>

scholarly journals Cryoprotectant treatment tests on three morphologically diverse marine dinoflagellates and the cryopreservation of Breviolum sp. (Symbiodiniaceae)

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Joseph Kanyi Kihika ◽  
Susanna A. Wood ◽  
Lesley Rhodes ◽  
Kirsty F. Smith ◽  
Lucy Thompson ◽  
...  
Keyword(s):  
Genetic Drift ◽  
Rapid Freezing ◽  
Diverse Group ◽  
Limited Success ◽  
The Family ◽  
Agricultural Applications ◽  
Marine Dinoflagellates ◽  
Long Term Preservation ◽  
Cryopreservation Protocol

AbstractDinoflagellates are among the most diverse group of microalgae. Many dinoflagellate species have been isolated and cultured, and these are used for scientific, industrial, pharmaceutical, and agricultural applications. Maintaining cultures is time-consuming, expensive, and there is a risk of contamination or genetic drift. Cryopreservation offers an efficient means for their long-term preservation. Cryopreservation of larger dinoflagellate species is challenging and to date there has been only limited success. In this study, we explored the effect of cryoprotectant agents (CPAs) and freezing methods on three species: Vulcanodinium rugosum, Alexandrium pacificum and Breviolum sp. A total of 12 CPAs were assessed at concentrations between 5 and 15%, as well as in combination with dimethyl sulfoxide (DMSO) and other non-penetrating CPAs. Two freezing techniques were employed: rapid freezing and controlled-rate freezing. Breviolum sp. was successfully cryopreserved using 15% DMSO. Despite exploring different CPAs and optimizing the freezing techniques, we were unable to successfully cryopreserve V. rugosum and A. pacificum. For Breviolum sp. there was higher cell viability (45.4 ± 2.2%) when using the controlled-rate freezing compared to the rapid freezing technique (10.0 ± 2.8%). This optimized cryopreservation protocol will be of benefit for the cryopreservation of other species from the family Symbiodiniaceae.

scholarly journals Droplet-vitrification methods for apical bud cryopreservation of yacon [Smallanthus sonchifolius (Poepp. and Endl.) H. Rob.]

2021 ◽  
Author(s):  
Stacy Denise Hammond Hammond ◽  
Iva Viehmannova ◽  
Jiri Zamecnik ◽  
Bart Panis ◽  
Milos Faltus
Keyword(s):  
Osmotic Dehydration ◽  
Survival Rates ◽  
Ms Medium ◽  
Smallanthus Sonchifolius ◽  
Apical Bud ◽  
Apical Buds ◽  
Tuberous Roots ◽  
Long Term Preservation ◽  
Cryopreservation Protocol

Abstract This study aimed to develop a cryopreservation protocol for the long-term preservation of yacon [Smallanthus sonchifolius (Poepp. and Endl.)], an Andean crop with high fructooligosaccharide content in its tuberous roots. Initially, the cryopreservation protocol was developed using a yacon clone originated from Ecuador classified as ECU 41. Osmotic dehydration of apical buds (2–3 mm long) was carried out by assessing two plant vitrification solutions, PVS2 (15, 30, and 60 min) at 0°C and PVS3 (30, 45, 60, and 75 min) at 22°C. After cryopreservation, the apical buds were thawed and placed on MS medium ± 0.1 mg l− 1 N6-benzyladenine (BA). The survival rates ranged from 37 to 90% within all treatments, with those subjected to PVS2 and PVS3 for 60 min showing the highest survival rates on MS medium without BA (87 and 90%, respectively). At 12 weeks post cryopreservation, these treatments also provided the highest regrowth rates, both reaching 73% of normally growing (shooting, rooting) plantlets. Survival rates on MS + 0.1 mg l− 1 BA regrowth medium reached up to 90%; however, regrowth into normally rooted plantlets did not exceed 67% post cryopreservation. The optimized protocols were then applied to 4 additional yacon clones originated from Bolivia and Peru, classified as BOL 22, BOL 23, PER 12, and PER 14. This resulted in survival and regeneration rates ranging between 79.7–94.1% and 66.3–75.4% respectively. Our study shows that optimal cryopreservation protocols for the long-term conservation of yacon can be based on both PVS2 and PVS3 vitrification solutions.

scholarly journals Effects of skim-milk supplementation on the quality and penetrating ability of boar semen after long-term preservation at 15 °C

2014 ◽  
Vol 62 (1) ◽  
pp. 106-116 ◽  
Author(s):  
Zhao Namula ◽  
Risa Kodama ◽  
Fuminori Tanihara ◽  
Yasuhiro Morita ◽  
Yoko Sato ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Milk Powder ◽  
Skim Milk ◽  
Skim Milk Powder ◽  
Control Group ◽  
Fertilisation Rate ◽  
Boar Semen ◽  
Long Term Preservation

This study investigated the effects of skim-milk supplementation on the quality and penetrating ability of boar semen preserved at 15 °C. When boar semen samples were preserved in Modified Modena extender supplemented with various concentrations (0, 7.5, 15, 30 and 50 mg/mL) of skim milk powder at 15 °C for 4 weeks, higher sperm motility and viability were observed in the case of 7.5 mg/mL skim-milk supplementation compared with the control group (0 mg/mL) during the preservation (P < 0.05). When in vitro matured oocytes were co-incubated with boar sperm that had been preserved in Modified Modena extender with three different concentrations (0, 7.5 or 15 mg/mL) of skim milk powder at 15 °C for two weeks, there were no apparent effects of skim-milk supplementation on the rates of fertilisation and development to blastocysts of oocytes after co-incubation. However, the monospermic fertilisation rate of sperm preserved with 15 mg/mL skim milk powder was higher (P < 0.05) than that of fresh non-preserved sperm, but did not differ among the preservation groups. The results indicate that the supplementation of Modified Modena extender with 7.5 mg/mL skim milk powder improves the motility and viability, but not the penetrating ability, of sperm after liquid preservation for at least two weeks.

Reactivity of Pulmonary Alveolar Cells to Crocidolite Asbestos Fibers

1982 ◽  
Vol 40 ◽  
pp. 334-335
Author(s):  
Charles L. Sanders ◽  
Roy R. Adee
Keyword(s):  
Osmium Tetroxide ◽  
Intratracheal Instillation ◽  
Uranyl Acetate ◽  
Fiber Suspension ◽  
Nacl Solution ◽  
Alveolar Cells ◽  
Asbestos Fibers ◽  
Fischer Rats ◽  
Mineral Silicates

Asbestos is a generic name for a group of hydrated mineral silicates that occur naturally in a fibrous form. The early interactions of asbestos fibers with alveolar cells in large part determines their long-term toxicity. Young adult, SPF, Fischer rats were given a single intratracheal instillation of 2 mg crocidolite asbestos suspended in 0.5 ml of 0.9% NaCl solution. About 80% of the fibers had lengths of less than 10 ym as measured on light micrographs of the fiber suspension. Two rats were killed at 3 hr, 1 d and 1, 4, 8, 12 and 16 wk after instillation and the lungs instilled with 8 ml McDowell - Trumps at 20 cm H2O. Lung tissue was dehydrated and sputtered coated with palladium-gold for SEM or post-fixed in osmium tetroxide, embedded in epoxy resin and sections stained with uranyl acetate and lead citrate for TEM.

Problems of treatment of primary openangle glaucoma in combination with thrombosis of the central retinal vein

GlaucomaNews ◽  
2020 ◽  
pp. 65-69
Author(s):  
T.E. Lipatkina ◽  
◽  
Е.V. Karlova ◽  
A.V. Zolotarev ◽  
◽  
...  
Keyword(s):  
Macular Edema ◽  
Primary Open Angle Glaucoma ◽  
Antihypertensive Drugs ◽  
Open Angle Glaucoma ◽  
Vascular Wall ◽  
Open Angle ◽  
Retinal Edema ◽  
Long Term Preservation ◽  
Central Retinal Vein

Patients with primary open-angle glaucoma (POAG) and ophthalmic hypertension have an increased likelihood of developing occlusions (thrombosis) of the central retinal vein. Different groups of antihypertensive drugs differ in their mechanism of action and may affect concomitant ocular pathology, in particular, retinal edema, which occurs, for example, in occlusion of the central retinal vein. Used in most patients with glaucoma, prostaglandin analogs can contribute to the long-term preservation of macular edema due to the effect on the permeability of the vascular wall. Preparations of other pharmacological groups, reducing the production of aqueous humor, on the contrary, may contribute to its regression. Therefore, the question of choosing a drug for antihypertensive therapy in patients with primary open-angle glaucoma and concomitant macular edema is relevant and is for further study.

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