Effective freezing rate for semen cryopreservation in endangered Mediterranean brown trout (Salmo trutta macrostigma) inhabiting the Biferno river (South Italy)

Zygote ◽  
2015 ◽  
Vol 24 (5) ◽  
pp. 668-675 ◽  
Author(s):  
Nicolaia Iaffaldano ◽  
Michele Di Iorio ◽  
Angelo Manchisi ◽  
Stefano Esposito ◽  
Pier Paolo Gibertoni
Keyword(s):  
Liquid Nitrogen ◽  
Brown Trout ◽  
Salmo Trutta ◽  
Semen Quality ◽  
Egg Yolk ◽  
Freezing Rate ◽  
Frozen Semen ◽  
Fresh Semen

SummaryThis study was designed to determine: (i) the in vitro effects of different freezing rates on post-thaw semen quality of Mediterranean brown trout (Salmo trutta macrostigma) from the Biferno river; and (ii) the in vivo fertilization and hatching percentage of freezing rate giving rise to the best post-thaw semen quality.Pooled semen samples were diluted 1:3 (v:v) in a freezing extender composed of 300 mM glucose, 10% egg yolk and 10% dimethyl sulfoxide (DMSO). The extended semen was packaged in 0.25 ml plastic straws and frozen at different heights above the liquid nitrogen surface (1, 5 or 10 cm) for 10 min to give three different freezing rates. Semen samples were thawed at 30°C for 10 s. The variables assessed after thawing were sperm motility, duration of motility and viability.Our results clearly indicate a significant effect of freezing rate on post-thaw semen quality. Semen frozen 5 cm above the liquid nitrogen surface showed the best quality after freezing/thawing. Based on these in vitro data, 2 groups of 200 eggs were fertilized with fresh semen or semen frozen 5 cm above the liquid nitrogen surface. Fertilization and hatching rates recorded for eggs fertilized with frozen semen were significantly lower (25.4% and 22.5%, respectively) than the ones obtained using fresh semen (87.8% and 75.5%, respectively). An effective freezing protocol will allow for the creation of a sperm cryobank to recover the original population of Mediterranean brown trout in the Biferno river.

scholarly journals Optimization of Sperm Cryopreservation Protocol for Mediterranean Brown Trout: A Comparative Study of Non-Permeating Cryoprotectants and Thawing Rates In Vitro and In Vivo

Animals ◽  
2019 ◽  
Vol 9 (6) ◽  
pp. 304 ◽  
Author(s):  
Giusy Rusco ◽  
Michele Di Iorio ◽  
Pier Paolo Gibertoni ◽  
Stefano Esposito ◽  
Maurizio Penserini ◽  
...  
Keyword(s):  
Brown Trout ◽  
Semen Quality ◽  
Sperm Quality ◽  
Egg Yolk ◽  
Dna Integrity ◽  
Experimental Conditions ◽  
C 30 ◽  
Thawing Rate

The aim of our study was to test the effects of different non-permeating cryoprotectants (NP-CPAs), namely low-density lipoproteins (LDLs), sucrose, and egg yolk, and thawing rates on the post-thaw semen quality and fertilizing ability of the native Mediterranean brown trout. Pooled semen samples were diluted 1:3 (v:v) with 2.5%, 5%, 10%, or 15% LDL; 0.05, 0.1, or 0.3 M sucrose; or 10% egg yolk. At the moment of analysis, semen was thawed at 30 °C/10 s or 10 °C/30 s. The post-thaw semen quality was evaluated, considering motility, the duration of motility, viability, and DNA integrity. Significantly higher values of motility and viability were obtained using egg yolk/10 °C for 30 s, across all treatments. However, LDL and sucrose concentrations affected sperm cryosurvival, showing the highest post-thaw sperm quality at 5% LDL and 0.1 M sucrose. Based on the in vitro data, egg yolk, 5% LDL, and 0.1 M sucrose thawed at 10 °C or 30 °C were tested for the in vivo trial. The highest fertilization and hatching rates were recorded using egg yolk/10 °C (p < 0.05). According to these in vitro and in vivo results, egg yolk emerged as the most suitable NP-CPA and 10 °C/30 s as the best thawing rate for the cryopreservation of this trout sperm, under our experimental conditions.

scholarly journals A Simple and Efficient Semen Cryopreservation Method to Increase the Genetic Variability of Endangered Mediterranean Brown Trout Inhabiting Molise Rivers

Animals ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 403
Author(s):  
Giusy Rusco ◽  
Michele Di Iorio ◽  
Roberta Iampietro ◽  
Stefano Esposito ◽  
Pier Paolo Gibertoni ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Brown Trout ◽  
Sperm Concentration ◽  
Mediterranean Area ◽  
Fertilization Rate ◽  
Semen Cryopreservation ◽  
Nitrogen Vapor ◽  
C 30

The aim of our study was to test the effectiveness of a simple semen cryopreservation procedure, developed for cultivated salmonid, on the wild salmonid of the Mediterranean area and to evaluate the effect of different thawing rates and sperm-to-egg ratios. The semen of five individual males was diluted into a final extender concentration of 0.15 M glucose and 7.5% methanol and loaded into 0.25 mL plastic straws, and a final sperm concentration of 3.0 × 109 sperm/mL was obtained. After equilibration, the straws were frozen by exposure to liquid nitrogen vapor at 3 cm above the liquid nitrogen level for 5 min. The semen was thawed at 40 °C/5 s or 10 °C/30 s. The sperm cryosurvival was evaluated by examining in vitro the sperm motility parameters using the CASA system, followed by fertilization trials in vivo, using three different sperm-to-egg ratios 6 × 105, 4.5 × 105 and 3 × 105:1. The applied cryopreservation procedure resulted in remarkably high (85.6%) post-thaw sperm total motility, when the semen was thawed at 40 °C/5 s, whilst the highest fertilization rate (53.1%) was recorded for a sperm-to-egg ratio of 4.5 × 105:1. According to these outcomes, the cryopreservation procedure that was tested turned out to be effective for the wild population of Mediterranean brown trout and practical for the creation of the first European semen cryobank foreseen as part of our “LIFE” Nat.Sal.Mo. project.

scholarly journals Effect of preservation time on the quality of frozen semen in indigenous rams

2015 ◽  
Vol 44 (1) ◽  
pp. 10-15 ◽  
Author(s):  
BBA Mahmuda ◽  
Azizun Nesa ◽  
BF Zohara ◽  
MGS Alam ◽  
FY Bari
Keyword(s):  
Semen Quality ◽  
Egg Yolk ◽  
Volume Density ◽  
Normal Morphology ◽  
Mean Values ◽  
Frozen Semen ◽  
Significant Difference ◽  
The Mean ◽  
Fresh Semen

The study was carried out to observe the effects of preservation time on the quality of frozen semen of indigenous rams. Semen was collected using AV once a week from 4 rams. Tris based with 10% egg yolk and 7% glycerol extender was used to extend and freezing the semen. Fresh semen was evaluated for volume, density, mass motility and concentration, and mean values were observed as 0.8±0.2ml, 3.0±0.3, 3.2±0.7, 3.9±0.7×109/ml, respectively. Significant difference (p<0.05) was found for all the parameters among the rams. Mean values of motility, viability and normal morphology percentages were 83.3±4.3%, 88.2±4.4%, 84.2±3.5% in fresh semen while those of chilled semen at 40C were 74.7±2.3, 78.8±4.9 and 79.2±2.9%, respectively. For all the parameters, significant (p<0.05) difference was found among the rams. Frozen sperm motility was observed after thawing at 39-400C for 14-15 seconds. The mean motility, viability and normal morphology percentages after freezing for 24hrs, 7, 15 and 30 days of duration were 39.8±3.1, 41.1±4.3, 40.1±4.1 and 39.4±2.9%; 44.5±2.5, 45.3±2.8, 44.6±2.8 and 43.9±2.8%; 71.0±2.0, 71.7±1.5, 70.7±1.7 and 70.3±1.8%, respectively and values did not decrease significantly (p>0.05) with the increasing time of preservation. Non significantly decrease of the semen quality with advance of preservation time indicates the suitability of the protocol used for freezing of indigenous ram semen in Bangladesh.DOI: http://dx.doi.org/10.3329/bjas.v44i1.23113            Bang. J. Anim. Sci. 2014. 44 (1): 10-15

In vitroeffect of various cryoprotectants on the semen quality of endangered Oravka chicken

Zygote ◽  
2017 ◽  
Vol 26 (1) ◽  
pp. 33-39 ◽  
Author(s):  
Andrea Svoradová ◽  
Lenka Kuželová ◽  
Jaromír Vašíček ◽  
Andrej Baláži ◽  
Emília Hanusová ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Semen Quality ◽  
Semen Analysis ◽  
Lower Percentage ◽  
Computer Assisted ◽  
Spermatozoa Motility ◽  
Progressive Movement ◽  
Animal Genetic Resources

SummaryWe aimed to compare the effect of three different permeating cryoprotectants on the post-thaw spermatozoa quality. Pooled semen from Oravka cock line (n= 6) was diluted inKobidil+extender and frozen in cryoprotectant solutions containing 8% dimethylsulfoxide (DMSO), 8% ethylene glycol (EG) or 8% glycerol (GL) in liquid nitrogen vapours before being plunged into the liquid nitrogen. Spermatozoa motility parameters were assessedin vitroafter freezing–thawing by a computer-assisted semen analysis (CASA) system and viability status was examined using fluorescent probes. The lower percentage (P <0.05) of motile and progressively moving spermatozoa immediately after thawing were obtained in all experimental groups (DMSO, EG, GL) compared with the control. Significant (P< 0.05) differences in total motility and progressive movement between GL and DMSO, EG groups were observed. However, the higher number (P <0.05) of acrosome damaged spermatozoa was found in the DMSO and EG groups and no significant differences were observed in the GL group compared with the control. Differences (P< 0.05) between experimental groups and the control in the results of spermatozoa necrosis were observed. No significant differences in the percentage of apoptotic spermatozoa were found between control and experimental groups. However, significant differences (P< 0.05) in number of live and necrotic spermatozoa between GL and DMSO, EG groups were examined. The findings of the present study indicate that glycerol seems to be suitable for semen cryopreservation in the gene banks. In addition, fertility evaluationin vivois needed in order to evaluate the possible contribution for the bank of animal genetic resources.

Kualitas Semen Cair Kambing Boer Berbahan Pengencer Air Kelapa Muda Varietas Viridis Setelah Simpan Dingin

2019 ◽  
Vol 6 (1) ◽  
pp. 78
Author(s):  
Muhammad Ade Salim ◽  
Muhammad Nur Ihsan ◽  
Nur Isnaini ◽  
Trinil Susilawati
Keyword(s):  
Semen Quality ◽  
Block Design ◽  
Animal Husbandry ◽  
Egg Yolk ◽  
Coconut Water ◽  
Boer Goat ◽  
Randomized Block Design ◽  
Frozen Semen ◽  
Artificial Vagina

ABSTRAKAir kelapa muda varietas viridisdapat dijadikan pengencer aletrnatif semen cair bagi program IB di daerah minim sarana semen beku. Tujuan penelitian ini untuk menguji pengaruh penggunaan air kelapa muda viridissebagai bahan pengencer terhadap kualitas semen cair kambing Boer setelah didinginkan. Dilaksanakanselama 3 bulan di Laboratorium Fakultas Peternakan UBUnit SumberSekar,Malang. Metodenya yaitu eksperimen. Semen dari  3 pejantan Boer umur 3-5 tahun, dikoleksi seminggu sekali dengan VB. Air kelapa mudaviridis umur 5-7 bulan serta tris aminomethane sebagai kontrol. Didesain menggunakan Rancangan Acak Kelompok (RAK) dengan 2 perlakuan yaitu P0 (tris aminomethane + 10% KT) dan  P1 (air kelapa muda viridis + 10% KT) masing-masing diulang 10 kali. Data dianalisis dengan analisis Ragam (Anova) dengan software Genstat 18. Variabelnya yaitu motilitas individu, viabilitas dan abnormalitas. Hasil penelitian yaitu motilitas individu pada P1bertahan sampai 4 hari (40,5± 24,3%), viabilitas terbaik sampai hari ke-5 (42±24,6%), abnormalitas terendah di hari ke-7(1,31± 0,6). Kesimpulannya, Pengencer air kelapa muda viridis dapat mempertahankan kualitas semen cair kambing Boer selama 4 hari untuk motilitas dan 5 hari untuk viabilitas.Kata Kunci:pengencer, air kelapa, varietas viridisABSTRACTYoung viridis coconut water could be used as an alternative to liquid semen diluent for artificial insemination program in the area with limited facility for frozen semen production. This study evaluated the use of young coconut water as a diluent on liquid semen quality of Boer goat after cold storage. This study was carried out for 3 months at Sumber Sekar Laboratory, Faculty of Animal Husbandry, University of Brawijaya, Malang. The semen was collected from 3 Boer bucks aged at 3 to 5 years old. The semen collection was done once a week with the aid of artificial vagina. The diluents used were young Viridis coconut (5 to 7 months old) and tris aminomethane. The method used was an experiment in a randomized block design with 2 treatments and 10 replicates. The treatments used were T0: tris aminomethane + 10% egg yolk (control) and T1:  young Viridis coconut water + 10% egg yolk. Data were analyzed by analysis of variance using Genstat 18 software. The variables measured were sperm individual motility, viability, and abnormality. The results showed that the sperm individual motility in T1 survived up to 4 days (40.5± 24.3%), the best viability at 5 days (42.0±24.6%),  while the lowest abnormality at 7 days (1.31±0.6). It could be concluded that: 1. Tris aminomethane diluent has higher quality with the storage length up to 9 days, 2. Young Viridis coconut water diluent could preserve liquid semen quality of Boer goat up to 4 days for sperm motility and 5 days for sperm viability.Keywords: diluents, coconut water, viridis variety

scholarly journals Effect of Oral Administration of Selenium and Vitamin E on the Quality of Fresh, Refrigerated and Frozen Semen in French Bulldog Breed Dogs

2017 ◽  
Vol 45 (1) ◽  
pp. 7
Author(s):  
Marcelo George Mungai Chacur ◽  
Mariana Grandis Ripari de Souza ◽  
Camila Dutra de Souza ◽  
Camila Pires Cremasco
Keyword(s):  
Vitamin E ◽  
Citric Acid ◽  
Oral Administration ◽  
Liquid Nitrogen ◽  
Sperm Concentration ◽  
Oral Supplementation ◽  
Frozen Semen ◽  
French Bulldog ◽  
Fresh Semen

Background: New methodologies have been developed seeking to maximize pregnancy rate in female dogs created in commercial kennels, and also in order to maintain the quality of canine semen after dilution, refrigeration or freezing. One of the main factors that generate damage to sperm is oxidative stress, to minimize sperm damage, selenium and antioxidants like vitamin E are administered, by oral administration, seeking to improve the quality of semen. The objective was to study the effect of vitamin E and selenium, by oral administration, in the quality of fresh, refrigerated and frozen semen in adult dogs French Bulldog breed.Materials, Methods & Results: Semen samples were collected from 5 adult dogs, French Bulldog breed, being 2 semen drawing before the daily oral supplementation with vitamin E and selenium (ESE®) and semen drawing at 20, 40 and 60 days after the beginning of oral supplement. The ejaculated samples were diluted in TRIS - fructose citric acid (3.28 g TRIS-hydroxy-methyl-amino-methane, 1.78 g of citric acid monohydrate and 1.25 g of D - fructose, dissolved in 100 mL of distilled water and added of 20% egg yolk and 6% of glycerol. The characteristics evaluated in fresh semen were: volume (mL), color, appearance, concentration (x106 / mL), sperm motility (%), sperm strength (1 to 5) and morphology (%). For refrigerated and frozen semen were analyzed: sperm motility (%), sperm strength (1-5) and morphology (%). Diluted semen samples were centrifuged at: 1500 g/10 min and “pellets” formed by sperm of each ejaculated, detached from the tube wall were diluted homogeneously in the diluent TRIS type up to the final volume of 1.5 mL. After that, packaged in 0.5 mL French straws, kept under refrigeration at 5ºC/4 h, placed in nitrogen vapor at -120ºC/15 min, and dipped in liquid nitrogen at -196ºC and then stored on identified rachis and stored in liquid nitrogen container until the time of thawing in  water bath at 37°C/30 s for semen microscopic analysis. Data from fresh, refrigerated and frozen semen were statistically analyzed by analysis of variance and the average compared by 5% of Tukey test. Fresh semen sperm concentration differed (P < 0.05) between the samples, rising after 40 days after the beginning of oral supplementation with selenium and vitamin E. For the spermatic strength, better score (P < 0.05) was observed at collection 4, in 40 days after the beginning of oral supplementation to dogs. For fresh and refrigerated semen, the total defects, defects of head, acrosome and tail did not differ (P > 0.05) between the samples. Total sperm defects and minor head and tail defects did not differ (P > 0.05) between the samples in post-thawing. Regarding the acrosome defects after thawing, there was a significant reduction (P < 0.05) in samples performed 40 and 60 days after the beginning of oral supplementation with selenium and vitamin E.Discussion: Attention should be paid for what purpose the extenders within the refrigeration or freezing biotech will be used. The managed supplement, by oral administration, containing selenium and vitamin E, influenced beneficially raising the sperm concentration in fresh semen and decreasing the acrosome defects in frozen semen. Oral administration of supplementation with selenium and vitamin E is recommended for improving the quality of fresh and frozen semen in dogs.

Surface culture of Steinernema sp. on two solid media and their pathogenicity against Galleria mellonella

2021 ◽  
Vol 95 ◽  
Author(s):  
C.I. Cortés-Martínez ◽  
A.I. Rodríguez-Hernández ◽  
M.R. López-Cuellar ◽  
N. Chavarría-Hernández
Keyword(s):  
Galleria Mellonella ◽  
Egg Yolk ◽  
Infective Juvenile ◽  
Surface Culture ◽  
Solid Media ◽  
Significant Difference ◽  
Bacterial Lawn ◽  
The Mean

Abstract The use of native entomopathogenic nematodes as biocontrol agents is a strategy to decrease the environmental impact of insecticides and achieve sustainable agriculture crops. In this study, the effect of the surface culture of Steinernema sp. JAP1 over two solid media at 23–27°C on infective juvenile (IJ) production and pathogenicity against Galleria mellonella larvae were investigated. First, the bacterial lawn on the surface of the media with egg yolk (P2) or chicken liver (Cl) were incubated in darkness at 30°C for 48 and 72 h, and 100 surface-sterilized IJs were added. Four harvests were conducted within the next 35 days and the mean accumulated production was superior on Cl (210 × 103 IJs) than on P2 (135 × 103 IJs), but the productivity decreased up to 10% when the incubation time of the bacterial lawn was of 72 h. The mean pathogenicity of in vitro- and in vivo-produced IJs were of 47–64% and 31%, respectively. It is worth noting that none of the two solid media had a statistically significant difference in IJ pathogenicity. Considering that the maximum multiplication factor of IJs on solid media was 2108 and that the pathogenicity against G. mellonella was outstanding, Steinernema sp. has a good potential for in vitro mass production.

scholarly journals Viability of Peranakan Etawah Liquid Semen Preserved in Tris Substituted with Various Energy Sources

2020 ◽  
Vol 25 (2) ◽  
pp. 68
Author(s):  
Nurul Afzan Hilda Zakiya ◽  
A H Yanti ◽  
T R Setyawati
Keyword(s):  
Energy Source ◽  
Artificial Insemination ◽  
Block Design ◽  
Egg Yolk ◽  
Energy Sources ◽  
Randomized Block Design ◽  
Frozen Semen ◽  
Semen Preservation ◽  
Semen Extender ◽  
Fresh Semen

The use of liquid semen for artificial insemination program of Etawah crossbreed goat (PE) is an alternative to replace frozen semen which is constrained by limited and expensive facilities. Production of liquid semen is faster than frozen semen, but the viability of liquid semen which preserved with a standard extender such as tris egg yolk is very short. The purpose of this study was to determine the viability of PE goat semen in egg yolk tris substituted with energy sources such as glucose, galactose, and mannose and to determine the most efficient energy source for semen preservation. This research was conducted from August to September 2018 at the Artificial Insemination Center in Lembang, West Java. This study was designed in a randomized block design (RBD) consist of three experimental groups divided into five groups. Fresh semen of PE goats were preserved using extender which energy source has been modified. Results showed that using glucose in PE goat semen extender produced the best motility among other groups (64.29 ± 9.2%). The highest viability was found in extender with fructose substitution (86.76 ± 2.3%). The longest viability of liquid semen was found in the extender with glucose substitution. It lasted for six days.

scholarly journals Repeatability and Phenotypic Correlation Among Semen Quality Traits in Holstein Bulls

2018 ◽  
Vol 42 (4) ◽  
Author(s):  
Argi Argiris ◽  
Siswanto Imam Santoso ◽  
Yon Supri Ondho ◽  
Edy Kurnianto
Keyword(s):  
Artificial Insemination ◽  
Semen Quality ◽  
Intraclass Correlation ◽  
Correlation Method ◽  
Phenotypic Correlation ◽  
Quality Traits ◽  
Frozen Semen ◽  
Holstein Bulls ◽  
Fresh Semen ◽  
Selection Of

The purpose of this research was to analysis the value of repeatability and correlation among the traits affecting the production of frozen semen from Holstein’s bull in Indonesia. Repeatability and correlation were calculated based on the data of frozen semen production of 15.699 records from 44 Holstein bulls at Singosari Artificial Insemination Center (SAIC) and 8.935 records from 39 Holstein bulls at Lembang Artificial Insemination Center (LAIC). Repeatability for volume, motility, fresh semen concentration and frozen semen production was evaluated by intraclass correlation method. The repeatability values of LAIC for volume, motility, fresh semen concentration and frozen semen production were 0.60; 0.54; 0.37 and 0.47. The repeatability values of SAIC for volume, motility, fresh semen concentration and frozen semen production were 0.54; 0.30; 0.43 and 0.29. The linear correlation value between volume, motility and fresh semen concentration with the amount of semen produced per collections were 0.41, 0.36, and 0.58. Concentration was the most factors influencing the number of frozen semen produced. The effectiveness of the selection of Holstein's frozen semen producing could be determined by the value of repeatability and the phenotypic correlation among semen quality traits such as volume, motility, concentration and frozen semen production.

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