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

Iaffaldano, Nicolaia; Iorio, Michele Di; Manchisi, A.; Esposito, Stefano; Gibertoni, Pier Paolo

doi:10.1017/s0967199415000647

Cited by 14 publications

(29 citation statements)

References 34 publications

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“…The freezing procedure tested here turned out to have a positive impact on sperm total motility post-thawing, recording values above 80%. These results were remarkably better compared to that found in our previous studies, which ranged between 33% and 52% [5][6][7]. However, a significant decrease in the total motility after thawing compared to the fresh semen was recorded, although this is expected following the cryopreservation process.…”

Section: Discussioncontrasting

confidence: 61%

“…However, a significant decrease in the total motility after thawing compared to the fresh semen was recorded, although this is expected following the cryopreservation process. In fact, it is known that the freezing and thawing of sperm is a complex process that causes several forms of cellular damages [2,5]. These injuries have been mainly attributed to cold shock, extreme osmotic changes, intracellular ice crystals and reactive oxygen species, with the consequent reduction of the membrane's permeability and integrity, motility, viability and fertilizing ability [14].…”

Section: Discussionmentioning

confidence: 99%

“…Thus, the conservation of cell structure and its functionality strictly depends on the cryopreservation protocol used. In this regard, in the past years our research group has been perfecting an effective semen cryopreservation protocol for the Mediterranean trout of the Molise rivers, with the aim of reducing cell damage and preserving the fertilizing ability of sperm [5][6][7]. However, despite our efforts and promising results obtained so far, we still aim to obtain a freezing procedure reporting a fertilization rate as similar as possible to that of fresh semen.…”

Section: Introductionmentioning

confidence: 99%

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

Rusco

Iorio

Iampietro

et al. 2020

Animals

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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.

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Section: Discussioncontrasting

confidence: 61%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

Rusco

Iorio

Iampietro

et al. 2020

Animals

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“…Most of the current protocols still use DMSO as permeable cryoprotectant, and DMSO is still considered as a suitable candidate for the development of new protocols. For instance, DMSO was used lately in the development of a protocol for the endangered Mediterranean brown trout Salmo trutta macrostigma [5]. However, methanol was suggested as an alternative to DMSO and tested in three salmonids including brown trout [6].…”

Section: Introductionmentioning

confidence: 99%

Comparing methanol-glucose and dimethyl-sulfoxide based extender for milt cryopreservation of brown trout (Salmo trutta)

Nusbaumer

Wedekind

2018

Preprint

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20The potential importance of sperm cryopreservation for aquaculture and conservation 21 management seems still undervalued, probably because the available protocols often lead to 22 reduced fertilization success. We experimentally compared the effectiveness of two different 23 freezing extenders for cryopreservation of brown trout (Salmo trutta) semen, controlling for 24 possible male and female effects. The methanol-glucose based extender that we tested was 25 significantly more effective than a common dimethyl-sulfoxide based extender (a commercial 26 cryopreservation kit). We then studied the effectiveness of the methanol-glucose based 27 extender at different sperm-egg ratios and found no significant differences in fertilization 28 ability of fresh and cryopreserved milt at a sperm-egg ratio of at least 110,000:1. We conclude 29 that brown trout sperm cryopreserved with this extender can be used even at low sperm-egg 30 ratios without significant effects on fertilization rates. 31 32 Key words 33 Sperm, cryopreservation, methanol, glucose, brown trout 34 35 36 49 Since the first attempts, much progress has been made and effective protocols now exist for a 50 variety of fishes, mainly freshwater fishes [1]. Salmonids have been a focus of these research 51 efforts due to their commercial and cultural importance [1]. The first successful 52 cryopreservation was achieved using glycerol as a cryoprotectant [4], but it was quickly 53 replaced by dimethyl sulfoxide (DMSO). Most of the current protocols still use DMSO as 54 permeable cryoprotectant, and DMSO is still considered as a suitable candidate for the 55 development of new protocols. For instance, DMSO was used lately in the development of a 56 protocol for the endangered Mediterranean brown trout Salmo trutta macrostigma [5]. 57However, methanol was suggested as an alternative to DMSO and tested in three salmonids 58 including brown trout [6]. Permeable cryoprotectants are most often associated with complex 59 saline solutions and non-permeable cryoprotectants, such as egg yolk. Recently, a very simple 60 extender consisting only of 9% methanol and 0.15M glucose was shown to be effective in 61 rainbow trout (Oncorhynchus mykiss) and in brown trout [7,8]. 62 63Here we compare the effectiveness of this methanol-glucose based extender to a DMSO-64 based extender on brown trout while controlling for potentially confounding parental or 65 population effects on fertilization success. The DMSO-based extender we use here is a 66 commercial product that use DMSO as permeable cryoprotectant, egg yolk derived lipids as 67 non-permeable cryoprotectant, and a saline solution. We ran two experiments. The first one 68 aimed to assess which of the two candidate extenders would produce the highest fertilization 69 success. In the second experiment, we tested the post-thaw fertilizing ability of sperm at 70 various dilutions. 16.5%, 1.65% and 0.165%, implying an absolute volume of milt of 66, 6.6 and 0.6 µL of milt 131 in extender or Storfish, respectively. We used 200 µL of...

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“…A widely cited and relatively recent taxonomic assessment of European trouts (Kottelat & Freyhof, 2007) considers Salmo cettii as the taxon including most of the autochthonous trout of the Tyrrhenian basin (with the exception of the restricted endemism Salmo fibreni, from Lake Fibreno, central Italy) and suggests avoiding the name S. (trutta) macrostigma, which was (and still is) often applied to some Italian populations (see Gandolfi, 1991;Iaffaldano, Di Iorio, Manchisi, Esposito & Gibertoni, 2016;Querci, Pecchioli, Leonzio, Frati & Nardi, 2013). There is no current consensus about the validity of S. cetti (sensu Kottelat & Freyhof, 2007) as a species (or even as a distinct taxon within the S. trutta complex) and some authors prefer to simply include it into a more comprehensive "Mediterranean" S. trutta (e.g.…”

Section: Introductionmentioning

confidence: 99%

Investigating the genetic structure of trout from the Garden of Ninfa (central Italy): Suggestions for conservation and management

Fabiani

Gratton

Zappes

et al. 2017

Fisheries Management Eco

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Mediterranean populations of brown trout (Salmo trutta L. complex) have lost a large part of their genetic distinctiveness, mostly due to massive restocking, and the waters of the Gardens of Ninfa (province of Latina, central Italy, Site of Community Importance since 2013) are regarded as one of a few potential reservoirs of autochthonous trout lineages in the Tyrrhenian drainage of the Italian peninsula. In this study, nuclear and mitochondrial markers were used on brown trout samples from Ninfa to estimate non-Mediterranean influence in the population gene pool, potential changes of genetic structure over time and genetic relationships with other sites known (or suspected) to host native trout gene pools. Striking changes in both microsatellite and mtDNA allele frequencies over a 9-year time span were found and provided evidence of unrecorded stocking from the nearby Lake Fibreno. Results are analysed in the light of potential ecological consequences of such events on a longer time scale and provide a scientific background for fisheries management and conservation programmes in the area

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

Cited by 14 publications

References 34 publications

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

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

Comparing methanol-glucose and dimethyl-sulfoxide based extender for milt cryopreservation of brown trout (Salmo trutta)

Investigating the genetic structure of trout from the Garden of Ninfa (central Italy): Suggestions for conservation and management

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