Use of glucose or BTS™ combined with DMSO or methylglycol under two differentfreezing protocols for the cryopreservation of sperm from the common curimatã(Prochilodus brevis)

This study evaluated the effect of glucose orBeltsville Thawing Solution (BTS™) combined withdimethyl sulfoxide (DMSO) or methylglycol (MG)under two different freezing protocols on the kineticsand morphology of cryopreserved Prochilodus brevissperm. The semen samples were diluted using one of fourdifferent treatments (glucose+DMSO, glucose+MG,BTS™+DMSO, and BTS™+MG), loaded into 0.25-mlstraws and subjected to two different freezing processes(programmed freezing machine and dry shipper). After10 days, the semen samples were thawed, and the spermmorphology and kinetics were evaluated. Thephysicochemical parameters of the semen in naturawere similar to those observed in other studies ofCharaciformes, indicating the feasibility of semencryopreservation. Glucose, when used as a diluentwith the cryoprotectant MG (glucose+MG), yieldedhigher percentages of mobile spermatozoa afterfreezing in a dry shipper (76.88 ± 4.84%) and in aprogrammed freezing machine (70.95 ± 1.76%)compared with the combination of glucose and DMSO.Moreover, the glucose+MG treatment yielded a highersperm velocity (curvilinear velocity: 79.52 ± 2.88 µm s-1; straight-line velocity: 45.46 ± 3.01 µm s-1; averagepath velocity: 67.92 ± 3.08 µm s-1) than the otherstudied treatments, and a higher amount of normalsperm (74.56 ± 0.77%) was observed in the semensamples cryopreserved using a programmed freezingmachine. The sperm abnormalities observed included abent tail morphology. Therefore, the use ofglucose+MG in combination with either a dry shipper ora programmed freezing machine is recommended forthe cryopreservation of P. brevis sperm because thesemethods yielded high numbers of motile andmorphologically normal spermatozoa.

Use of glucose or BTS™ combined with DMSO or methylglycol under two differentfreezing protocols for the cryopreservation of sperm from the common curimatã(Prochilodus brevis)

This study evaluated the effect of glucose orBeltsville Thawing Solution (BTS™) combined withdimethyl sulfoxide (DMSO) or methylglycol (MG)under two different freezing protocols on the kineticsand morphology of cryopreserved Prochilodus brevissperm. The semen samples were diluted using one of fourdifferent treatments (glucose+DMSO, glucose+MG,BTS™+DMSO, and BTS™+MG), loaded into 0.25-mlstraws and subjected to two different freezing processes(programmed freezing machine and dry shipper). After10 days, the semen samples were thawed, and the spermmorphology and kinetics were evaluated. Thephysicochemical parameters of the semen in naturawere similar to those observed in other studies ofCharaciformes, indicating the feasibility of semencryopreservation. Glucose, when used as a diluentwith the cryoprotectant MG (glucose+MG), yieldedhigher percentages of mobile spermatozoa afterfreezing in a dry shipper (76.88 ± 4.84%) and in aprogrammed freezing machine (70.95 ± 1.76%)compared with the combination of glucose and DMSO.Moreover, the glucose+MG treatment yielded a highersperm velocity (curvilinear velocity: 79.52 ± 2.88 µm s-1; straight-line velocity: 45.46 ± 3.01 µm s-1; averagepath velocity: 67.92 ± 3.08 µm s-1) than the otherstudied treatments, and a higher amount of normalsperm (74.56 ± 0.77%) was observed in the semensamples cryopreserved using a programmed freezingmachine. The sperm abnormalities observed included abent tail morphology. Therefore, the use ofglucose+MG in combination with either a dry shipper ora programmed freezing machine is recommended forthe cryopreservation of P. brevis sperm because thesemethods yielded high numbers of motile andmorphologically normal spermatozoa.(AU)

Cinética e morfologia de espermatozoides de carpa comum criopreservados em água de coco em pó ACP-104 / Kinetics and morphology of common carp sperm cryopreserved in powdered coconut water ACP-104

O objetivo deste estudo foi avaliar o efeito de diferentes taxas de diluição seminal sobre a motilidade, velocidade e presença de anormalidades espermáticas no sêmen descongelado de carpa comum (Cyprinus carpio). Utilizaram-se 20 machos sexualmente maduros, pertencentes ao Departamento Nacional de Obras Contra as Secas (DNOCS). A partir do sêmen coletado, foi formado um total de nove pools. O sêmen de cada pool foi avaliado quanto à motilidade, velocidade e morfologia espermática antes e depois da criopreservação. A criopreservação seminal foi realizada em meio ACP-104 + DMSO 10% diluído em 1:1 ou 1:3 (sêmen:diluidor). As amostras foram envasadas em palhetas de 0,25mL, congeladas em vapor de nitrogênio líquido (caixa de poliestireno) e estocadas em nitrogênio líquido (-196 °C). Não houve diferença significativa (P>0,05) entre as taxas de diluições sobre os parâmetros cinéticos e morfológicos de sêmen descongelado de carpa comum. Entretanto, esses parâmetros apresentaram diferença significativa (P<0,05) em relação ao sêmen fresco (controle). Baixos valores de velocidade foram registrados no sêmen descongelado (VCL 42,6-46,5μm/s; VSL 33,2-37,1μm/s; VAP 38,9-43,2μm/s); contudo, observaram-se motilidades acima de 59% (59,8-67,8%) e baixos índices de anormalidades espermáticas (19,3-19,5%), sugerindo que o ACP pode ser usado como um meio favorável à criopreservação do sêmen de carpa comum em qualquer uma das taxas de diluição testadas.(AU)

The aim of this study was to determine the effects of various dilution ratios on thaw sperm motility, velocity and presence of spermatic abnormalities in the common carp. 20 males with three years of age, raised at the Departamento Nacional de Obras Contra as Secas (DNOCS) were used. From the collected semen a total of nine pools were formed. The semen from each pool was evaluated for motility, velocity and spermatic morphology before and after cryopreservation. The sperm cryopreservation was performed in medium ACP-104 + DMSO 10% diluted in 1:1 or 1:3 (sperm:extender). The samples were loaded into 0.25mL straws, frozen in liquid nitrogen vapor (Styrofoam) and stored in liquid nitrogen (-196 °C). There was no significant difference (P>0.05) between the ratios of dilution on the kinetic and morphological parameters of thawed semen from common carp. However, these parameters were significantly different (P<0.05) compared to fresh semen (control). Low velocity values were recorded in thawed semen (VCL 42.6-46.5μm/s; VSL 33.2-37.1μm/s; VAP 38.9-43.2μm/s), however, observed motility above 59% (59.8-67.8%), and low levels of spermatic abnormalities (19.3-19.5%) suggested that the ACP-104 can be used as a suitable medium for cryopreservation of common carp semen in any of the tested dilution ratios.(AU)

Avaliação de diferentes crioprotetores e taxas de diluição na criopreservação seminal de Prochilodus brevis / Evaluation of different crioprotectants and dilution ratios on Prochilodus brevis sperm cryopreservation

Este estudo objetivou caracterizar o sêmen de Prochilodus brevis e avaliar os efeitos de diferentescrioproterores e taxas de diluição sobre a cinética e a morfologia do sêmen criopreservado. Inicialmente,amostras seminais de P. brevis (n = 40) foram analisadas quanto ao volume, pH, osmolaridade, concentração,motilidade e morfologia espermáticas. Para a criopreservação, o material coletado foi distribuído em 10 pools desêmen (n = 10), submetidos a seis tratamentos compostos pela combinação de glicose 5%, dois crioprotetores(DMSO ou MG) e três taxas de diluição (1:3, 1:6 ou 1:9 sêmen:diluidor). As amostras, in natura e pósdescongeladas,foram analisadas quanto à sua morfologia e cinética espermática utilizando o CASA. O sêmen deP. brevis apresentou características semelhantes a demais espécies de Prochilodus. O sêmen congelado comglicose e MG apresentou resultados significativamente superiores (P < 0,05) comparado àquele congeladoutilizando DMSO. As diluições 1:3 e 1:6 apresentaram melhores resultados para glicose + MG, enquanto 1:9 foimelhor para glicose + DMSO. Portanto, a associação glicose + MG, numa taxa de diluição de 1:3 ou 1:6, é amais indicada para a criopreservação do sêmen de Prochilodus brevis.(AU)

This study aimed to characterize Prochilodus brevis sperm and evaluate the effects of different diluentsand dilution ratios on kinetics and morphology of cryopreserved sperm. Initially, sperm samples of P. brevis(n = 40) were assessed for volume, pH, osmolality, spermatic concentration, motility and morphology. Forcryopreservation, the collected material was distributed in ten pools of semen (n = 10), submitted to sixtreatments composed by a combination of glucose 5% with two cryoprotectants (DMSO or MG) and threedilution ratios (1:3, 1:6 or 1:9 - sperm:diluent). In natura and post-thawed samples were assessed for spermaticmorphology and kinetics using CASA. The sperm of P. brevis showed similar characteristics to otherProchilodus species. Sperm cryopreserved in glucose plus MG presented significantly higher results (P < 0.05)compared to sperm frozen in DMSO. The dilution ratios of 1:3 and 1:6 yielded better results for glucose + MG,while 1:9 was the best dilution for glucose + DMSO. In conclusion, the association of glucose + MG, in adilution ratio of 1:3 or 1:6, is the most indicated for Prochilodus brevis sperm cryopreservation.(AU)

Avaliação de diferentes crioprotetores e taxas de diluição na criopreservação seminal de Prochilodus brevis / Evaluation of different crioprotectants and dilution ratios on Prochilodus brevis sperm cryopreservation

Este estudo objetivou caracterizar o sêmen de Prochilodus brevis e avaliar os efeitos de diferentescrioproterores e taxas de diluição sobre a cinética e a morfologia do sêmen criopreservado. Inicialmente,amostras seminais de P. brevis (n = 40) foram analisadas quanto ao volume, pH, osmolaridade, concentração,motilidade e morfologia espermáticas. Para a criopreservação, o material coletado foi distribuído em 10 pools desêmen (n = 10), submetidos a seis tratamentos compostos pela combinação de glicose 5%, dois crioprotetores(DMSO ou MG) e três taxas de diluição (1:3, 1:6 ou 1:9 – sêmen:diluidor). As amostras, in natura e pósdescongeladas,foram analisadas quanto à sua morfologia e cinética espermática utilizando o CASA. O sêmen deP. brevis apresentou características semelhantes a demais espécies de Prochilodus. O sêmen congelado comglicose e MG apresentou resultados significativamente superiores (P < 0,05) comparado àquele congeladoutilizando DMSO. As diluições 1:3 e 1:6 apresentaram melhores resultados para glicose + MG, enquanto 1:9 foimelhor para glicose + DMSO. Portanto, a associação glicose + MG, numa taxa de diluição de 1:3 ou 1:6, é amais indicada para a criopreservação do sêmen de Prochilodus brevis.

This study aimed to characterize Prochilodus brevis sperm and evaluate the effects of different diluentsand dilution ratios on kinetics and morphology of cryopreserved sperm. Initially, sperm samples of P. brevis(n = 40) were assessed for volume, pH, osmolality, spermatic concentration, motility and morphology. Forcryopreservation, the collected material was distributed in ten pools of semen (n = 10), submitted to sixtreatments composed by a combination of glucose 5% with two cryoprotectants (DMSO or MG) and threedilution ratios (1:3, 1:6 or 1:9 - sperm:diluent). In natura and post-thawed samples were assessed for spermaticmorphology and kinetics using CASA. The sperm of P. brevis showed similar characteristics to otherProchilodus species. Sperm cryopreserved in glucose plus MG presented significantly higher results (P < 0.05)compared to sperm frozen in DMSO. The dilution ratios of 1:3 and 1:6 yielded better results for glucose + MG,while 1:9 was the best dilution for glucose + DMSO. In conclusion, the association of glucose + MG, in adilution ratio of 1:3 or 1:6, is the most indicated for Prochilodus brevis sperm cryopreservation.