Rodrigo Muíño

Universidade de Santiago de Compostela

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Joaquín Hernández Bermúdez

Fundación Hospital San Pedro

C. Castillo

Universidade de Santiago de Compostela

J. L. Benedito

Universitat Politècnica de València

C.O. Hidalgo

Servicio Regional de Investigación y Desarrollo Agroalimentario

Ana I. Peña

Universidade de Santiago de Compostela

Víctor Pereira

Universidade de Santiago de Compostela

Carolina Tamargo

Servicio Regional de Investigación y Desarrollo Agroalimentario

Marc Yeste

University of Girona

José Luis Benedito Castellote

Universidade de Santiago de Compostela

Joan E. Rodríguez‐Gil

Universitat Autònoma de Barcelona

Cooperative Institutions

Universidade de Santiago de Compostela

Universitat Autònoma de Barcelona

University of Girona

Universidad de Murcia

Centro Regional de Selección y Reproducción Animal

Institute for Animal Reproduction

Universidade Federal do Rio Grande do Sul

Ceres

Universidad Complutense de Madrid

Consejo Superior de Investigaciones Científicas

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Cryopreservation of bovine sperm causes single-strand DNA breaks that are localized in the toroidal regions of chromatin

Journal of Animal Science and Biotechnology/Journal of animal science and biotechnology (2024)

Jordi Ribas‐Maynou Rodrigo Muíño Carolina Tamargo Marc Yeste

Abstract Background Sperm cryopreservation is widely used in the cattle industry, as it allows for disassociating the localization of sires and the collection of semen from the timing of artificial insemination. While freeze-thawing is known to impair sperm DNA integrity, whether the damage induced consists of single- (SSB) or double-strand breaks (DSB) has not been determined. In addition, no previous study has addressed if DNA breaks preferentially reside in specific genome regions such as those forming the toroid linker regions, or are rather spread throughout the regions linked to protamines. The main aim of the present work, therefore, was to elucidate the type and localization of the DNA damage generated by cryopreservation and to evaluate its impact on artificial insemination outcomes in cattle. Results The incidence of SSB and DSB was evaluated in 12 ejaculates before and after cryopreservation with the Comet assay, and the localization of the DNA breaks was assessed using pulsed-field gel electrophoresis (PFGE). Before cryopreservation, the incidence of SSB was 10.99% ± 4.62% and involved 20.56% ± 3.04% of sperm cells, whereas these figures significantly ( P < 0.0001) increased up to 34.11% ± 3.48% and 53.36% ± 11.00% in frozen-thawed sperm. In contrast, no significant differences in the incidence of DSB were observed ( P > 0.990) before and after cryopreservation (before: incidence of 13.91% ± 1.75% of sperm DNA affecting 56.04% ± 12.49% of sperm cells; after: incidence of 13.55% ± 1.55% of sperm DNA involving 53.36% ± 11.00% of sperm cells). Moreover, PFGE revealed that the percentage of sperm DNA fragments whose length was shorter than a toroid (< 31.5 kb) was greater ( P < 0.0001) after (27.00% ± 4.26%) than before freeze-thawing (15.57% ± 4.53%). These differences indicated that the DNA breaks induced by cryopreservation affect the regions condensed in protamines, which are structured in toroids. On the other hand, in vivo fertility rates were associated to the incidence of SSB and DSB in frozen-thawed sperm ( P = 0.032 and P = 0.005), but not with the size of the DNA fragments resulting from these breaks ( P > 0.05). Conclusion Cryopreservation of bovine sperm generates single-strand DNA breaks, which are mainly located in protamine-condensed toroidal regions. The incidence of DNA breaks in cryopreserved sperm has an impact on cattle fertility, regardless of the size of generated fragments.

10.1186/s40104-024-01099-0

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Is adenosine deaminase (ADA) activity in saliva and serum a more accurate disease detection tool than traditional redox balance parameters in early-lactating dairy cows?

Veterinary Research Communications (2023)

C. Castillo Joaquín Hernández Bermúdez Juan Sotillo Rodrigo Muíño J. L. Benedito

Abstract Enzyme adenosine deaminase (ADA) is a marker of inflammation in domestic animals, but it is unclear whether it is a reliable marker of oxidative stress, especially in the transition period in dairy cows. This study aims to assess if ADA and redox status measurements in saliva provide the same utility to detect disease condition as that obtained from serum. Sixty-eight multiparous Holstein cows, between 2 and 3 weeks postpartum were selected. Five study groups were established: control (healthy), and cows with ketosis, mastitis, laminitis, and metritis. The parameters measured were ADA activity, total oxidants (TOS), antioxidants (TAC), and OSi ratio. Regarding redox status, no significant differences arise in both saliva and serum being the correlations negative and not significant. In saliva, ADA activity in healthy cows differs from those with pathological processes, having the lowest activities. In serum, ADA activity is similar in the healthy and ketosis cows, showing the lowest activities meanwhile animals with mastitis, laminitis, or metritis have significantly higher activities. In conclusion, the measurement of ADA activities and redox status in saliva does not give consistent results, being preferable to measure them in serum during the transition period.

10.1007/s11259-023-10069-2

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Evaluation of Anogenital Distance and Anti-Müllerian Hormone Plasmatic Concentration as Potential Phenotypes to Predict Reproductive Performance in Holstein Heifers

Veterinary Sciences (2024)

L. Vidal Jacobo Álvarez Uxía Yáñez J. Caínzos Rodrigo Muíño

Anogenital distance (AGD) is a marker of the degree of prenatal exposure to androgens in multiple species, and it has been suggested that there is an inverse association between AGD and fertility. Therefore, the aim of this study was to determine the usefulness of AGD and anti-Müllerian hormone (AMH) concentrations, an indirect marker of the follicular population, as predictors of future reproductive potential in Holstein cattle. The AGD was measured in 566 females from 9 dairy farms in Galicia (Spain). A group of 172 females underwent a second measurement 9 months after. Additionally, data on the age at first insemination (1stAI age), number of AI (AI-PREG), age at first pregnancy (1stPREG age), age at first calving (1stCAL age), and calving-pregnancy (CAL-PREG) and calving-calving (CAL-CAL) intervals were collected. Blood samples were collected from 80 heifers to determine AMH concentrations. Our results showed that AGD varied minimally with age, and that cows with short AGD had earlier 1stAI age, 1stPREG age, and 1stCAL age (

Anogenital distance

Anti-Müllerian hormone

10.3390/vetsci11100495

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Evaluation of sperm subpopulation structure in relation to in vitro sperm–oocyte interaction of frozen-thawed semen from Holstein bulls

Theriogenology (2014)

Marcia de Almeida Monteiro Melo Ferraz Roser Morató Marc Yeste Núria Arcarons Ana I. Peña

10.1016/j.theriogenology.2014.01.033

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Citations (54)

Suspected botulism in a herd of Holstein heifers.

Albéitar (2012)

Rodrigo Muíño C. Castillo Joaquín Hernández Bermúdez Marta López‐Alonso Víctor Pereira

Botulism

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Relationship of BCS prepartum with reproductive performance and lipomobilization in Holstein dairy cows

Pakistan Veterinary Journal (2017)

José Chapel Rodrigo Muíño Víctor Pereira C. Castillo Joaquín Hernández Bermúdez

Body condition score (BCS) changes and non-esterified fatty acids (NEFA) are good indicators for measuring lipomobilization during the transition period. The aim of this study was to establish the possible correlation between cows with a higher BCS during this period and their reproductive performance postpartum. A total of 47 Holstein dairy cows were randomly allotted into two groups according to their prepartum BCS, including: G1 (BCS 3.5 to 3.75, n = 20) and G2 (BCS > 3.75, n=27). Moreover, another two groups were established at 14th day postpartum: G3, 37 cows lost more than 0.75 points of BCS and G4, 10 cows lost 0.75 points or less. Blood samples were obtained from 10 to 7 days before expected calving, and 14 days postpartum in all groups and serum concentrations of glucose, s-hydroxybutyrate, NEFA, urea, calcium and hepatic enzymes, and reproductive parameters were determined. NEFA values showed that cows of G2 and G4 had a higher lipomobilization than G1 and G3. Moreover, cows with a BCS of more than 3.75 and NEFA values of more than 495 μmol/L prepartum had a higher risk of non-pregnancy at first AI, and there was an increase in the number of open cows at 150 DIM, this being the reason why the elimination rate due to non-pregnancy increases (20%) and their reproductive lives shorten.

NEFA

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BOTULISMO TIPO C EM UM REBANHO DE NOVILHAS LEITEIRAS NO NOROESTE DA ESPANHA

Archives of Veterinary Science (2012)

Rodrigo Muíño Víctor Pereira J. L. Benedito C. Castillo Joaquín Hernández Bermúdez

Intoxicações por toxina botulínica dos tipos B e C tem sido relatadas em bovinos como consequência do consumo de alimento contaminado com carcaças putrefatas de animais ou cama de frango, mas silagem de pasto embalada em fardos com cobertura plástica, também pode ser fonte de intoxicações botulínicas. No presente trabalho, realizado em um rebanho de novilhas leiteiras no Noroeste da Espanha, a assistência veterinária foi procurada devido a que 21 novilhas apresentavam decúbito com suas cabeças flexionadas lateralmente, letárgicas, relutantes a movimentar-se e com os membros contraídos. Em poucos animais foi possível observar salivação excessiva e dificuldade para retrair a língua. No exame físico não se encontrou motivo para o decúbito e as novilhas foram tratadas para aliviar os sinais clínicos. Uma semana depois do surto, 8 novilhas morreram e 5 novilhas foram eutanasiadas por motivos éticos, tendo sobrevivido apenas 8 novilhas. As novilhas estavam alimentadas com ração misturada a vontade constituída por silagem fresca, feno e ração comercial. Foi verificado que a cobertura plástica da silagem estava danificada aos lados, estragada e com mofo. Em análise posterior foi encontrada toxina tipo C de Clostridium botulinum a partir de amostras de silagem mofada. Os resultados junto com a história clínica e os sinais clínicos levaram ao diagnóstico de botulismo tipo C.

10.5380/avs.v17i3.25775

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Post‐thaw Survival and Longevity of Bull Spermatozoa Frozen with an Egg Yolk‐based or Two Egg Yolk‐free Extenders after an Equilibration Period of 18 h

Reproduction in Domestic Animals (2007)

Rodrigo Muíño M. Fernández AI Peña

The aim of the present study was to determine the suitability of using two egg yolk-free commercial extenders, Andromed and Biociphos Plus as compared with the Tris-egg yolk based diluent Biladyl, for the cryopreservation of bull spermatozoa when the freezing protocol involved holding the extended semen at 4 degrees C for 18 h before the freezing. Six ejaculates from each of 10 Holstein bulls were collected by using artificial vagina. The ejaculates were evaluated for volume, sperm concentration and motility, divided in to three equal volumes, and diluted, respectively, with the three extenders as specified above. Extended semen was equilibrated for 18 h at 4 degrees C and frozen in 0.25-ml straws. After thawing, 100-mul aliquots of semen were labelled with SYBR-14, PI and PE-PNA (Phycoerythrin-conjugated Peanut agglutinin) and analysed by flow cytometry at 0, 3, 6 and 9 h after incubation at 37 degrees C. A General Lineal Model procedure for repeated measures was used to determine the effects of extender, bull, replicate and the interaction between them, on sperm viability and acrosomal integrity. Semen samples frozen with Biladyl showed higher (p < 0.001) sperm survival after 0 h (47.9%) and 9 h (30.3%) of incubation than those frozen with Andromed (38.5% and 17.3%, after 0 and 9 h respectively) or Biociphos Plus (34.9% and 21.6%, after 0 and 9 h respectively). The bull and replicate had significant effects (p < 0.001) on both sperm viability and acrosomal integrity, but the interactions between bull and extender and between replicate and extender were not significant. It was concluded that, when holding the semen overnight before freezing, the use of Biladyl results in higher sperm survival and longevity than the use of Andromed or Biociphos Plus.

Yolk

10.1111/j.1439-0531.2006.00784.x

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Characterization of the Germplasm Bank for the Spanish Autochthonous Bull Breed “Asturiana de la Montaña”

Animals (2023)

Carolina Tamargo Amer Salman J. N. Caamaño Felipe Martínez‐Pastor Ángel Fernández

Semen cryobanks are critical for preserving autochthonous and rare breeds. Since sperm cryopreservation has been optimized for commercial breeds, non-commercial ones (often endangered) must be characterized to ensure the germplasm's viability. This study reports an investigation of the "Asturiana de la Montaña" breed (AM), a valuable Spanish autochthonous cattle breed adapted to the mountainous Atlantic environment. The survey included cryopreserved semen doses from 40 bulls stored at the Principado de Asturias Germplasm Bank. Data were obtained from the routine fresh semen analysis, CASA (motility), and flow cytometry analyses of fresh and post-thawing semen, and the 56-day non-return-rate (NRR) in heifers and cows (all results as 1st and 3rd quartiles). Fresh samples (artificial vagina) were within the normal range for cattle (4-6 mL, 5-10 × 109/mL; mass motility 5). Post-thawing results showed motility below typical for commercial breeds (total motility 26-43%, progressive 14-28%), with higher values for viability (47-62%). Insemination results showed a good performance for this breed (NRR: 47-56%; higher for heifers). Sperm volume increased with age, with little or no effects on sperm quality. Few associations were found between post-thawing quality or freezability and NRR, LIN being the variable more strongly associated (positively). The AM semen bank shows a good prospect for preserving and disseminating the genetics of this breed. This survey indicates that dedicated research is needed to adapt freezing protocols to this breed, optimizing post-thawing results.

Germ plasm

Semen quality

10.3390/ani13081402

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Relationship among blood indicators of lipomobilization and hepatic function during early lactation in high-yielding dairy cows

Journal of Veterinary Science (2011)

Félix González Rodrigo Muíño Víctor Pereira Rómulo Campos Gaona J. L. Benedito

Blood indicators are used as a tool to diagnose metabolic disorders. The present work was conducted to study the relationships among blood indicators of lipomobilization and hepatic function in high-yielding dairy cows. Two groups of Holstein cows were studied: 27 early lactation cows and 14 mid lactation cows from four different herds with similar husbandry characteristics in Galicia, Spain. Blood samples were obtained to measure beta-hydroxybutyrate (BHB), non-esterified fatty acids (NEFA), triglycerides (TG), and the activity of aspartate transaminase (AST) and gamma-glutamyl transferase. Cows in early lactation had higher levels of BHB and NEFA than mid lactation cows. High lipomobilization (NEFA > 400 µmol/L) was detected in 67% and 7% of early lactation and mid lactation cows, respectively, while subclinical ketosis (BHB > 1.2 mmol/L) was detected in 41% and 28% of the early lactation and lactation cows, respectively. TG concentrations were low in all cows suffering subclinical ketosis and in 61% of the cows with high lipomobilization. During early lactation, 30% of cows suffered hepatic lipidosis as detected by levels of AST. Compromised hepatic function was observed in early lactation cows as shown by lower concentrations of glucose, total protein, and urea.

NEFA

Ketosis

Transaminase

10.4142/jvs.2011.12.3.251

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Citations (225)