Animal Reproduction (AR)
https://animal-reproduction.org/article/doi/10.1590/1984-3143-AR2023-0113
Animal Reproduction (AR)
ORIGINAL ARTICLE

Development of assays for the characterization of sperm motility parameters, viability, and membrane integrity in the epididymis and vas deferens of the greater rhea (Rhea americana)

Luana Grasiele Pereira Bezerra; Andréia Maria Silva; Maiko Roberto Tavares Dantas; Romário Parente dos Santos; Samara Sandy Jeronimo Moreira; Ana Glória Pereira; Moacir Franco de Oliveira; Pierre Comizzoli; Alexandre Rodrigues Silva

Downloads: 0
Views: 74

Abstract

Abstract: The objectives of the study were to (1) describe the kinematic parameters of spermatozoa (2) compare methods of evaluating sperm viability (3) validate assays of functionality and integrity of the sperm membrane and (4) evaluate possible changes between spermatozoa from the epididymis and the vas deferens of the greater rhea. Semen samples were recovered from 7 adult individuals. Sperm motility was characterized by adjusting the set-up for Computer-assisted semen analysis (CASA) to that new species. For sperm viability evaluation, smears of bromophenol blue and eosin-nigrosine dyes were used. Five solutions of different osmolarities were then tested for the hypoosmotic swelling test (HOST). The combination of fluorescent probes (propidium iodide - IP and Hoechst 33342) was also used to assess plasma membrane integrity. Data were presented as mean ± SEM. Rhea spermatozoa from the vas deferens had an overall motility of 14.6 ± 2.5%. The bromophenol blue staining technique revealed that 64.6 ± 5.2% sperm were viable, while that proportion was 72.1 ± 2.5% using eosin-nigrosine. An average of 77.6 ± 4.8% of spermatozoa reacted to the HOST with distilled water at 0 mOsm/l. Fluorescent probes indicated that 65.3 ± 2.6% of spermatozoa had intact membranes. Interestingly, no statistical differences were observed between the parameters analyzed in the epididymal spermatozoa and the vas deferens. These new assays set reference values that can now be used to further exploration of sperm handling conditions and freezing protocols in rheas.

Keywords

ratites, sperm pathways, avian sperm, bird semen, wildlife

References

Ahammad MU, Nishino C, Tatemoto H, Okura N, Kawamoto Y, Okamoto S, Nakada T. Maturational changes in the survivability and fertility of fowl sperm during their passage through the male reproductive tract. Anim Reprod Sci. 2011;128(1-4):129-36. http://dx.doi.org/10.1016/j.anireprosci.2011.09.010. PMid:21996579.

Amann RP, Waberski D. Computer-assisted sperm analysis (CASA): capabilities and potential developments. Theriogenology. 2014;81(1):5-17.e1-3. http://dx.doi.org/10.1016/j.theriogenology.2013.09.004. PMid:24274405.

Arruda RPD, Celeghini EDC, Alonso MA, Carvalho HF, Oliveira LZ, Nascimento J, Silva DF, Affonso FJ, Jaimes JD. Methods of the assessment of morphology and function of sperm: actual moment and future challenges. Rev Bras Reprod Anim. 2011;35(2):145-51.

Bernal B, Behnamifar A, Álvarez-Rodríguez C, Toledano-Díaz A, Castaño C, Velázquez R, Gil MG, Gutiérrez-Adán A, Woelders H, Blesbois E, Santiago-Moreno J. Transit along the vas deferens results in a high percentage of filiform spermatozoa with compacted chromatin in the rooster (Gallus domesticus). Reprod Fertil Dev. 2022;34(10):699-712. http://dx.doi.org/10.1071/RD21209. PMid:35366958.

Bezerra JA, Silva AM, Peixoto GC, Silva MA, Oliveira MF, Silva AR. Influence of recovery method and centrifugation on epididymal sperm from collared peccaries (Pecari tajacu Linnaeus, 1758). Zool Sci. 2014;31(5):338-42. http://dx.doi.org/10.2108/zs130149. PMid:24832907.

Bezerra LGP, Silva AM, Jurema AP, Dantas MRT, Pereira AG, Oliveira MF, Comizzoli P, Silva AR. Changes in sperm morphology, morphometry, and motility from the epididymis to the vas deferens in rheas (Rhea americana, Linnaeus, 1758). Animals (Basel). 2023a;13(9):1483. http://dx.doi.org/10.3390/ani13091483. PMid:37174520.

Bezerra LGP, Silva AM, Moreira SSJ, Souza CMP, Silva AR. Establishment of methods to analyze the structural and functional integrity of the quail (Coturnix coturnix japonica) sperm plasma membrane. Br Poult Sci. 2023b;64(3):429-34. http://dx.doi.org/10.1080/00071668.2022.2163615. PMid:36628926.

Brock M. Semen collection and artificial insemination in the hispaniolan parrot (Amazona ventralis). J Zoo Wildl Med [serial on the Internet]. 1991 [cited 2023 Jul 14];22:107-14. Available from: https://www.jstor.org/stable/20095125

Bucevičius J, Lukinavičius G, Gerasimaitė R. The use of Hoechst dyes for DNA staining and beyond. Chemosensors (Basel). 2018;6(2):18. http://dx.doi.org/10.3390/chemosensors6020018.

Celeghini ECC, Arruda RP, Albuquerque R, Silva FHA, Faria DE, Andrade AFC, Nascimento J, Raphael CF. Utilization of fluorescent probe association for simultaneous assessment of plasmatic, acrosomal, and mitochondrial membranes of rooster spermatozoa. Rev Bras Cienc Avic. 2007;9(3):143-9. http://dx.doi.org/10.1590/S1516-635X2007000300001.

Chalah T, Brillard JP. Comparison of assessment of fowl sperm viability by eosin-nigrosin and dual fluorescence (SYBR-14/PI). Theriogenology. 1998;50(3):487-93. http://dx.doi.org/10.1016/S0093-691X(98)00155-1. PMid:10732141.

Chatdarong K, Thuwanut P, Morrell JM. Single-layer centrifugation through colloid selects improved quality of epididymal cat sperm. Theriogenology. 2010;73(9):1284-92. http://dx.doi.org/10.1016/j.theriogenology.2009.12.009. PMid:20171724.

Convention on International Trade in Endangered Species of Wild Fauna and Flora – CITES. Appendices I, II and III. Geneva: CITES; 2023. p. 35. Available from: https://cites.org/sites/default/files/eng/app/2023/E-Appendices-2023-05-21.pdf

Cornwall GA, Hann SR. Specialized gene expression in the epididymis. J Androl. 1995;16(5):379-83. http://dx.doi.org/10.1002/j.1939-4640.1995.tb00548.x. PMid:8575976.

Dogliero A, Rota A, Lofiego R, von Degerfeld MM, Quaranta G. Semen evaluation in four autochthonous wild raptor species using computer-aided sperm analyzer. Theriogenology. 2016;85(6):1113-7. http://dx.doi.org/10.1016/j.theriogenology.2015.11.023. PMid:26725000.

du Plessis L, Soley JT. Light microscopic features and morphometry of sperm in the emu (Dromaius novaehollandiae). Theriogenology. 2014;81(2):203-9. http://dx.doi.org/10.1016/j.theriogenology.2013.08.016. PMid:24112875.

Fattah A, Sharafi M, Masoudi R, Shahverdi A, Esmaeili V. L-carnitine is a survival factor for chilled storage of rooster semen for a long time. Cryobiology. 2017;74:13-8. http://dx.doi.org/10.1016/j.cryobiol.2016.12.011. PMid:28042051.

Fischer D, Schneider H, Failing K, Meinecke‐Tillmann S, Wehrend A, Lierz M. Viability assessment of spermatozoa in large falcons (Falco spp.) using various staining protocols. Reprod Domest Anim. 2020;55(10):1383-92. http://dx.doi.org/10.1111/rda.13785. PMid:32722862.

Góes PAA, Cavalcante AKS, Nichi M, Perez EGA, Barnabe RC, Barnabe VH. Reproductive characteristics of captive greater rhea (Rhea americana) males reared in the state of São Paulo, Brazil. Rev Bras Cienc Avic. 2010;2(1):57-62. http://dx.doi.org/10.1590/S1516-635X2010000100009.

Jeyendran RS, van der Vem HH, Perez-Pelaez M, Crabo BG, Zaneveld LJ. Development of an assay to assess the functional integrity of the human sperm membrane and its relationship to other semen characteristics. J Reprod Fertil. 1984;70(1):219-28. http://dx.doi.org/10.1530/jrf.0.0700219. PMid:6694140.

Jones R. Plasma membrane structure and remodelling during sperm maturation in the epididymis. J Reprod Fertil Suppl. 1998;53:73-84. PMid:10645268.

Madeddu M, Berlinguer F, Ledda M, Leoni GG, Satta V, Succu S, Rotta A, Pasciu V, Zinellu A, Muzzeddu M, Carru C, Naitana S. Ejaculate collection efficiency and post-thaw semen quality in wild-caught griffon vultures from the sardinian population. Reprod Biol Endocrinol. 2009;7:18. http://dx.doi.org/10.1186/1477-7827-7-18. PMid:19228408.

Matson P, Kappelle W, Malecki I. The use of a hypo-osmotic swelling (HOS) test on sperm of the pig (Sus scrofa domesticus), emu (Dromaius novaehollandiae), asian elephant (Elephas maximus), hamadryas baboon (Papio hamadryas hamadryas), and central rock rat (Zyzomys pedunculatus). Reprod Biol. 2009;9(2):181-87. http://dx.doi.org/10.1016/S1642-431X(12)60025-4.

Morton BE, Sagadraca R, Fraser C. Sperm motility within the mammalian epididymis: species variation and correlation with free calcium levels in epididymal plasma. Fertil Steril. 1978;29(6):695-8. http://dx.doi.org/10.1016/S0015-0282(16)43348-0. PMid:658483.

Muvhali PT, Bonato M, Malecki IA, Cloete SWP. Mass sperm motility is correlated to sperm motility as measured by Computer-Aided Sperm Analysis (CASA) technology in farmed ostriches. Animals (Basel). 2022;12(9):1104. http://dx.doi.org/10.3390/ani12091104. PMid:35565531.

Nield AP, Nathan R, Enright NJ, Ladd PG, Perry GL. The spatial complexity of seed movement: animal‐generated seed dispersal patterns in fragmented landscapes revealed by animal movement models. J Ecol. 2019;108(2):687-701. http://dx.doi.org/10.1111/1365-2745.13287.

Parizzi RC, Miglino MA, Maia MO, Souza JA, Santos JM, Oliveira MF, Santos TC. Morfologia do ovário da ema (Rhea americana). Pesq Vet Bras. 2007;27(3):89-94. http://dx.doi.org/10.1590/S0100-736X2007000300002.

Peixoto JV. Criopreservação de sêmen e avaliação histológica e funcional do testículo de periquitos australianos (Melopsittacus undulatus Shaw, 1805) [thesis]. Viçosa: Universidade Federal de Viçosa; 2010.

Phillipis DM, Asa CS. Development of spermatozoa in the rhea. Anat Rec. 1989;223(3):276-82. http://dx.doi.org/10.1002/ar.1092230306. PMid:2923278.

Pollock C, Carpenter JW, Antinoff N. Birds. In: Carpenter JW, editors. Exotic animal formulary. St. Louis, MO: Elservier Saunders; 2005. p. 135-344.

Quintela AT, Oliveira IRS, Souza AO, Silva AR. Water-induced hypo-osmotic test for the evaluation of canine sperm membrane integrity. Anim Reprod. 2010;7:70-4.

Rijsselaere T, Vansoom A, Maes D, Verberckmoes S, Kruif A. Effect of blood admixture on in vitro survival of chilled and frozen-thawed canine spermatozoa. Theriogenology. 2004;61(7-8):1589-02. http://dx.doi.org/10.1016/j.theriogenology.2003.09.008. PMid:15036988.

Sandhu LC, Warters RL, Dethlefsen LA. Fluorescence studies of Hoechst 33342 with supercoiled and relaxed plasmid pBR322 DNA. Cytometry. 1985;6(3):191-4. http://dx.doi.org/10.1002/cyto.990060304. PMid:3996135.

Santiago-Moreno J, Castaño C, Coloma MA, Gómez-Brunet A, Toledano-Díaz A, López-Sebastián A, Campo JL. Use of the hypo-osmotic swelling test and aniline blue staining to improve the evaluation of seasonal sperm variation in native Spanish free- range poultry. Poult Sci. 2009;88(12):2661-9. http://dx.doi.org/10.3382/ps.2008-00542. PMid:19903966.

Santos TC, Sousa JA, Oliveira MF, Santos JM, Parizzi RC, Miglino MA. Morfologia dos órgãos genitais masculinos e da cloaca da ema (Rhea americana americana). Pesq Vet Bras. 2011;31(5):430-40. http://dx.doi.org/10.1590/S0100-736X2011000500011.

Sibley CG, Monroe BL. Birds of the world, version 2. Cincinnati: Thayer Birding Software: 1996.

Słowińska M, Paukszto Ł, Jastrzębski JP, Bukowska J, Kozłowski K, Jankowski J, Ciereszko A. Transcriptome analysis of turkey (Meleagris gallopavo) reproductive tract revealed key pathways regulating spermatogenesis and post-testicular sperm maturation. Poult Sci. 2020;99(11):6094-118. http://dx.doi.org/10.1016/j.psj.2020.07.031. PMid:33142529.

Smith AMJ, Bonato M, Dzama K, Malecki IA, Cloete SWP. Liquid storage of ostrich (Struthio camelus) semen at 5° C through intermediate dilution. Anim Reprod Sci. 2023;249:107148. http://dx.doi.org/10.1016/j.anireprosci.2022.107148. PMid:36621189.

Sood S, Malecki IA, Tawang A, Martin GB. Survival of emu (Dromaius novaehollandiae) sperm preserved at subzero temperatures and different cryoprotectant concentrations. Theriogenology. 2012;78(7):1557-69. http://dx.doi.org/10.1016/j.theriogenology.2012.06.025. PMid:22980087.

Stelzer G, Crosta L, Bürkle M, Krautwald-Junghanns ME. Attempted semen collection using the massage technique and semen analysis in various psittacine species. J Avian Med Surg. 2005;19(1):7-13. http://dx.doi.org/10.1647/1082-6742(2005)019[0007:ASCUTM]2.0.CO;2.

Tartaglione CM, Ritta MN. Prognostic value of spermatological parameters as predictors of in vitro fertility of frozen-tawed bull semen. Theriogenology. 2004;62(7):1245-52. http://dx.doi.org/10.1016/j.theriogenology.2004.01.012. PMid:15325551.

Tingari MD. On the structure of the epididymal region and ductus deferens of the domestic fowl (Gallus domesticus). J Anat. 1971;109(Pt 3):423-35. PMid:4949288.

Vazquez JM, Martinez EA, Martinez P, Garcia-Artiga C, Roca J. Hypoosmotic swelling of boar spermatozoa compared to other methods for analyzing the sperm membrane. Theriogenology. 1997;47(4):913-22. http://dx.doi.org/10.1016/S0093-691X(97)00046-0. PMid:16728040.

Verstegen J, Iguer-Ouada M, Onclin K. Computer assisted semen analyzers in andrology research and veterinary practice. Theriogenology. 2002;57(1):149-79. http://dx.doi.org/10.1016/S0093-691X(01)00664-1. PMid:11775967.
 


Submitted date:
07/14/2023

Accepted date:
12/06/2023

659d36e2a95395140d54c243 animreprod Articles
Links & Downloads

Anim Reprod

Share this page
Page Sections