Animal Reproduction (AR)
Animal Reproduction (AR)

Evaluation of fertilization capability of frozen-thawed completely immotile spermatozoa collected from a white bengal tiger after interspecific ICSI with bovine oocytes

Hai-Jun Liu; Jing-Hua Ma; Ru-Ming Liu; Zhang-Guo Liu; Hai-Jun Huang; Jian-Qiang Zou; Jian-Xun Liu; Xian-Fu Zhang

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The objective of this study was to evaluate the fertilization capability of White Bengal Tiger frozen-thawed completely immotile spermatozoa after interspecific intracytoplasmic sperm injection (ICSI) with bovine oocytes. The fertilization status of presumptive zygotes was assessed 18 h after ICSI by immunofluorescence staining and confocal microscopy. The fertilization rate was 34.8% (8/23), as confirmed by the extrusion of two polar bodies, or male and female pronuclei formation. For unfertilized oocytes (65.2%, 15/23), one activated oocyte had an activated spermatozoon but most were unactivated oocytes with unactivated spermatozoa (1/15, 6.7% vs 10/15, 66.7%, respectively, p < 0.05). These results showed that White Bengal Tiger frozen-thawed completely immotile spermatozoa retained the capacity to fertilize bovine oocytes after interspecific ICSI. This is the first report of in vitro produced zygotes using tiger immotile sperm with bovine oocytes by interspecific ICSI technique, which provides an efficient and feasible method for preservation and utilization of endangered feline animals.


White Bengal Tiger, cryopreservation, immotile spermatozoa, interspecific ICSI, bovine oocytes


Buarpung S, Tharasanit T, Comizzoli P, Techakumphu M. Feline spermatozoa from fresh and cryopreserved testicular tissues have comparable ability to fertilize matured oocytes and sustain the embryo development after intracytoplasmic sperm injection. Theriogenology. 2013;79(1):149-58. PMid:23138024.

Chen H, Feng G, Zhang B, Zhou H, Wang C, Shu J, Gan X, Lin R, Huang D, Huang Y. A new insight into male fertility preservation for patients with completely immotile spermatozoa. Reprod Biol Endocrinol. 2017;15(1):74. PMid:28923067.

Donoghue AM, Johnston LA, Seal US, Armstrong DL, Simmons LG, Gross T, Tilson RL, Wildt DE. Ability of thawed tiger (Panthera tigris) spermatozoa to fertilize conspecific eggs and bind and penetrate domestic cat eggs in vitro. J Reprod Fertil. 1992;96(2):555-64. PMid:1339836.

Donoghue AM, Johnston LA, Seal US, Armstrong DL, Tilson RL, Wolf P, Petrini K, Simmons LG, Gross T, Wildt DE. In vitro fertilization and embryo development in vitro and in vivo in the tiger (Panthera tigris). Biol Reprod. 1990;43(5):972-80. PMid:2291909.

Gambini A, Duque Rodríguez M, Rodríguez MB, Briski O, Flores Bragulat AP, Demergassi N, Losinno L, Salamone DF. Horse ooplasm supports in vitro preimplantation development of zebra ICSI and SCNT embryos without compromising YAP1 and SOX2 expression pattern. PLoS One. 2020;15(9):e0238948. PMid:32915925.

Geber S, Lemgruber M, Taitson PF, Valle M, Sampaio M. Birth of healthy twins after intracytoplasmic sperm injection using ejaculated immotile spermatozoa from a patient with Kartagener’s syndrome. Andrologia. 2012;44(Suppl. 1):42-4.

Goto K, Kinoshita A, Takuma Y, Ogawa K. Fertilization of bovine oocytes by the injection of immobilized, killed spermatozoa. Vet Rec. 1990b;127(21):517-20. PMid:2281585.

Kimura Y, Yanagimachi R, Kuretake S, Bortkiewicz H, Perry ACF, Yanagimachi H. Analysis of mouse oocyte activation suggests the involvement of sperm perinuclear material. Biol Reprod. 1998;58(6):1407-15. PMid:9623599.

Lin F, Huang CJ, Liu CS, Guo LL, Liu G, Liu HJ. Laminin-111 inhibits bovine fertilization but improves embryonic development in vitro, and receptor integrin-beta1 is involved in sperm-oocyte binding. Reprod Domest Anim. 2016;51(5):638-48. PMid:27491353.

Liu H, Liu R. Dynamic changes in chromatin and microtubules at the first cell cycle in SCNT or IVF goat embryos. Cell Biol Int. 2018;42(10):1401-9. PMid:29993158.

Moro LN, Sestelo AJ, Salamone DF. Evaluation of cheetah and leopard spermatozoa developmental capability after interspecific ICSI with domestic cat oocytes. Reprod Domest Anim. 2014;49(4):693-700. PMid:24966115.

Nordhoff V, Schuring AN, Krallmann C, Zitzmann M, Schlatt S, Kiesel L, Kliesch S. Optimizing TESE-ICSI by laser-assisted selection of immotile spermatozoa and polarization microscopy for selection of oocytes. Andrology. 2013;1(1):67-74. PMid:23258632.

Parrington J, Swann K, Shevchenko VI, Sesay AK, Lai FA. Calcium oscillations in mammalian eggs triggered by a soluble sperm protein. Nature. 1996;379(6563):364-8. PMid:8552195.

Suzuki R, Okada M, Nagai H, Kobayashi J, Sugimura S. Morphokinetic analysis of pronuclei using time-lapse cinematography in bovine zygotes. Theriogenology. 2021;166:55-63. PMid:33689928.

Wakayama T, Uehara T, Hayashi Y, Yanagimachi R. The response of mouse oocytes injected with sea urchin spermatozoa. Zygote. 1997;5(3):229-34. PMid:9460907.

Wayan Kurniani Karja N, Fahrudin M, Setiadi MA, Tumbelaka LI, Sudarwati R, Hastuti YT, Mulia BH, Widianti A, Sultan K, Terazono T, Namula Z, Taniguchi M, Tanihara F, Takemoto T, Kikuchi K, Sato Y, Otoi T. Characteristics and fertility of Sumatran tiger spermatozoa cryopreserved with different sugars. Cryo Lett. 2016;37(4):264-71. PMid:27925009.

Zahmel J, Jänsch S, Jewgenow K, Sandgreen DM, Skalborg Simonsen K, Colombo M. Maturation and fertilization of African lion (Panthera leo) oocytes after vitrification. Cryobiology. 2021;98:146-51. PMid:33248046.

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