Muzaffer TAŞ, Kemal AK, Ümit CİRİT, Süleyman BACINOĞLU, İrfan Kamuran İLERİ

Boğaların potansiyel fertilitelerinin belirlenmesinde yeni bir servikal mukus penetrasyon test tekniği (Payet yöntemi)

Bu çalışmada boğaların potansiyel fertilitelerinin belirlenmesinde kullanılabilecek yeni bir servikal mukus penetrasyon test (CMPT) tekniği geliştirildi.Araştırmanın ilk aşamasında 10 Holstein boğanın dondurulmuş spermaları ile suni tohumlamalar yapıldı. Ortalama non return rate (NRR) değerleri arasında istatistiksel farklılık bulunan ilk 3 ve son 3 sıradaki boğalar sırasıyla yüksek ve düşük fertilite gruplarını oluşturdu ve dondurulmuş spermaları çalışmanın 2. aşamasında (CMPT) kullanıldı. Yaygın olarak kullanılan CMPT yöntemlerinden farklı olarak servikal mukus 0.25 ml’lik şeffaf payetlere çekildi. Daha sonra eritilmiş sperma ile 3 farklı ısı (37, 39 ve 41ºC) ve iki farklı sürede (15 ve 45 dakika) inkübasyona tabi tutuldu. İnkübasyonlardan sonra payetler, sıvı azot buharında dondurularak -20ºC’de saklandı. İnceleme gününde payetlerin uç kısmından itibaren ölçülerek 15–17.5 mm (1. penetrasyon aralığı; PA1), 32.5–35 mm (PA2) ve 50–52.5 mm’ler (PA3) arasındaki 2.5’er mm’lik parçalar kesilerek ayrıldı. Daha sonra kesilen parçalar içerisindeki mukus bir tel vasıtasıyla itilerek donmuş halde lamlar üzerine aktarıldı ve lamel kapatılarak faz-kontrast mikroskop (200x) spermatozoonlar sayıldı.Boğaların bireysel NRR skorları ile PA 3’te saptanan spermatozoon sayıları (P

A new cervical mucus penetration test technique (Straw method for determination of potential fertility of bulls

A new cervical mucus penetration test (CMPT) technique was developed for determination of the potential fertility of bulls in this study.In the first stage of the experiment artificial inseminations were conducted by using the frozen semen of 10 Holstein bulls. The first and last 3 bulls whose mean non-return rates (NRR) scores had significantly difference formed the high and low fertility groups respectively, and their frozen semen were used at the 2nd stage (CMPT) of the study. Differently from the current CMPT methods the cervical mucus was drawn into transparent straws of 0.25 ml. Than incubated with thawed semen at 3 various temperature (37, 39 and 41ºC) and for 2 various periods (15 and 45 min). Following incubations, the straws were frozen at liquid nitrogen vapour and stored at -20ºC. On the examination day, the straws were cut into pieces by measuring the lengths from the open end as following; 15-15.5 mm (1st penetration interval; PA1), 32.5-35 mm (PA2) and 50-52.5 mm (PA3) 2.5 mm each. Frozen mucus portion in each cut straw part is then pushed out on slight glasses by means of a piece of wire and cover glass was placed for spermatozoon count under a phase-contrast microscope (200x).A significant positive correlation was found between the individual NRR scores of bulls and spermatozoon numbers at PA3 (P

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1) Paul G, Stumpf PG., Lloyd T. In vitro penetration of human sperm into bovine cervical mucus: effects of sperm washing and exposure to low temperature. Obstetrics and Gynecology 1985; 65(1): 42–45.
2) Correa JR, Pace MM, Zavos PM. Relationships among frozen-thawed spermcharacteristics assessed via the routine semen analysis, sperm functional tests and fertility of bulls in an artificial insemination program. Theriogenology 1997; 48: 21–731.
3) Graham EF, Schmehl MKL, Nelson DS. Problems with laboratory assays. Proc 8th Tech Conf AI Reprod NAAB 1980; 1–8.
4) Stålhammar EM, Janson L, Philipsson J. The impact of sperm motility on non-return rate in preselected dairy bulls. Reprod Nutr Dev 1994; 34: 37–45.
5) Hirano Y, Shibahara H, Obara H, et al. Relationships between sperm motility characteristics assessed by the computer-aided sperm analysis (CASA) and fertilization rates in vitro. J Assist Reprod Genet 2001;18: 213–8.
6) Eggert-Krause W, Gerhard I, Tilgen W, et al. Clinical Significance of crossed in vitro sperm-cervical mucus penetration test in infertility investigation. Fertil Steril 1989; 52 (6): 1032–1040.
7) Keel BA, Schalue TK. Correlation of the bovine cervical mucus penetration test with human sperm characteristics in 1406 ejaculates. Arch Androl 2000; 44: 109–115.
8) Kremer J. A simple sperm penetration test. Int J Fertil 1965; 10 (3): 209–215.
9) Matousek J, Říha J, Sršeň V, et al. Penetration of cervical mucus and other body fluids by bull sperm in capillary tubes. Anim Reprod Sci 1989; 18: 161–166.
10) Galli A, Basetti M, Balduzzi D, et al. Frozen bovine semen quality and bovine cervical mucus penetration test. Theriogenology 1991; 35 (4, abst.): 837–844.
11) Anilkumar R, Devanathan TG, Pattabiraman SR, et al. Correlation between the spermatozoal characteristics and sperm penetration distance in polyacrylamide gel and bovine cervical mucus. Theriogenology 2001; 55: 685–691.
12) Yogev L, Gamzu R, Paz G., et al. Improvement in the cervical mucus penetration test by using standart sperm control. Arch Androl 1999; 43: 253–257.
13) Pandya IJ, Mortimer D, Sawers RSA. Standardized approach for evaluating the penetration of human spermatozoa into cervical mucus in vitro. Fertil Steril 1986; 45 (3): 357-365.
14) Alexander NJ. Evaluation of male infertility with an in vitro cervical mucus penetration test. Fertil Steril 1981; 36: 201–
15) Hull MGR, McLeod FN, Joyce DN, et al. Human in vitro fertilization, in vivo sperm penetration of cervical mucus and unexplained infertility. Lancet 1984; 2: 245–248.
16) Verberckmoes S, Van Soom A, De Pauw I, et al. Migration of bovine spermatozoa in Synthetic medium and its relation to in vivo bull fertility. Theriogenology 2002; 58: 1027– 1037.
17) Anilkumar R, Devanathan TG. Correlation between white side test for quality of cervıcal mucus and sperm penetration test. Indian Vet J 1996; 73: 1099-1100.
18) Okuda K, Murase S, Sato K, et al. Penetration ability of bull spermatozoa into bovine cervical mucus. 11th International Congress on Animal Reproduction and Artificial Insemination 1988; 3: 279–280.
19) Herman HA, Madden FW. Evaluation of semen-incubation test. In: The artificial insemination of dairy and beef cattle (including techniques for goats, sheep, horses and swine), Fourth Edition. Sherman, Texas, 1972.
20) Fiser PS, Hansen C, Underhill L, et al. New thermal stress test to assess the viability of cryopreserved boar sperm. Cryobiology 1991; 28: 454–459.
21) Tardif S, Laforest JP, Cormier N, et al. The importance of porcine sperm parameters on fertility in vivo. Theriogenology 1999; 52: 447–458.
22) Doğan, İ. Döl Tutmayan inek ve düvelerde penetrasyon testinin kullanım olanakları. Uludağ Üniversitesi Sağlık Bilimleri Enstitüsü, Doktora Tezi, Bursa 1994.
23) Dabas VPS, Maurya SN. A field method of collection of bovine cervical mucus for microbiological studies. Indian J Anim Reprod 1988; 9: 138–139.
24) Hallap T, Hård M, Jaakma Ü, et al. Does cleansing of frozen-thawed bull semen before assessment provide samples that relate better to potential fertility. Theriogenology 2004; 62: 702–713.
25) Saacke RG. Components of semen quality. J Anim Sci 1982; 55 (Suppl 2):1–13.
26) Cox JF., Martínez C., Lagos S, et al. Sperm migration in cervical mucus in goats. II relationship with colonization of the oviduct and fertilization efficiency. Theriogenology, 1997; 47 (Abst.): 254.