三倍體虹鱒魚（Oncorhynchus mykiss）紅細(xì)胞改變的觀察

Abstract. 

The subject of this study was investigation of red blood cell alterations in diploid and triploid rainbow trout (Oncorhynchus mykiss). The triploid rainbow trout were obtained through the application of heat shock of 26.5°C for 20 min to eggs 20-min following fertilization. The diploids were not subjected to shock. The 3N and 2N fingerlings of rainbow trout were reared under the same condition (environment, feeding). Fish ploidal level was determined by erythrocyte size measurement and the number of active nucleoli in epithelial gill cells (NORs). Triploidy was induced in 80.86% in heat shocked treatment. It was found that the percentage of erythrocytes with segmented nuclei in triploid fish was 13.52% and was significantly higher (P<0.01) than that in diploid fish (1.06%). In addition amitosis of red blood cells only seen in triploids (0.57%). It seems that increase of nuclear segmentation is one of the important characteristics for triploid rainbow trout.

本研究的主題是研究二倍體和三倍體虹鱒魚 （Oncorhynchus mykiss） 的紅細(xì)胞改變。三倍體虹鱒魚是通過在受精后 20 分鐘對卵施加 26.5 C 的熱休克 20 分鐘獲得的。二倍體沒有受到電擊。虹鱒魚3N和2N魚種在相同條件下（環(huán)境、攝食）飼養(yǎng)。通過紅細(xì)胞大小測量和上皮鰓細(xì)胞 （NOR） 中活性核仁的數(shù)量來測定魚倍體水平。熱休克處理中三倍體誘導(dǎo)率為80.86%。研究發(fā)現(xiàn)，三倍體魚類中具有分段細(xì)胞核的紅細(xì)胞比例為13.52%，顯著高于二倍體魚類（1.06%）（P< 0.01）。此外，紅細(xì)胞的有絲分裂僅見于三倍體（0.57%）?？磥恚朔指畹脑黾邮侨扼w虹鱒魚的重要特征之一。

Key words: amitotic cell, red blood cell, rainbow trout, triploid, segmented nuclei

                 無絲分裂細(xì)胞、紅細(xì)胞、虹鱒魚、三倍體、分節(jié)核

INTRODUCTION 

Triploidy induction is an effective way in salmonid fish cultivation. Triploids have three sets of homologous chromosomes in their somatic cells instead of the normal diploid number of two [Smith and Benfey 2001]. This results in the abnormal pairing of chromo somes during meiotic reduction divisions, and the resultant inability of most triploid germ cells to complete gametogenesis and produce euploid gametes. Thus, triploids typically show impaired gonadal development and are sterile and therefore metabolic energy is di verted from reproduction to somatic growth [Benfey 1999]. They have also reduced if not eliminated other problems associated with sexual development. Because of polyploidy induction, some physiological alterations may be resulted in some fish species. Blood cells structure has some differences between diploid and triploid Salmonids.

For example, total blood hemoglobin and mean cellular hemoglobin concentration were lower in triploid Atlantic salmon as compared with diploids [Benfey and Sutterlin 1984]. There was no significant difference in total blood hemoglobin between diploid and triploid brook trout (Salvelinus fontinalis), although total number of erythrocytes was lower and cell size was larger in triploids [Benfey and Biron 2000]. Erythrocyte size was larger in triploid rainbow trout but total erythrocytes and their hemoglobin content was lower as compared with diploids [Suresh and Sheehan 1998]. In this research, some structural abnormalities like erythrocyte amitosis and nuclei segmentation was observed during the examination of blood smears. The objective of this research was accurate determination of these alterations and their probable effect on ery throcyte functions in diploid and triploid rainbow trout (Oncorhynchus mykiss).

MATERIALANDMETHODS 

Triploid rainbow trout were obtained through the application of heat shock of 26.5oC for 20 min to eggs 20-min post fertilization. Diploids (control) received no heat shock, and fingerlings were reared at the same nutritional and environmental conditions. Fish ploidy level was determined by erythrocyte size measurement [Perovic et al. 2003] and by count ing of gill epithelial cells active nucleoli (NORs) [Philips et al. 1986]. Blood samples for the smears were taken from the caudal vein of 42 triploids and 35 diploids, fixed in 95% methanol for 3 minutes, left to air-dry and stained with 10% Giemsa solution for 15 min utes. One hundred erythrocytes per slide were studied at 400* magnification to determine the frequency of cell amitosis and cells with segmented nuclei. Statistical evaluation was carried out in SPSS using non-parametric t-test

通過向受精后20分鐘的卵子施加26.5°C的熱沖擊20分鐘，獲得了三倍體虹鱒魚。二倍體(對照組)未接受熱沖擊，魚苗在相同營養(yǎng)和環(huán)境條件下飼養(yǎng)。魚的多倍體水平通過紅細(xì)胞尺寸測量【Perovic等2003】和上皮細(xì)胞活性核仁組織染色體的計數(shù)【Philips等1986】來確定。從42條三倍體和35條二倍體魚的尾靜脈采集血樣，固定在95%的甲醇中3分鐘，然后自然風(fēng)干，用10%的Giemsa溶液染色15分鐘。每張玻片上研究100個紅細(xì)胞，在400*倍顯微鏡下觀察，以確定細(xì)胞有絲分裂和細(xì)胞出現(xiàn)分段核的頻率。在SPSS中使用非參數(shù)t檢驗進(jìn)行統(tǒng)計評估