三倍体虹鳟鱼（Oncorhynchus mykiss）红细胞改变的观察
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.
本研究的主题是研究二倍体和三倍体虹鳟鱼 （Oncorhynchus mykiss） 的红细胞改变。三倍体虹鳟鱼是通过在受精后 20 分钟对卵施加 26.5 C 的热休克 20 分钟获得的。二倍体没有受到电击。虹鳟鱼3N和2N鱼种在相同条件下（环境、摄食）饲养。通过红细胞大小测量和上皮鳃细胞 （NOR） 中活性核仁的数量来测定鱼倍体水平。热休克处理中三倍体诱导率为80.86%。研究发现，三倍体鱼类中具有分段细胞核的红细胞比例为13.52%，显著高于二倍体鱼类（1.06%）（P< 0.01）。此外，红细胞的有丝分裂仅见于三倍体（0.57%）。看来，核分割的增加是三倍体虹鳟鱼的重要特征之一。
Key words: amitotic cell, red blood cell, rainbow trout, triploid, segmented nuclei
无丝分裂细胞、红细胞、虹鳟鱼、三倍体、分节核
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
通过向受精后20分钟的卵子施加26.5°C的热冲击20分钟，获得了三倍体虹鳟鱼。二倍体(对照组)未接受热冲击，鱼苗在相同营养和环境条件下饲养。鱼的多倍体水平通过红细胞尺寸测量【Perovic等2003】和上皮细胞活性核仁组织染色体的计数【Philips等1986】来确定。从42条三倍体和35条二倍体鱼的尾静脉采集血样，固定在95%的甲醇中3分钟，然后自然风干，用10%的Giemsa溶液染色15分钟。每张玻片上研究100个红细胞，在400*倍显微镜下观察，以确定细胞有丝分裂和细胞出现分段核的频率。在SPSS中使用非参数t检验进行统计评估
