副溶血弧菌、脂多糖和嗜酸小球菌对凡纳滨对虾血清一氧化氮合酶的影响

Effects of Vibrio parahaemolyticus, LPS and Pediococcus acidilacticion nitric oxide synthase activity of Litopenaeus vannamei

探讨了凡纳滨对虾Litopenaeus vannamei血细胞中是否存在一氧化氮合酶(NOS)活性,进行了副溶血弧菌Vibrioparahaemolyticus和哈维氏弧菌Vibrioharveyi脂多糖(LPS)体外孵育实验,采用亚硝酸盐法对凡纳滨对虾血细胞中NOS活性进行了测定。结果表明,副溶血弧菌和LPS离体孵育血细胞组的一氧化氮(NO)浓度显著高于对照组,而且这一反应能被NOS抑制剂亚硝基L精氨酸甲酯(LNAME)所阻断,证实了凡纳滨对虾血细胞中存在NOS活性。同时进行了副溶血弧菌感染凡纳滨对虾对其血清NO浓度和NOS活性的影响试验。试验表明,副溶血弧菌注射感染凡纳滨对虾后12h血清中NOS活性极显著高于对照组;NO浓度在注射后24h开始升高,72h后各组NO浓度都显著高于对照组。说明副溶血弧菌感染凡纳滨对虾可以诱导NOS的表达,并产生NO来抵抗副溶血弧菌的入侵,从而增强了对虾的免疫力。此外将嗜酸小球菌Pediococcusacidilactici按10.0mg·kg-1添加到饲料里投喂凡纳滨对虾后,血清中溶菌酶活力和NOS活性比对照组有显著提高,且溶菌酶活力和NOS活性存在显著正相关性(r=0.629,r0.05[1,12]=0.532)。从而进一步说明了NOS活性可以作为凡纳滨对虾免疫力高低的有效指标。

Litopenaeus vannamei haemocytes were stimulated by 100g·ml^-1 bacterial lipopolysaeeharide （LPS） or 5×10^6CFU· ml^-1 Vibrio parahaemolyticus to study whether they have nitric oxide synthase ac tivity or not. The results showed that shrimp haemocytes induced by Vibrio parahaemolyticus or LPS could significantly produce nitric oxide, and this phenomenon could be blocked by inducible nitric oxide synthase （iNOS） reversible inhibitor, N-ω-nitro-L-arginine-methyl ester （L-NAME）, demonstrating that Litopenaeus vannamei haemocytes have NOS activity. In addition, effects of Vibrio parahaemolyticus on nitric oxid concentration and nitric oxide synthase activity of shrimp serum were also studied. The results showed that NOS activity in shrimp serum of the experimental group was significantly higher than that of the control group after 12 hours of post infeetion; and NO concentration in shrimp serum increased slightly after 24 hours of post-infection, and was significantly higher than that of the control group after 72 hours of post-infection. These demonstrated that Vibrio parahaemolyticus infection can stimulate NOS synthesis in Litopenaeus vannamei and produce NO to resist Vibrio parahaemolyticus. Another experiment was conducted to examine the effect of Pediococcus acidilaetiei on the activities of shrimp serum lysozyme and nitric oxide synthase. Four practical diets were formulated to contain 0.0, 10.0, 50.0, and 100.0 mg Pediococcus acidilactici per kg of diet. After 8-week feeding, the activities of lysozyme and NOS with 10.0mg Pediococcus acidilaetici per kg of diet were significantly higher than those of the control group. The activity of lysozyme was correlated positively with the activity of NOS in shrimp serum. The correlation coefficientwas0.629 （r0.05[l,12]=0.532）. Therefore, NOS activity can be regarded as an immune index of Litope naeus vannamei.