机构地区:[1]华中科技大学同济医学院公共卫生学院环境医学研究所教育部环境与健康重点实验室,武汉430030 [2]华中科技大学同济医学院公共卫生学院护理学系,武汉430030
出 处:《中华预防医学杂志》2014年第7期622-627,共6页Chinese Journal of Preventive Medicine
基 金:国家自然科学基金(20977938)
摘 要:目的 自天然水体中筛选分离微囊藻毒素-RR(microcystin-RR,MC-RR)产毒藻株并对其培养特征、产毒性质与急性毒性进行初步探讨.方法 采用毛细吸管法从水华样本中分离微囊藻株,经BG11培养基扩大培养,反复分离纯化藻种.用PCR方法筛查微囊藻毒素合成酶基因,再将藻毒素合成酶基因呈阳性的藻株扩大培养,用高效液相色谱(high performance liquid chromatography,HPLC)鉴定其毒素种类及产毒能力;藻细胞起始浓度为1.5×10^5个/ml,光照强度2 000 lx,光暗比12 h∶12 h,(25±1)℃的条件下,监测藻液在680 nm处的吸光度值而测定藻株生长曲线.采用序贯法测定MC-RR对小鼠的经口染毒半数致死量(LD50),并检测染毒后小鼠的血清酶学、脏器系数及脏器病理改变.结果 接种的198份样品中,分离到5株微囊藻毒素合成酶基因检测呈阳性的微囊藻株,其中一株经高效液相色谱检测鉴定以产MC-RR为主;经形态学与系统发育分类初步确定为铜绿微囊藻,该藻株对数生长期为55 ~60 d,整个生长周期为81 d,藻细胞最大浓度为5.52×10^ 7个/ml.小鼠MC-RR的经口染毒LD50为(12.10±1.35)mg/kg;观测不同作用时间的血清酶学等变化,发现丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转氨酶(AST)、碱性磷酸酶(ALP)、乳酸脱氢酶(LDH)与对照组比较差异具有统计学意义[染毒45 min时小鼠ALT值为(157.08±20.38) U/L,AST值为(333.00±68.53)U/L,ALP值为(392.70±89.59) U/L,LDH值为(1 071.13±160.22) U/L;4 h时ALT值为(514.68±156.87) U/L,AST值为(593.15±40.41) U/L,ALP值为(618.55±208.76) U/L,LDH值为(2 281.72±866.67) U/L;对照组ALT值为(40.30±4.89) U/L,AST值为(142.70±26.59)U/L,ALP值为(56.90±11.89)U/L,LDH值为(509.50 ±94.75) U/L;染毒45 min时各血清酶学变化的t值分别为-11.20、-5.77、-7.38、-6.60,染毒4h时各血清酶学变化的t值分别为-6.04、-20.21、-5.35、-4.07,P�Objective Attempting to isolate and cultivate the microcytin-RR-producing cyanobacteria from natural blooms as well as to further investigate some characteristics of their growth and metabolite toxicity.Methods Capillary-pipette method was used to isolate wild Microcystis strains collected from eutrophicated lakes.The isolated strains were cultured in BG11 media at (25 ± 1) ℃,under 2 000 lx illumination of fluorescent light with a light-dark rhythm of 12-12 h.The growth curve was observed by measuring optical density of culture suspension,toxin-related genes and the metabolite toxins were identified separately by PCR and HPLC,and its acute toxicity was carried out by orally administered toxins to Kunming (KM) mice.Results One of five toxigenic strains from 198 collected samples was confirmed to be a MCRR producing blue-green alga by existing two specific toxin-synthesized enzyme genes and showing specific chromatographic peak of the toxin compared with standard MC-RR through both PCR and HPLC methods.The toxic strain was classified as Microcystin aeruginosa by morphologic and phylogenetic tree analysis.The growth length of the strain lasted nearly 81 days with 55-60 days' exponential phase and the maximal concentration of 5.52 × 10^7 cell/ml.The LD50 of the MC-RR to the KM mice ranged from 10.75 mg/kg to 13.45 mg/kg of body weight.As a result of the acute toxicity,the enzymatic indexes in serum such as alanine aminotransferase (ALT),aspartate aminotransferase (AST),alkaline phosphatase (ALP),lactate dehydrogenase(LDH) were significantly higher than those in the control group.The levels of ALT,AST,ALP and LDH in the treated group at 45 min were (157.08 ± 20.38),(333.00 ± 68.53),(392.70 ±89.59) and (1 071.13 ±160.22) U/L respectively,and at 4 h were (514.68±156.87),(593.15 ±40.41),(618.55 ± 208.76) and (2 281.72 ± 866.67) U/L respectively,and meanwhile the values of ALT,AST,ALP and LDH in the control group were (40.30 ±4.89),(142.70 ±26.5
分 类 号:R114[医药卫生—卫生毒理学]
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