Inhibiting fungus on largemouth bass eggs with copper sulfate and its toxicity to fry and juveniles

This study determined the effectiveness of copper sulfate (CuSO₄) to inhibit fungal growth (caused by Saprolegnia spp.) on largemouth bass (LMB) eggs spawned on/in fiber mats in very high-alkalinity/-hardness waters; experiments also determined the toxicity of CuSO₄ to LMB fry and juveniles. An untreated control and three CuSO₄ concentrations (10, 20, and 40 mg/L) were tested under a flow-through scenario in the effectiveness experiment. Eggs were treated daily until hatching began. Fungal load at the time of hatch had a total area > 3.0 cm² in the untreated controls, total area 1.0 to <2.0 cm² in the 10 and 20 mg/L CuSO₄ treatments, and total area > 1.0 cm² in the 40 mg/L CuSO₄ treatments. Fungus samples were identified as Saprolegnia australis. The 24-hr median lethal concentration (LC50) values on the LMB yolk-sac and swim-up fry were 32.0 and 4.6 mg/L CuSO₄, respectively; the No Observed Effect Concentrations (NOEC) were 16.0 and 0.125 mg/L CuSO₄, respectively. Juvenile LMB were extremely tolerant to CuSO₄, and their 24-hr LC50 value was 185.5 mg/L; the NOEC was 64 mg/L. This study indicates that CuSO₄ can be an important resource for hatcheries to control egg fungus, especially in high-alkalinity/-hardness waters.     

5种药物对异育银鲫"中科3号"受精卵孵化的影响

在水温21~23℃,pH 6.2~6.5条件下,采用半静水试验法,按等对数间距设置药物质量浓度梯度,日换水50%,补充药物50%,24 h观察记录1次,比较美婷Ⅱ、霉灵、纳他霉素、杀毒矾和高锰酸钾5种药物对异育银鲫"中科3号"受精卵水霉抑制作用及孵化率的影响。试验结果显示,试验药物对受精卵水霉生长均有一定抑制作用,其抑霉率为美婷Ⅱ>霉灵>高锰酸钾、纳他霉素和杀毒矾,但杀毒矾对异育银鲫"中科3号"受精卵的毒副作用较大,显著降低了受精卵孵化率,不宜使用;美婷Ⅱ、霉灵、纳他霉素和高锰酸钾对异育银鲫"中科3号"受精卵的适宜使用质量浓度分别为4.90~6.50、1.97~3.87、1.50、2.50 mg/L,使用方法为长时间浸浴,可有效抑制受精卵孵化过程中水霉的生长,显著提高受精卵孵化率,适合在异育银鲫"中科3号"受精卵孵化过程中使用。     

水霉属菌(Saprolegnia)分离培养技术研究

 对水霉(Saprolegnia)的分离培养技术进行了探索,筛选出大麻籽、油菜籽、红花籽、苍蝇、蚜虫、小麦粒、玉米片、茶叶、葵花仁9种材料作为诱发水霉菌的诱饵。其中油菜籽能防止腐霉菌(Pythium)污染,是初步纯化菌种、排除杂菌的适宜饵料。同时发现控制温度能使水霉在最短时间内分别产生无性或有性繁殖。在水培条件下,25℃左右适合菌丝及无性器官的发育,有性器官却不能产生,而只有在15~17℃条件下才能使水霉很快进入有性生殖阶段;在加入200mg/L链霉素的CMA培养基上促发有性阶段则需要在25℃左右的温度下进行,方能缩短培养时间。     

血根碱抗水霉菌体外活性研究

采用测OD值和琼脂稀释法,以孔雀石绿、氯化钠作为对照,分别进行了3种药物对水霉菌孢子和菌丝的体外抑菌活性研究,其中血根碱以2%DMSO(二甲亚砜)为溶剂,孔雀石绿和NaCl均以水为溶剂。结果表明:血根碱对水霉孢子的MIC80为200μg/mL,对菌丝的MIC为256μg/mL;孔雀石绿对水霉菌孢子的MIC80为6μg/mL,对菌丝的MIC为12μg/mL;氯化钠对水霉菌孢子的MIC80为15 000μg/mL,对菌丝的MIC为20 000μg/mL,说明血根碱具有开发为抗水霉菌药物应用于渔药领域的前景。     

15种中草药超临界CO_2流体萃取物对2种水霉菌的抑制作用研究

研究了15种中草药超临界CO2流体萃取技术,从中草药粉碎的粒度、超临界CO2流体萃取仪的萃取温度、萃取压力对中草药提取条件进行了优化,获得了15种中草药超临界CO2流体萃取技术方案;将15种中草药超临界CO2流体萃取物添加到PDA培养基中研究了15种中草药萃取物对多子水霉(Saprolegniales ferax)和澳大利亚水霉(Saprolegnia australis)菌丝的抑制作用,结果显示:中草药粉碎成20目时,适宜采用超临界CO2流体萃取仪进行萃取,当超临界CO2流体萃取仪的萃取温度一定的条件下萃取物产率随萃取压力的提高而增加,当萃取压力一定的条件下萃取物产率随萃取温度的增加先增加后降低;15种中草药中川楝子、土槿皮、五倍子、山苍子的萃取物对2种水霉菌菌丝的生长抑制效果相对较好,当萃取物浓度大于200 mg/L时才能较好抑制2种水霉菌菌丝的生长。     

Do fathead minnows,Pimephales promelas Rafinesque,alter their club cell investment in responses to variable risk of infection from Saprolegnia?

Fish in the Superorder Ostariophysi possess large epidermal club cells that release chemical cues warning nearby conspecifics of danger. Despite the long‐held assumption that such club cells evolved under the selective force of predation, recent studies demonstrated that predation has no effect on club cell investment. Rather, club cells have an immune function and cell production may be stimulated by skin‐penetrating pathogens and parasites. The current work investigates whether fathead minnows, Pimephales promelas, alter their club cell characteristics based on variation in infection risk. In a 2 × 3 design, we exposed minnows to infective cysts of two oomycete species (Saprolegnia ferax and S. parasitica) at three different concentrations (2, 20 or 200 cysts L^?1). Club cell characteristics (number and size) were quantified 12 days after exposure. Saprolegnia parasitica is thought to be more pathogenic than S. ferax, hence we predicted greater club cell investment and a larger turnover rate of cells by minnows exposed to S. parasitica than S. ferax. We also predicted that minnows exposed to higher numbers of cysts should invest more in club cells and have a higher turnover rate of cells. We found no difference in club cell density or size between fish exposed to the two Saprolegnia species; however, fish exposed to high concentrations of pathogens had smaller club cells than those exposed to low concentrations, indicating a higher rate of turnover of cells in the epidermis.     

山女鳟水霉病病原的分离鉴定及其生物学特性

用传统方法从患病山女鳟体表病灶部位分离纯化一株致病性水霉菌株,将其命名为SN,并对其生物学特性进行研究。通过观察附着在油菜籽上水霉菌的形态,其菌丝有适量的分支,呈透明管状,无横隔;菌丝顶端产生棒状孢子囊,游动孢子成熟后,从孢子囊中迅速释放,新孢子囊以层出方式或侧生出芽方式发生。在一定的生长条件下,菌丝顶端或中间可见球形或不规则形状的厚垣孢子。通过与Gen Bank基因库中水霉属菌株ITS r DNA序列的同源性比较,确定SN菌株与鲑菌株(Gen Bank登录号EU551152)的亲缘关系最近。因此,结合形态特征及ITS序列鉴定结果,可推测菌株SN为鲑水霉(Saprolegnia salmonis)。研究不同生长条件对菌株的生物学特性的影响,菌株SN的适宜生长温度为10~30℃,最适生长温度为25℃;适宜p H为5~11,最适生长p H为8;适宜盐度为0.5%~2%,最适生长盐度为0.5%。通过分离鉴定山女鳟水霉病致病性病原,并对其生物学特性进行初步研究,为冷水鱼水霉病的防治提供理论依据。     

患病北极红点鲑的病原分离与鉴定

对2004年5月四川省某养殖基地养殖的北极红点鲑患病鲑进行了病理学检查和病原分离,从病灶处分离到细菌和真菌。分离到的细菌经API生化鉴定为杀鲑气单胞菌杀鲑亚种(Aeromonas salmonicidasubsp.salmonicida)。用引物AP1和AP2对纯化后的细菌进行PCR扩增,结果扩增出长度为421 bp的DNA片段,对扩增片段进行克隆、测序,用NCBI-BLASTn在GenBank中搜寻相似序列,结果与杀鲑气单胞菌各株A层蛋白部分编码基因有99%以上的序列同源性。用引物EUS-F和EUS-R对分离到的真菌进行PCR扩增,结果扩增出长度为755 bp的DNA片段,对扩增片段进行克隆、测序,用NCBI-BLASTn在GenBank中搜寻相似序列,结果与水霉属各株的DNA片段(包括18S核糖体RNA基因部分序列、内转录间隔区1全序列、5.8S核糖体RNA基因全序列、内转录间隔区2全序列和28S核糖体RNA部分序列)有93%-100%的同源性;而与属于丝囊霉菌属的EUS各株相对应的DNA片段仅有60%的序列同源性。因此判定所分离的真菌为水霉。杀鲑气单胞菌是引起疖疮病和溃疡病等病的病原菌,而水霉广泛存在于水中,当鱼受伤后,水霉孢子容易从患病部位侵入鱼体,加重感染。因此判断该病主要是由杀鲑气单胞菌引起,而水霉则引起继发感染。     

Morphological and physiological characteristics of Saprolegnia spp. strains pathogenic to Atlantic salmon, Salmo salar L

Seventeen strains of Saprolegnia spp. were examined for morphological and physiological characteristics, and seven were examined for their pathogenicity to Atlantic salmon, Salmo salar L. Two of the Saprolegnia strains tested caused 89 and 31% cumulative mortality in challenged salmonids and were significantly more pathogenic than the other strains tested. The positive control (Saprolegnia parasitica ATCC 90213) caused 18% mortality, but this was not significantly higher than non-pathogenic strains (0-3% cumulative mortality). All the pathogenic Saprolegnia strains and two non-pathogenic strains had secondary cysts with long, hooked hairs, a characteristic which is claimed to be typical of S. parasitica. This characteristic is apparently necessary, but does not in itself determine the ability to cause mortality in Atlantic salmon. However, all the pathogenic Saprolegnia strains in the present study showed a significantly higher initial growth rate of cysts in sterilized tap water than did non-pathogenic strains. The results of the present study suggest that initial growth rate of germinating cysts in pure water, together with the presence of long hooked hairs on the secondary cysts, may be indicators of pathogenicity of Saprolegnia strains to Atlantic salmon.