宽体金线蛭室内繁殖的初步研究

在土壤温度为24℃～26℃下,采用网箱湿土养殖法对具有生殖环带的宽体金线蛭（Whitmania pigra）进行了繁殖试验,统计了水蛭减重率、成活率和产茧率。在室内温度为26℃～28℃下,采用水箱孵化法孵化宽体金线蛭所产的卵茧,统计卵茧孵化率。结果表明：产茧后的宽体金线蛭平均减重率和成活率分别为43.5±1.41%和87.5±6.36%;宽体金线蛭平均产茧量和卵茧孵化率分别为1.22±0.04枚/条和73.1±0.57%。     

宽体金线蛭对稻草基质产卵适应性的研究

为研究宽体金线蛭对稻草基质的产卵适应性,分别建造以稻草作基质的产卵平台和以土壤作基质的产卵平台以及稻草基质产卵平台和土壤基质产卵平台交错分布的混合型产卵平台,并进行了对比试验。分别统计稻草基质组和土壤基质组中每个平台采集的卵的总数量、总体质量和孵化出苗的卵的数量,计算每尾宽体金线蛭的平均产卵数、卵体质量和孵化率；分别统计混合组中稻草基质产卵平台和土壤基质产卵平台采集的卵的总数。结果显示:稻草基质组和土壤基质组在平均产卵数、卵平均质量和孵化率方面差异不显著；混合组中,稻草基质产卵平台与土壤基质产卵平台的产卵数差异显著,说明宽体金线蛭对稻草基质的产卵适应性良好,相对于土壤基质,宽体金线蛭产卵时对稻草基质存在选择性偏好。     

膨胀蛭石对宽体金线蛭卵孵化的影响

为研究膨胀蛭石对宽体金线蛭卵孵化的影响,在(25±1)℃的恒温条件下,分别用膨胀蛭石和土壤(对照组)作为孵化基质孵化宽体金线蛭卵,并进行了对比试验。试验组和对照组各设5个重复,测量并计算不同基质条件下宽体金线蛭卵的平均孵化期、平均孵化率和孵出幼体的平均体质量。结果显示:膨胀蛭石组中宽体金线蛭卵的平均孵化期、平均孵化率和孵出幼体的平均体质量分别为11.16、11.40、11.08、11.52和11.32 d,98.04%、100%、94.12%、96.08%和96.08%,0.025 6、0.025 9、0.025 4、0.025 2和0.025 7g;土壤组中宽体金线蛭卵的平均孵化期、平均孵化率和孵出幼体的平均体质量分别为12.12、12.26、12.16、12.28和11.94 d,88.24%、92.16%、90.20%、94.12%和90.20%,0.025 1、0.024 8、0.024 5、0.024 7和0.025 0 g。各组对应数据之间均存在显著性差异(P0.05)。试验结果表明:以膨胀蛭石作孵化基质,对宽体金线蛭卵的孵化期、孵化率和孵出幼体的体质量有显著影响,膨胀蛭石适宜作为宽体金线蛭卵的加温孵化基质。     

宽体金线蛭人工繁殖技术初步研究

为研究宽体金线蛭的人工繁殖技术,以野外采捕的宽体金线蛭成体作试验材料进行室外繁殖和室内孵化试验,统计其产茧量、空茧数、孵化率。试验结果:约5 000 kg种蛭共产卵茧351.5 kg;当种蛭投入产茧床时的土壤湿度为60 %时,产茧量达到最大值;当孵化箱的土壤湿度为40 %时,空茧数显著低于湿度为20 %、50 %、70 %的组(P<0.05),孵化率显著高于湿度20 %、50 %、70 %组(P<0.05),空茧数、孵化率与湿度30 %组差异不显著(P>0.05)。结果表明:宽体金线蛭人工繁殖采用室外繁殖、室内孵化的方案可行。当投入产茧床时,60 %的土壤湿度有利于种蛭钻进土壤中产茧,显著提高种蛭的产茧量。孵化箱的土壤湿度控制在30 %~40 %时,孵化率最高。     

人工养殖宽体金线蛭食性偏好初探

为进一步了解宽体金线蛭的食性偏好,在室内养殖条件下,分别用螺(环棱螺)、鸭血、菲牛蛭配合饲料等3种饲料投喂宽体金线蛭,并对其生长情况进行了对比。结果表明,在室内养殖的水温、pH、溶解氧近似的条件下,经过129 d养殖,投喂螺组宽体金线蛭的体质量呈上升趋势,平均体质量增长率为247.12%,平均日增长0.04 g/d;投喂鸭血组和配合饲料组宽体金线蛭的体质量均呈下降趋势,平均体质量增长率分别为-50.71%和-41.71%。试验表明,3种饲料中,螺较适合作为宽体金线蛭的饲料,鸭血和菲牛蛭配合饲料则不适合作为其饲料。     

不同孵化基质和开口饵料对宽体金线蛭生长发育的影响

比较有机孵化基质对宽体金线蛭卵茧孵化效果和不同开口饵料对苗种生长发育的影响。孵化基质分别为土壤、牛粪有机肥、蘑菇菌渣,开口饵料分别为螺蛳、漂螺。试验结果显示,蘑菇菌渣+土壤组卵茧孵化率高达98.89%,显著高于土壤组和蘑菇菌渣组(P<0.05);幼蛭各生长指标显著高于土壤组、蘑菇菌渣组、有机肥组及有机肥+土壤组(P<0.05);混合组幼蛭各生长指标均显著高于未添加土壤组(P<0.05)。5种不同孵化基质孵化率依次为蘑菇菌渣+土壤>土壤>有机肥+土壤>有机肥>蘑菇菌渣。不同开口饵料的试验比较发现,漂螺组幼蛭成活率为72.50%,显著高于螺蛳组和螺蛳+漂螺组(P<0.05),漂螺组和螺蛳+漂螺组幼蛭体质量显著高于螺蛳组(P<0.05)。生长效果依次为漂螺组>螺蛳+漂螺组>螺蛳组。试验结果表明,蘑菇菌渣可以作为宽体金线蛭卵茧孵化基质;无厣结构的漂螺更适宜作宽体金线蛭幼蛭的开口饵料。     

南四湖宽体金线蛭生物学观察及人工繁殖初探

初步观察和了解了宽体金线蛭的形态特征、生活习性、繁殖特性，并进行了人工繁殖试验。在10m^2的面积内，放养种蛭100条，产卵茧200余枚，繁殖幼蛭5000余条。     

水蛭无土产卵技术

<正>水蛭俗称蚂蟥。中药材使用的水蛭种类主要是宽体金线蛭、菲牛蛭和日本医蛭。目前这三种水蛭的人工养殖均已获得初步成功,但仍有很多不完善的地方。仅水蛭繁殖产卵这方面而言,目前水蛭繁殖产卵都是采用仿野生的有土方式,就是在水蛭养殖环境中,用泥土人工筑造高出水面的产卵平台,让水蛭自动钻入产卵平台的泥土中交配、产卵。在水蛭产卵10~20天后,用工具人工挖开平台泥土,取出水蛭产的卵茧,再将卵茧集中孵化。     

放养时间段对宽体金线蛭产茧率的影响

为探究不同放养时间段对宽体金线蛭(Whitmania pigra Whitman)繁殖时产茧率的影响,找到繁殖时最佳需要放养的时间,选用体重规格为20.00 g±2.00 g的宽体金线蛭亲本,按5 d间隔设放养时间不同的7个试验组,其放养时间分别为10 d、15 d、20 d、25 d、30 d、35 d、40 d,将亲本放养于聚乙烯泡沫箱中,观察其繁殖产茧及孵化情况。结果表明:30 d及35 d组产茧率最高,分别为11.71%和14.63%;10 d组产茧率最低,为3.97%;40 d组孵化率最低为95.38%;其余组都为100%。因此,放养繁殖时间在30~35 d时产茧率和孵化率较好,经济效益最佳。     

宽体金线蛭苗种培育试验

正宽体金线蛭卵茧孵化出苗1～2 d后开口摄食,摄食方式是将头部钻入幼螺体内吸食汁液。因宽体金线蛭苗特殊的形态和运动方式,其苗种培育方式与鱼苗不同。目前主要采用聚乙烯网布制作育苗箱,因其体积小,环境易污染及换水次数频繁等原因,蛭苗易发生应激反应而死亡,且苗种培育过程中网箱壁遇水后易发生大范围逃苗。营造适合水蛭苗种生长的养殖环境和易于收集苗种的育苗设施是宽体金线蛭苗种培育成功的关键。在2015—2017年实施江苏省水产三新工程"宽体金线蛭高效养殖及苗种繁育技术集成与示范"项目过程中,对宽体金线蛭苗种培育技术做了一些探讨。     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Evaluation of a Fast Method Based on the Presence of Two Restriction Sites in the Mitochondrial ND5 (mt ND5) Gene for the Identification of Scomber Species

The purpose of this work was to evaluate the suitability of a method based on the presence of two restriction sites (for Hae III and Hindf I) in the mitochondrial NADH dehydrogenase subunit 5 (mt ND5) gene to identify Scomber species. The evaluation was performed on 144 reference and market samples by sequencing of the entire 505-bp fragment of the mt ND5 gene and of a 464-bp fragment of the Kocher fragment of the cytochrome b gene (mt Cytb). Sequence analysis of any of the two fragments allows the identification of each of the four Scomber species, but S. japonicus and S. colias had the same restriction sites at the ND5 amplicon and would not have been differentiated by this analysis. Similarly, loss of the Hae III site in some S. scombrus individuals would have misidentified them as not being Scomber. All the market products were correctly labeled except one acquired in Spain labeled as originating in the Atlantic and containing S. japonicus.     

Nitrogenous waste excretion and accumulation of urea and ammonia inChalcalburnus tarichi (Cyprinidae), endemic to the extremely alkaline Lake Van (Eastern Turkey)

The endemic, anadromous cyprinidChalcalburnus tarichi is the only fish species known to occur in alkaline Lake Van (Eastern Anatolia, Turkey). EightC. tarichi were maintained individually in Lake Van water (17 – 19°C; pH 9.8; 153 mEq·I^–1 total alkalinity; 22 total salinity) and tank water samples analyzed for 24 h in 2 to 4 h intervals. At zero time, < 1µM ammonia was present and urea was undetectable in the tank water; at 24 h, total ammonia and urea made up 114±32 and 35±25µM, respectively. Over the experimental period, ammonia-N and urea-N excretion averaged 1041±494 and 607±169moles·kg^–1 fish·h^–1, respectively. The extent of urea excretion was highly variable between specimens. Uric acid excretion was not detectable.Urea was present at high concentrations in all tissues and plasma (25 – 35moles·g^–1·ml^–1) of freshly caughtC. tarichi; total ammonia content of the tissues was by a factor of 1.9 (liver) to 3.0 (brain) lower. High arginase activity (2.4±0.2 U·min^–1·g^–1) was detected in the liver ofC. tarichi but ornithine carbamoylphosphate transferase, a key enzyme of the ornithine-urea-cycle, was absent. Ureagenesis is likely through degradation of arginine and/or uricolysis. High glutamine synthetase activity (11±0.6 U·min^–1·g^–1) and low ammonia content in brain suggest that, like other teleosts,C. tarichi has an efficient ammonia detoxification in the brain, but in no other tissue.Nitrogenous waste excretion at alkaline pH is discussed. The ability ofC. tarichi to excrete high levels of ammonia at extremely alkaline pH is unique among teleosts studied so far. The mechanism of ammonia excretion under Lake Van conditions remains to be elucidated.     

Imported ornamental fish are colonized with antibiotic‐resistant bacteria

There has been growing concern about the overuse of antibiotics in the ornamental fish industry and its possible effect on the increasing drug resistance in both commensal and pathogenic organisms in these fish. The aim of this study was to carry out an assessment of the diversity of bacteria, including pathogens, in ornamental fish species imported into North America and to assess their antibiotic resistance. Kidney samples were collected from 32 freshwater ornamental fish of various species, which arrived to an importing facility in Portland, Oregon from Colombia, Singapore and Florida. Sixty‐four unique bacterial colonies were isolated and identified by PCR using bacterial 16S primers and DNA sequencing. Multiple isolates were identified as bacteria with potential to cause disease in both fish and humans. The antibiotic resistance profile of each isolate was performed for nine different antibiotics. Among them, cefotaxime (16% resistance among isolates) was the antibiotic associated with more activity, while the least active was tetracycline (77% resistant). Knowing information about the diversity of bacteria in imported ornamental fish, as well as the resistance profiles for the bacteria will be useful in more effectively treating clinical infected fish, and also potential zoonoses in the future.     

Effect of iodophor disinfection of non‐hardened Salmo trutta eggs on their bacterial and fungus load

This study investigated the efficiency of iodophor disinfection (135 ppm active iodine for 15–30 min) of non‐hardened Salmo trutta eggs against different groups of bacteria and against fungus. Egg samples were taken from non‐disinfected and from disinfected eggs, microorganisms were cultured on specific nutrient media and their mass was measured by turbidimetric methods. Bacteria and fungus mass of non‐hardened eggs could be reduced but not eliminated by iodophor disinfection with 135 ppm active iodine for 15 min. The extent of reduction was 47–65% (Experiment 1). The efficiency of disinfection increased with disinfection time as the reduction in bacteria and fungus mass was 40–55% after 15 min and 58–74% after 30 min (Experiment 2). Disinfection efficiency of iodophor solution diluted in water (reduction 49–57%) and of iodophor solution diluted in sodium chloride solution iso‐osmolar to the oocytes (reduction 52–61%) was similar (Experiment 3). The reduction in bacteria and fungus mass was persistent as it was 39–72% lower in embryos deriving from disinfected eggs than in embryos deriving from non‐disinfected ones (Experiment 4). In conclusion, the tested disinfection method is inadequate to eliminate pathogens completely but it could positively influence immune defence of eggs and embryos.