喀斯特深水型湖泊网箱养殖对水体细菌群落结构的影响--以万峰湖为例

贵州省万峰湖（104.5°E至105.1°E,24.7°N至24.9°N）分别于网箱上游1000m处（对照点）以及网箱内、网箱下游200m、400m、600m、800m处设置采样点,每个采样点均采集表层、中层（25m）、底层（50m）水体水样。 利用高通量测序技术研究了水样中 细菌群落多样性,以探究网箱养殖对喀斯特深水型湖泊水体细菌多样性的影响。结果显示：18个水样中共鉴定出细菌门类38个,微生物细菌属699个,其中Flavobacterium（黄杆菌属）和Pseudomonas（假单胞菌属）是万峰湖水体中优势菌属,然网箱底层水体中分布有Lactobacillus（乳杆菌属）、Barnesiella、Shewanella（希瓦氏菌属）、Sphingomonas（鞘氨醇单胞菌）、Shewanella、Sphingomonas等特异性优势菌属。对不同采样点细菌多样性分析表明：万峰湖水体与网箱的水平距离是影响细菌群落结构的关键,影响范围在800m以内。网箱养殖造成了底层水体大量特异性细菌种属的产生。pH、总磷和COD是影响网箱水体细菌群落的关键因子。本研究结果可为湖泊渔业养殖可持续性发展和湖泊生态环境保护提供参考。     

巢湖后生浮游动物群落及不同湖区营养状态评价

为了给巢湖水质管理及富营养化防治提供科学依据，根据水系特征，在巢湖东、西湖区湖心、裕溪河出湖口、丰乐河、派河和南淝河共设置8个采样点，2013年12月至2014年11月共采样12次，分别代表冬季(12月-次年2月)、春季(3-5月)、夏季(6-8月)和秋季(9-11月)，对巢湖水体理化因子和后生浮游动物（枝角类、桡足类、轮虫）群落进行逐月周年调查。基于叶绿素(Chl-a)、总磷(TP)、总氮(TN)、化学需氧量(CODMn)、透明度(SD)等水体理化指标，运用综合营养状态指数(TLI)和生物多样性指数，对不同湖区受污染程度和营养状态进行综合评价。结果显示，巢湖共检出后生浮游动物42种（属），优势种矩形龟甲轮虫(Keratella quadrata)、萼花臂尾轮虫(Brachionus calyciflorus)为富营养化水体或有机质较多水体的常见种，不同季节的优势种存在较大差异；总体上，后生浮游动物全年平均丰度371.6 个/L（191.5~600.0 个/L）和全年平均生物量3.0 mg/L（2.0~4.6 mg/L）均呈现西半湖高于东半湖、夏秋季高于冬春季的分布特点；其群落结构也存在显著的季节差异和空间差异，矩形龟甲轮虫、萼花臂尾轮虫等是造成差异的主要种类。水温(WT)和Chl-a是影响枝角类丰度的重要因子，桡足类丰度与WT呈显著正相关，而轮虫丰度受TLI(∑)指数的显著影响。TLI(∑)指数均值为60.33（52.18~66.28），指示巢湖总体处于轻度至中度富营养化状态。Shannon-Wiener多样性指数和Margalef丰富度指数表明，巢湖总体处于中度污染状态，不同湖区水体具有明显的不均衡性。     

网箱养殖对刘家峡水库浮游细菌群落组成及影响因素分析

为了评价网箱养殖对刘家峡水库浮游细菌群落结构和多样性的影响,于2016年10月份对刘家峡水库6个生态位点进行水样采集,采用高通量测序技术分析了浮游细菌群落特征。结果表明,刘家峡水库浮游细菌主要由变形菌门、放线菌门、蓝藻门、拟杆菌门等11个门,ACK-M1-norank、聚球藻属、鞘氨醇单胞菌属、红球菌属、β变形菌属和Pelagibacteraceae-uncultured等73个属组成,浮游细菌丰度和多样性指数由水库上游至下游依次降低,网箱养殖活动使附近水体的理化性质发生变化,并导致浮游细菌多样性降低。聚球藻属、黄杆菌属、放线菌属、Limnohabitans属和不动杆菌属的种类相对丰度受网箱养殖活动影响敏感。冗余分析表明NO-3-N、TP和TN是影响刘家峡水库浮游细菌群落结构的主要水环境因子,共同解释了55.11%的群落变异。     

白洋淀水域夏季细菌群落结构及与环境因子的关系

为了解白洋淀水域细菌多样性,于夏季采集水样进行水质和微生物群落结构分析。水质分析显示,白洋淀平均酸碱值(pH)为7.95,溶解氧(DO)为16.67 mg/L,属于偏碱性湖泊且含氧量丰富。通过Illumina NovaSeq6000高通量测序分析微生物群落结构表明:白洋淀不同采样点Chao1指数、Ace指数均较高,Shannon指数为7.151~8.443,Simpson指数为0.986~0.992,反映出白洋淀各采样点微生物群落多样性丰富、稳定性好,且细菌丰富度和多样性均以光淀(GD)最高。变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)、蓝细菌门(Cyanobacteria)、拟杆菌门(Bacteroidetes)和放线菌门(Actinobacteria)是湖水的优势菌门,丰度之和能达到90%,且以变形菌门(Proteobacteria)为绝对优势菌门,丰度为44.15%~60.18%;在属水平上烧车淀(SCD)和枣林庄(ZLZ)以多核杆属(Polynucleobacter)为第一优势菌属,其余采样点优势菌属为unidentified_Cyanobacteria。CCA(canonial correspondence analysis)分析表明,pH和DO是影响白洋淀细菌群落结构的主要环境因子。     

凡纳滨对虾不同养殖密度高位池水体细菌群落动态

通过对养殖水体环境基因组DNA中细菌16S rRNA基因V4–V5区的高通量测序和生物信息学分析,研究了两种养殖密度的凡纳滨对虾(Litopenaeus vannamei)高位池水体中细菌群落在养殖过程中的动态。结果显示,养殖过程中各菌群相对丰度变化明显,细菌多样性随时间逐渐提高,优势菌群为变形菌门(Proteobacteria)、蓝藻门(Cyanobacteria)、拟杆菌门(Bacteroidetes)、放线菌门(Actinobacteria)和浮霉菌门(Planctomycetes)。随着养殖时间增长,蓝藻丰度所占比例逐渐减少,而变形菌、拟杆菌和浮霉菌丰度逐渐增大,同时养殖前期高密度池浮霉菌丰度显著高于低密度池(P0.01),而其他菌群无显著差异。结果表明,养殖期前50 d不同养殖密度水体细菌群落差异较大,而后30 d内细菌群落的时间异质性大于空间异质性,这意味着高位池水体菌相被划分为两类,到养殖后期菌相快速转变,养殖密度所带来的影响被减弱。     

海浪河流域河夹村段细菌群落数量时空分布

本文于2009至2010年对海浪河河夹村段水体及底泥中细菌组成与时1空分布进行研究与分析,结果表明,该流域分离出的细菌隶属于5个细菌菌属,其中微球菌属（Micrococcus）是优势菌属,其次是假单胞菌属（Pseudomonas）和动胶菌属（Zoogloeaitzigsohn）。在时间分布上,水样细菌群落数量最高值出现在2009年5月份,最小值出现在2010年1月份,泥样细菌群落数量最高值出现在2009年5月份,最小值出现在2009年7月份。在空间分布上,水样和泥样细菌群落数量总数在8、9、10号样点高于其它样点一个数量级,但各样点细菌菌属没有明显的差异。     

斑点叉尾鮰养殖池塘底泥微生物群落结构特征及其影响因素

微生物群落在水产养殖环境中起着重要作用,深入了解微生物群落组成及其构建机制对了解池塘生态功能具有重要意义。本研究通过高通量测序技术研究斑点叉尾鮰（）养殖池塘底泥中细菌群落机构特征,并分析了细菌群落的主要影响因素。结果显示,斑点叉尾鮰养殖池塘底泥细菌群落结构呈现出季节性变化,冬春季样品相似性较高。斑点叉尾鮰养殖池塘底泥优势菌群主要为变形菌门（Proteobacteria）、绿弯菌门（Chloroflexi）、拟杆菌门（Bacteroidetes）、厚壁菌门（Firmicutes）和酸杆菌门（Acidobacteria）,与池塘水体细菌群落间有明显的差别。线性判别分析显示,冬、春和秋季的特异性优势菌群较为丰富,冬季和春季分布较为集中,分别集中于浮霉菌、拟杆菌门、绿弯菌门和厚壁菌门、拟杆菌门,秋季特异性优势菌群分布特别分散,而夏季优势细菌类群最少,只有绿弯菌门厌氧绳菌属1个属具有显著优势。环境因子中,透明度和总溶解性悬浮物与斑点叉尾鮰养殖池塘底泥细菌群落结构有显著相关（<0.05）,对细菌群落结构的影响也最大。本研究旨为斑点叉尾鮰池塘微生物群落的变化和调控提供借鉴。     

工厂化循环水养殖系统微生物群落结构和功能分析

为比较循环水系统中各水处理单元细菌群落组成、物种多样性和功能的差异,通过高通量测序技术分析该系统养殖池、沉淀池、蛋白分离池、生物滤池和海马齿浮床池水体中的菌群组成,并进行菌群功能预测。结果显示,与其他池相比,生物滤池和海马齿浮床池中α-变形菌纲显著减少,拟杆菌门明显增高。疣微菌门在生物滤池中的相对丰度显著高于其他池。从属的水平分析,优势菌Loktanella在沉淀池和蛋白分离池的占比是其他池的2～3倍,Rubritalea在生物滤池的占比明显高于其他池。海马齿浮床池菌群丰度和菌群多样性均最高,生物滤池菌群丰度最低。在群落组成和结构方面,生物滤池和海马齿浮床池与其他样品相似度最低。PICRUSt功能预测结果显示,该循环水系统中细菌群落占优势的功能类群主要集中在新陈代谢方面。比较各单元细菌群落功能相似性发现,沉淀池和蛋白分离池较相似,生物滤池则与海马齿浮床池类似。另外氮代谢功能基因的相对丰度在海马齿浮床池中更高。该试验结果可为构建适合于中国南方地区的循环水养殖系统提供数据支撑和技术支持。     

巢湖流域河湖系统水环境因子分布特征及营养状态评价

为解析巢湖流域水环境因子空间分布特征，监测了巢湖及其环湖河流33个采样点水环境因子，并采用水质指数（WQI）对其营养状态进行了评价。结果表明：（1）巢湖流域河湖系统巢湖与河流之间及河流与河流之间所有水质参数（除水温外）和WQI指数均存在显著性差异，巢湖流域河湖系统水环境在营养梯度上呈现明显的空间格局，表现在东巢湖水质优于西巢湖；杭埠河、柘皋河、裕溪河水质优于兆河、白石山河、派河、南淝河；依据WQI指数，东巢湖、西巢湖、杭埠河、柘皋河、裕溪河水质为较好，兆河、白石山河、派河、南淝河水质为中等；（2）相较以往研究，本研究中西巢湖水质无明显变化，东巢湖水质略有下降，白石山河和兆河由原来的清水型河流转变为污染严重型河流，污染程度加重，其余环湖河流水质无明显变化；（3）应加强巢湖入湖河流氮源的管理和控制，并加强对小型流域的持续监测，深入了解小型支流在整个营养物质输送中所起的作用。     

温度对拟穴青蟹循环水养殖系统微生物群落结构的影响

为探究温度对拟穴青蟹(Scylla paramamosain)循环养殖水体中微生物群落的影响,深入理解环境因子与养殖水体微生物群落之间的相关性。在水温分别为18℃、22℃、26℃、30℃和32℃的循环养殖系统中进行拟穴青蟹养殖,采用16S rRNA高通量测序技术对不同温度下的养殖水体微生物群落组成进行分析。结果表明:(1) Alpha多样性指数显示5组水样中细菌群落多样性指数大小为t26t22t30t32t18,物种丰度大小为t22t26t32t30t18。温度可以导致养殖水体中微生物群落多样性发生变化,但并未发现二者之间直接的相关性。(2) 5种温度下的养殖系统中细菌群落共有31门、43纲、94目、185科和351属,变形菌门(Proteobacteria)为循环养殖水体中绝对优势菌门。在养殖系统水温为26℃时,养殖水体中氮磷污染物含量相对较高,其水中脱氮除磷菌属的相对丰度也显著高于其他4组。(3)由花瓣图和UPGMA聚类分析图进行相似性分析可知,水体微生物群落对温度的变化有着良好的响应, 5组水样的细菌群落结构总体相似性较低,其中最高温度32℃和最低温度18℃的水体中细菌群落结构最为相似。(4)水体中微生物群落结构和与环境因子的相关性分析显示,氮磷营养盐中的总磷(TP)和硝酸盐(NO3–-N)对水中微生物群落特征具有显著性影响,而温度是导致养殖水体中氮磷营养盐差异化的主要原因。上述研究结果初步解释了温度对拟穴青蟹循环养殖水微生物群落的作用机制,为下一步深入研究拟穴青蟹循环水养殖系统中环境变化与微生物的相互关系提供参考。     

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).     

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.     

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.     

Influence of the antibacterial herbs, Solanum trilobatum, Andrographis paniculata and Psoralea corylifolia on the survival, growth and bacterial load of Penaeus monodon post larvae

The indiscriminate use of antibiotics and chemicals in shrimp hatcheries has led to biomagnification and that in turn could lead to rejection of a whole consignment. The application of the bioencapsulation technique as a tool for curative treatment in shrimp larvae was investigated. Herbs having antibacterial properties such as Solanum trilobatum, Andrographis paniculata and Psoralea corylifolia (methanolic extracts) were bioencapsulated in Artemia and fed to Penaeus monodon post larvae PL 1–25. The post larvae were reared in a medium inoculated with pathogenic bacteria such as Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella typhi and Vibrio sp. Post larvae reared in the non-inoculated water and fed with non-enriched Artemia exhibited 90% survival, highest specific growth rate (12.43%) and reduced bacterial load. P. monodon reared in the bacterial inoculated water and fed with the non-enriched Artemia exhibited the lowest survival (10–30%), specific growth rate (8.42–9.1%) and increased bacterial load (2.86 × 10³ to 3.76 × 10⁵ cfu/g). The methanolic extracts of the herbs helped to increase survival and specific growth rate and reduced bacterial load in the P. monodon culture system. Among the three herbal extracts, P. corylifolia enriched Artemia fed post larvae showed the tendency to higher survival (>50%), growth rate (11.5 averaged) and low bacterial load (1.12 × 10⁵ cfu/g). This revised version was published online in August 2006 with corrections to the Cover Date.