金鱼致病菌的分离鉴定与药敏特性

采用常规方法对患病金鱼(Carassius auratus)内脏组织(肝脏、肾脏、性腺)中致病菌进行了分离鉴定与药敏特性研究。结果显示:实验共分离了8株优势菌,经人工回归感染实验证实其中3株细菌(菌株LK-T、PL-R和PK-T)具有致病性。ATB细菌鉴定仪鉴定结果显示,菌株LK-T和菌株PL-R为温和气单胞菌(Aeromonas so-bria),菌株PK-T为豚鼠气单胞菌(Aeromonas punctata subsp.caviae)。这3株致病菌的主要特征为:革兰氏染色阴性,杆状,对奈替米星、头孢呋肟、头孢拉啶、庆大霉素、环丙沙星、新霉素、先锋必等药物均高度敏感。其中庆大霉素、环丙沙星、新霉素等常规渔药可作为防治用药的依据。     

牛蛙背部肿胀病的病原菌分离与鉴定及药敏实验

在广东广州番禺某牛蛙养殖场发现较为特殊的疾病且传染率和死亡率较高,为查明其致病原,从患病牛蛙(Rana catesbeiana)中抽取其背部的积液和对其他病变部位如肝、脾等进行细菌的分离纯化后得到两株优势菌,然后将两株优势菌进行16S rRNA基因测序、进化树构建、革兰氏染色细菌和生理生化实验进行鉴定和分析,结果显示一株菌株GPY923为嗜水气单胞菌(Aeromonas hydrophila),另一株菌株GPY924为海豚链球菌(Streptococcus iniae),进行人工感染后表明其具有较强的致病性。药敏试验结果显示嗜水气单胞菌对丁胺卡那、红霉素、头孢他啶、头孢呋辛、青霉素和氟苯尼考敏感,对多种药物耐药;海豚链球菌对林可霉素和头孢呋辛耐药,对头孢他啶中度敏感,其余药物均敏感。     

革胡子鲶出血性败血症病原菌的分离鉴定

从暴发性死亡的革胡子鲶(Clarias gariepinus)中分离出可疑病原菌株2010111403(简称1403),分别采用细菌全细胞脂肪酸鉴定系统、Biolog微生物自动鉴定系统及细菌16 S rDNA序列分析三种方法对其鉴定,结果表明菌株1403为嗜水气单胞菌(Aeromonas hydrophila)。分离菌株对健康革胡子鲶致病性测试表明其对革胡子鲶的半数致死剂量(LD50)为6.32×106菌落形成单位(CFU);实验感染革胡子鲶出现与自然发病相似症状,表明菌株1403是引起革胡子鲶发生暴发性死亡的主要致病病原。药敏试验表明,庆大霉素、四环素、阿奇霉素、氟哌酸、先锋霉素等9种抗生素对分离菌株有较强的抑制作用,但分离菌株对复方新诺明、克林霉素、罗红霉素等4种药物表现出耐药性。     

草鱼和银鲫肠道产消化酶细菌的研究

检测了分别从草鱼(Ctenopharyngodon idellus)和银鲫(Carassius auratus gibelio)肠道中分离的180株细菌的蛋白酶、脂肪酶、淀粉酶和纤维素酶的产酶能力。结果显示,两种鱼肠道内可分泌胞外消化酶的细菌包括Aero-monas(气单胞菌属,Aer.)、Vibrio(弧菌属,Vib.)、Bacillus(芽孢杆菌属,Bac.)、Pseudomonas(假单胞菌属,Pse.)四个种属的细菌,Aer.在其中占主要优势,45.71%的Aer.可分泌胞外消化酶。草鱼可分泌上述四种胞外消化酶的菌株共有33株,占肠道菌总数的36.67%;银鲫43株,占47.78%。产酶菌的分布上,草鱼中肠内产消化酶细菌数量显著多于前肠和后肠(P<0.05),前、中、后肠分别是6株、20株和7株;银鲫中肠和后肠数量差异不显著,前肠分布最少。草鱼分泌蛋白酶、脂肪酶、淀粉酶和纤维素酶的菌株分别有21株(23.33%)、10株(11.11%)、30株(33.33%)和16株(17.78%)。银鲫肠道内未检测到可分泌纤维素酶的细菌,蛋白酶、脂肪酶和淀粉酶菌株的数量分别是21株、37株和17株。可见鱼类肠道细菌对食饵消化有重要作用。     

天峡红鮰出血病的病原及其防治研究

从患出血病的天峡红蛔进行了病原菌的分离得出2株细菌,经过人工感染实验确立H-I菌株为致病菌株,并对该菌形态培养特性,生理生化鉴定,确定该菌为豚鼠产气单胞菌.药敏试验该菌对呋哺妥因,氯霉素、丁胺卡那最为敏感,经过生产防治试验取得了较好的效果.     

两株光合细菌的分离鉴定及其水质净化效果研究

为了筛选能够高效净化水质的光合细菌菌株,采用半固体试管法和双层平板法,从市售光合菌菌液中分离得到2株光合细菌,分别命名为GHJ-1、GHJ-2,对菌株进行了形态学观察、生理生化及16S rDNA鉴定,比较了这2株光合细菌对青鱼(Mylopharyngodon piceus)养殖污水的净化效果。结果表明:经鉴定这两株光合细菌为Rhodovulum sp菌株。菌株GHJ-1、GHJ-2对污水进行15 d的净化处理后,硝酸盐去除率分别为91.4%和96.3%;亚硝酸盐的去除率分别为68.9%和82.7%;氨氮去除率分别为56.8%和67.5%;COD Mn的去除率分别为61.7%和42.4%。     

养殖牙鲆致病性弧菌的鉴定及药敏分析

2020年丹东东港市某养殖厂养殖1龄牙鲆(Paralichthys olivaceus)突发病害,表现为腹水病症状。取患病的5尾养殖牙鲆的鳃、肝、脾、肠道、腹水为材料进行细菌分离及纯培养,选择优势菌株通过全自动细菌分析仪及16S rDNA序列分析对该菌株进行鉴定,结果表明分离纯培养的14株菌(DS200401-DS200414)共鉴定出2种,其中1种为该病的致病菌:弧菌属灿烂弧菌(Vibrio splendidus)。随后进行人工感染试验和药敏试验。用12种抗菌药物对3种菌进行药敏试验,结果显示在不同菌株间无明显的敏感与耐药性差异。     

黄鳝源嗜水气单胞菌DB01分离鉴定及药敏特性研究

从患病的黄鳝(Monopterus albus)体内分离到一株致病菌株DB01,对该菌株进行人工感染、生理生化鉴定及16S r DNA序列分析。结果显示:人工感染黄鳝出现与自然发病相同的症状,表明菌株DB01为发病黄鳝的病原菌。菌株DB01为革兰氏阴性杆菌,无芽孢和荚膜,在兔血平板上能形成明显的β-溶血。序列分析显示,菌株DB01与嗜水气单胞菌(Aeromonas hydrophila)的同源性达99%。药敏试验结果显示,菌株DB01对诺氟沙星、左氧氟沙星、阿奇霉素及氟苯尼考等抗生素高度敏感,五倍子、黄柏、地榆及乌梅等中草药对菌株DB01抑菌作用强。     

黑斑蛙歪头病病原米尔伊丽莎白菌的分离、鉴定及特性分析

为查明湖北石首大垸农场养殖的黑斑蛙（Rana nigromaculata）歪头病病原与致病原理，本研究应用微生物分离技术，从病蛙脑、肝、脾和肾脏组织中分离、纯化病菌，采用形态特征、生理生化鉴定、16S rRNA基因序列分析进行细菌种类鉴定，通过组织病理学方法分析患病蛙病理损伤，及人工回归感染确定其致病性，并利用纸片扩散法进行药物敏感性试验。结果显示：分离菌（BSH3）与米尔伊丽莎白菌（Elizabethkingia miricola）聚为一支，结合该菌的形态学特征和生理生化特征，综合鉴定为米尔伊丽莎白菌。组织病理切片结果表明分离菌（BSH3）会导致蛙脑组织基质水肿，基质中出现空泡和炎性细胞浸润的现象；肝、脾和肾脏组织出现变性、坏死和炎症细胞浸润。人工回归感染试验显示，菌株可使黑斑蛙出现与自然发病蛙相似症状，半数致死浓度为6.3×10⁴ CFU/mL。药敏试验结果表明菌株已呈现多重耐药性，仅对少数检测抗生素（米诺环素、氟苯尼考、利福平和万古霉素）敏感，根据药敏试验结果及农业部水产用药规定，推荐使用氟苯尼考治疗该疾病。     

锦鲤致病性维氏气单胞菌的分离、鉴定及药敏研究

新疆乌鲁木齐本地某养殖场锦鲤(Cyprinus carpio)突然大批死亡,在排除寄生虫感染后,怀疑是由细菌引起。为明确引起锦鲤大批死亡的致病菌,本实验对病鱼肝脏、肠道、肾脏等组织进行细菌分离纯化。通过革兰氏染色、微量生理生化反应及分子生物学鉴定,成功分离出两株菌,命名为CK5和CK9。两株菌都为革兰氏阴性短杆菌,在30℃正常生长,能利用葡萄糖、乳糖、纤维二糖,并且V-P、明胶、水杨苷实验阳性;PCR扩增其gyr B基因并测序,BLAST比对后,两株菌与维氏气单胞菌(Aeromonas veronii)的同源性最高,分别为99.6%和99.0%,在系统发育树上与维氏气单胞菌聚为一枝。病理组织切片显示肝脏脂肪空泡变性,肾小管变性坏死。药敏实验结果表明CK5、CK9对庆大霉素、氧氟沙星、头孢噻肟、氨曲南、诺氟沙星等药物敏感,对万古霉素、克林霉素、苯唑西林耐药。     

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.