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1.
Streptococcus iniae and Gyrodactylus niloticus are two common pathogens of cultured Nile tilapia, Oreochromis niloticus. We studied concurrent infection of tilapia by G. niloticus and S. iniae and evaluated whether parasitism in tilapia with Gyrodactylus increased susceptibility and mortality following immersion infection with S. iniae. Results showed that death mainly occurred in fish with G. niloticus and challenged with S. iniae (G-S group). The accumulative mortality (42.2%) was significantly higher in the G-S group than in fish not infected by the parasite (6.7%), but exposed to S. iniae. Bacteriological examination revealed S. iniae from > or =92% of dead or moribund fish challenged with S. iniae. Gyrodactylus not only damaged fish epithelium and provided entry for invasive bacteria but also was found to harbour viable cells of S. iniae for 24 and 72 h. Streptococcus iniae was isolated from 60% and 40% of G. niloticus collected from fish infected by intraperitoneal injection or immersion, respectively, at 24 h post-challenge. The present study confirms that parasitism of tilapia by G. niloticus increased host mortality following exposure to the bacterial pathogen S. iniae.  相似文献   

2.
尼罗罗非鱼(Oreochromis niloticus)是世界水产养殖业中的重要经济鱼类,但在养殖生产中易受海豚链球菌(Streptococcus iniae)感染而致病致死,使用疫苗是一种相对理想的防感染措施。该研究采用海豚链球菌simA、pgmA基因构建的真核表达载体作为DNA疫苗,肌肉注射罗非鱼评估疫苗保护效果。免疫后在DNA和RNA水平上,在注射鱼体内检测到2个目的基因。首次免疫后第7至第28天,鳃、肝、肾脏、头肾中疫苗组的白介素1 (Interleukin,IL-1β)与肿瘤坏死因子(Tumor Necrosis Factor,TNF-α)表达量高于PBS对照组;疫苗组的抗体滴度、血清抗菌活性显著(P<0.05)高于PBS对照组。攻毒后,注射pcDNA3.1-pgmA、pcDNA3.1-simA、pcDNA3.1-pgmA与pcDNA3.1-simA等比例混合疫苗的相对保护率(Relative percent survival,RPS)分别为60.7%、49.9%和75.0%。结果表明所制备的疫苗具有免疫保护效果,可作为候选疫苗。  相似文献   

3.
强俊  杨弘  王辉  徐跑  柒壮林  何杰 《水产学报》2012,36(6):958-968
以吉富罗非鱼、新吉富罗非鱼、埃及尼罗罗非鱼和红罗非鱼为研究对象,饲养100d后,进行海豚链球菌(2.95×108CFU/mL)感染试验,分析攻毒前后各品系罗非鱼的血液生化指标和肝脏HSP70 mRNA表达量的变化规律。另从各桶中取20尾鱼进行同样的攻毒试验,统计攻毒后各时间点的累积死亡率。结果表明,感染海豚链球菌96h后,吉富罗非鱼和新吉富罗非鱼对病原较为敏感,累积死亡率分别达到36.67%和38.33%;埃及尼罗罗非鱼对病原敏感性较差,试验期间未见死亡。吉富罗非鱼、新吉富罗非鱼和红罗非鱼血清皮质醇和葡萄糖水平以及肝脏HSP70 mRNA的表达量在攻毒后明显提高,血清谷丙转氨酶、谷草转氨酶与溶菌酶活力也呈上升趋势,碱性磷酸酶活力与甘油三酯和胆固醇水平低于攻毒前。埃及尼罗罗非鱼可以利用糖原和脂类产生的能量,提高了HSPS与一些特定免疫蛋白(溶菌酶、球蛋白等)的合成,增强了鱼体的非特异性免疫力。罗非鱼选育过程中,需要将抗病力与生长性能进行有效的结合,在注重生长速度的同时也要增强其抗应激能力,从而为罗非鱼产业的可持续发展提供保证。  相似文献   

4.
广东省养殖罗非鱼、海鲈、尖吻鲈海豚链球菌感染调查   总被引：4，自引：0，他引：4  
利用细菌分离培养方法结合特异PCR技术,对广东省珠三角地区养殖罗非鱼(Oreochromis spp.)、海鲈(Lateolabrax japonicus)及尖吻鲈(Lates calcarifer)的海豚链球菌(Streptococcus iniae)感染情况进行了周年调查。每月固定时间在特定养殖区域采集目标鱼的脑、肝、脾、肾和肌肉等组织,并对其进行海豚链球菌的细菌分离培养鉴定。仅从已经患病的尖吻鲈中分离到3株链球菌,经生理生化鉴定和16S rDNA测序确定为海豚链球菌。利用海豚链球菌特异PCR技术对上述养殖鱼类不同组织进行检测,发现罗非鱼、海鲈、尖吻鲈的海豚链球菌感染率分别为30.21%、23.53%、14.55%,其中罗非鱼脑和肌肉的感染率明显较其他组织高(P<0.05),分别为20.65%和23.75%;海鲈的脑部和肌肉感染率也较其他组织高(P<0.05),分别为12.1%和10%;而尖吻鲈各组织感染率没有较大差异(P>0.05)。另外,研究结果还表明采集样本的海豚链球菌感染率随着其体长的增加而呈现下降趋势。  相似文献   

5.
Abstract Channel catfish, Ictalurus punctatus (Rafinesque), were immunized with Ichthyophthirius multifiliis (Ich) theronts and trophonts, and the immune response and host protection against both homologous and heterologous serotypes of Ich were evaluated. Immunizations were done with two immobilization serotypes (ARS4 and ARS6) of live theronts by bath immersion (trial I) and with sonicated trophonts by intraperitoneal (i.p.) injection (trial II). Cutaneous and serum antibody titres against Ich following immunization were measured and survival of catfish was determined after theront challenge. Theronts were immobilized by the antiserum from fish immunized with homologous theronts or trophonts, but not by the serum of fish immunized with the heterologous serotype. Serum from fish immunized by immersion with live theronts showed higher enzyme-linked immunosorbent assay titres against both homologous and heterologous serotypes than fish immunized by i.p. injection of trophonts. Channel catfish immunized by immersion with live theronts or by i.p. injection with sonicated trophonts developed an immune response against Ich and provided cross-protection against challenge from both serotypes (ARS4 and ARS6) of the parasite. Sonicated trophont antigens in aqueous solution by i.p. injection could stimulate an immune response in fish, but the immunity was of short duration.  相似文献   

6.
Passive immunisation of fish was conducted to determine whether anti-Vibrio anguillarum whole sera (AVA) and affinity-purified AVA raised in sheep, rabbits and rainbow trout (Oncorhynchus mykiss) were persistent when injected and orally administered into rainbow trout. These responses were compared with active immunisation by immersion in, and intraperitoneal (i.p.) injection with, formalin-killed V. anguillarum cells. Sheep and rabbit AVA were detected in rainbow trout sera for up to 70 days (half-life 21 days) after i.p. injection as determined by an enzyme-linked immunosorbent assay (ELISA). The relative percentage survival (RPS) of passively immunised rainbow trout challenged with virulent V. anguillarum after an injection was comparable to that of active immunisation by immersion after 1 month post-immunisation (p.i). Affinity-purified sheep and rabbit AVA exhibited the same protective potential as whole serum in rainbow trout. Rabbit and sheep immune sera diluted 1:8 and 1:50, respectively, provided equivalent protection as undiluted fish immune serum. An active immune response against passively acquired heterologous immunoglobulins was demonstrated by ELISA, with responses against sheep AVA being less than those against rabbit AVA. Rainbow trout given purified sheep AVA conjugated to LTB (the GM-1-binding subunit of Escherichia coli heat-labile toxin) and administered orally had an RPS of 37.5% at 15 days and 27% at 1 month p.i. In contrast, fish given sheep AVA conjugated to TraT (an internal membrane of E. coli) or in micellar form with Quil-A had RPSs of only 18.7 and 6.2%, respectively, after 15 days, and 13.3 and 0% after 1 month, respectively. The protection conferred by immune sera was shown to be due to the immunoglobulin component alone. Heat inactivation of the complement in sera had no effect on the potency of immune sera.  相似文献   

7.
Specific antibody responses to Flavobacterium columnare (isolate ATCC 23463T) were characterized in plasma and mucus of tilapia following intraperitoneal (i.p.) injection or immersion immunization with formalin-killed sonicated or whole cell preparations. Fish (30 per treatment) received a primary immunization and were booster immunized 4 weeks later. An enzyme-linked immunosorbent assay was developed for detection and quantification of specific anti-F. columnare antibody, and it was found that formalin-killed sonicated cells in Freund's complete adjuvant (FCA) injected i.p. stimulated a significant systemic antibody response within 2 weeks (mean titre 11,200) which increased to 30,600 following secondary immunization. At 10 weeks post-immunization, the mean titre remained significantly elevated above the controls. Antibodies were also observed in cutaneous mucus of fish immunized i.p. with formalin-killed sonicated cells in FCA at 6 and 8 weeks post-immunization (mean titres 67 and 33, respectively). Although some individual fish responded, mean plasma and cutaneous mucus antibody titres were not significantly greater than controls in any of the other treatment groups. The results of this study demonstrate that tilapia can mount a significant humoral response in plasma and cutaneous mucus to F. columnare, but i.p. immunization with FCA is required to elicit this response.  相似文献   

8.
A previously developed monoclonal antibody (mAb) specific for the heavy chain of hybrid striped bass (HSB) Morone chrysops×M. saxatilis immunoglobulin was used in an assay to detect the humoral response to antigens of Streptococcus iniae. In order to validate this assay, an anti‐S. iniae antibody was produced in HSB by immunization with formalin‐killed cells of S. iniae mixed with Freund's complete adjuvant (FCA). After boosting with cells mixed with Freund's incomplete adjuvant (FIA), fish were challenged with live S. iniae by i.p. injection. This resulted in mortalities of 100%, 13% and 7% for non‐immunized, immunized and non‐challenged fish respectively. Live S. iniae were recovered only from moribund non‐immunized challenged fish, indicating that the immunization conferred protection against S. iniae. Sera were taken at 28 and 42 days post challenge and anti‐S. iniae antibody was measured by both agglutination and indirect ELISA. Titres of anti‐S. iniae antibody increased only in the immunized group as measured by agglutination and ELISA. Serum complement measured in the final bleeding was significantly higher in the immunized group. Western blotting using immune HSB serum indicated that the predominant antigen in this case was the high molecular weight polysaccharide of S. iniae.  相似文献   

9.
A multiplex nested-polymerase chain reaction (PCR)-based (m-nested PCR) method was developed for simultaneous detection of four important freshwater/marine fish pathogens in subtropical Asia, including Aeromonas hydrophila, Edwardsiella tarda, Photobacterium damselae and Streptococcus iniae . The specificity of the oligonucleotide primers used for PCR detection was confirmed to generate specific amplicons for the corresponding pathogens. Moreover, non-specific amplicons were observed when the primers were tested using pure DNA extracted from 31 related bacterial strains belonging to 23 species or tissue homogenates of infected tilapia. This m-nested PCR approach could detect 19 colony forming unit (CFU) for A. hydrophila , 62 CFU for E. tarda , 280 CFU for P. damselae subsp. piscicida and 179 CFU for S. iniae in infected tilapia kidney homogenates, consistent with the results derived from bacteriological methods. The assay described in this paper is a sensitive and effective method for simultaneous detection of multiple fish pathogens.  相似文献   

10.
Serum and mucosal antibody responses of juvenile rainbow trout, Oncorhynchus mykiss, were characterized by enzyme‐linked immunosorbent assay (ELISA) following immunization with various preparations of formalin‐killed Flavobacterium psychrophilum cells. The protective nature of these preparations was then determined by immunizing rainbow trout fry and challenging with the bacterium. Juvenile rainbow trout immunized intraperitoneally (i.p.) with formalin‐killed F. psychrophilum emulsified with Freund's complete adjuvant (FCA), and i.p. with formalin‐killed F. psychrophilum either with or without culture supernatant generated significant serum antibody responses by 6 and 9 weeks, respectively. Significant mucosal antibody responses were detected by 9 weeks only in fish immunized i.p. with killed F. psychrophilum/FCA. Following immunization and bacterial challenge of rainbow trout fry, protective immunity was conferred in F. psychrophilum/FCA and saline/FCA groups with relative per cent survival values of up to 83 and 51, respectively. Significant protection was not observed in treatment groups immunized by immersion or i.p. without adjuvant at the challenge doses tested. Results suggest that stimulation of non‐specific immune factors enhances the ability of fish to mount a protective immune response, but specific antibody appears necessary to provide near complete protection. In this study, an ELISA was developed to monitor anti‐F. psychrophilum antibody production in trout. The relationship of such responses to protective immunity suggests that future vaccination strategies against coldwater disease may require stimulation of both the innate and adaptive arms of the immune response.  相似文献   

11.
Francisella noatunensis subsp. orientalis is a pathogen of tilapia and other warm‐water fish for which no vaccines are commercially available. In this study, a whole cell formalin‐inactivated vaccine was developed for the first time using the highly virulent isolate STIR‐GUS‐F2f7 and the oil‐based adjuvant Montanide? ISA 763A VG. The efficacy of the vaccine was assessed in red Nile tilapia via intraperitoneal (i.p.) injection using homologous experimental infection and correlates of protection such as seral antibody production and bacterial loads in the spleen. For immunization, fish were i.p. injected with 0.1 ml of the vaccine, the adjuvant alone or PBS. At 840 degree days post‐vaccination, all fish were i.p. injected with 4.0 × 103 CFU/fish of pathogenic bacteria. The RPS at the end of the trial was 100% in the vaccinated group with significantly higher survival than in the adjuvant and control groups. The RPS in the adjuvant group was 42%, and no significant difference was seen in survival between this and the PBS group. Moreover, significantly higher antibody titres in the serum and significantly lower bacterial loads in the spleen were detected in the vaccinated fish by ELISA and qPCR, respectively. These findings highlight the potential of autogenous vaccines for controlling francisellosis in tilapia.  相似文献   

12.
Abstract. The level of protective immunity in rainbow trout, Salmo gairdneri Richardson, following vaccination by intraperitoneal (i.p.) injection or by direct immersion with Vibrio anguillarum was studied over a period of 46 weeks. Antibody titres to V. anguillarum O-antigen were determined with ELISA at the time of each experimental infection to study the relation between specific antibody production and protective immunity. Both vaccination by i.p. injection and vaccination by direct immersion induced a high degree of protective immunity against vibriosis which lasted for at least 46 weeks. At each experimental infection, the proportions of seropositive fish were lower than the proportions of fish with protective immunity. However, seropositive individuals were found in both experimental groups throughout the study.  相似文献   

13.
Ichthyophthirius multifiliis, commonly called “Ich,” is a protozoan parasite that infects the epidermis and gills of freshwater fish. Here, we used goldfish (Carassius auratus) to determine whether the four vaccine preparations (live trophonts, formalin‐fixed trophonts, freeze‐thawed trophont lysates, and trophont cilial and cell cortical fractions) of I. multifiliis (Fouquet) can elicit resistance of immunized fish against subsequent theront challenge, and whether a relationship exists between in vitro immobilization of theronts in sera or mucus from immunized fish and host protective immunity against the parasite. Experimental goldfish were randomly divided into five groups with five parallel controls, with each group or subgroup consisting of 20 individuals. Each test goldfish was immunized with one of the four Ich vaccine preparations administered by one of the three routes: live trophonts by gavage (Group 1), formalin‐fixed trophonts by injection (Group 2) or by immersion (Group 3), freeze‐thawed trophont lysates by injection (Group 4), and trophont cilial and cell cortical fractions by injection (Group 5). The fish were then challenged by a standard Ich theront challenge on Day 21 postimmunization. For every 10 d following immunization, we tested the in vitro immobilization of Ich theronts by sera or mucus from immunized and control fish. We found that although all control (nonimmunized) and Group 3‐immunized goldfish died following theront challenge, more than half (55–65%) of the immunized fish from Groups 1, 2, 4, and 5 survived. Furthermore, except for the fish from Groups 3 and 4, the number of mean days to death in each test group was significantly higher than that in the respective control. These results indicate that goldfish immunized by injection or by gavage with any of the four vaccine preparations gained resistance to Ich infection. Further analysis indicated that this protective immunity was closely associated with the in vitro immobilization of Ich theronts by fish sera or mucus.  相似文献   

14.
The humoral immune responses and host protection of channel catfish, Ictalurus punctatus (Rafinesque), against Ichthyophthirius multifiliis (Ich) were determined after immunization with live theronts and sonicated trophonts. Immunizations with live theronts or sonicated trophonts were carried out by both bath immersion and intraperitoneal (i.p.) injection. Cutaneous and serum immunoglobulin (Ig) levels and anti-Ich antibodies were measured 12 and 21 days post-immunization. The level of Ich infection and survival of catfish were determined after theront challenge. Cutaneous and serum anti-Ich antibodies were significantly higher (P < 0.05) in fish immunized with live theronts by immersion or i.p. injection, or with sonicated trophonts administered by i.p. injection, than in fish immunized with sonicated trophonts by immersion, with bovine serum albumin by i.p. injection, or non-immunized controls. Host protection was noted only in fish immunized with live theronts by immersion or i.p. injection or with sonicated trophonts by i.p. injection. There was a positive correlation between higher levels of anti-Ich antibodies and host survival in the immunized fish.  相似文献   

15.
Streptococcus agalactiae is a major bacterial pathogen that is the cause of serious economic losses in many species of freshwater, marine and estuarine fish worldwide. A highly efficacious S. agalactiae vaccine was developed using extracellular products (ECP) and formalin-killed whole cells of S. agalactiae. The vaccine efficacy following storage of S. agalactiae ECP and formalin-killed S. agalactiae cells at 4 degrees C for 1 year was determined. The stored ECP containing S. agalactiae formalin-killed cells failed to prevent morbidity and mortality among the vaccinated fish, and the relative percentage survival was 29. Serum antibody responses of the stored ECP and freshly prepared ECP against soluble whole cell extract of S. agalactiae indicated that significantly less antibody was produced in fish immunized with stored ECP and S. agalactiae cells than in those fish immunized with freshly prepared ECP and S. agalactiae cells at day 31 post-vaccination. Silver staining of sodium dodecyl sulphate-polyacrylamide gels and immunostaining of Western blots with tilapia antiserum to S. agalactiae revealed that predominant 54 and 55 kDa bands were present in the freshly prepared ECP fraction. The 55 kDa band was absent from the stored ECP and new bands below 54 kDa appeared on the Western blot. The results of this study on S. agalactiae ECP provide evidence for a correlation between protection and antibody production to ECP and for the importance of the 55 kDa ECP antigen for vaccine efficacy.  相似文献   

16.
Abstract— Streptococcus iniae was isolated from moribund pure strain Nile tilapia Oreochromis niloticus in a water recirculation aquaculture facility. The extent of the infection was determined in a number of separate culture systems at the facility. A pilot study was also conducted to gain information on potential stressors that may play a role in spread of the infection within such a water reuse facility. The conditions employed in the pilot stress test did not have any apparent impact on spread of S. iniae infection to fish in additional tanks within the facility.  相似文献   

17.
Abstract. The humoral immune response of turbot, Scophthalmus maximus (L.), to antigens from the microsporean parasite Tetramicra brevifilum Matthews & Matthews, 1980, was studied. Thirty days after intraperitoneal immunization with whole T. brevifilum spores in Freund's complete adjuvant, double indirect ELISA indicated that initial production of antibodies to parasite surface antigens was considerably higher than production of antibodies to the antigens contained in a crude extract (CE) of spores. Following re-immunization without adjuvant on day 30, levels of antibodies to surface antigens gradually declined, whilst levels of antibodies to CE antigens increased. The antibody response of intraperitoneally immunized fish was characterized by Western blotting of total soluble antigens obtained by heating and reduction of T. brevifilum spores at 95–100°C in Tris-HCl buffer containing SDS and dithiothreitol: a series of bands with molecular weights between 20 and 53 kDa was recognized by immunized turbot sera. Four additional bands (with molecular weights between 15 and 18kdA) were recognized by serum from re-immunized fish. ELISA studies of sera from naturally infected fish revealed a surprisingly low incidence of strong T. brevifilum seropositivity (61% individuals); antibodies to surface antigens predominated in seropositive individuals. The low background response levels and high sensitivity of the ELISA used in this study indicate that the assay is of value for the monitoring of serum antibody levels in turbot. However, given the relatively low seropositivities observed in naturally infected turbot, particularly to CE antigens, the use of anti- T. brevifilum serum antibody levels for the diagnosis of infection by this parasite may lead to false negative results.  相似文献   

18.
Japanese flounder, Paralichthys olivaceus (Temminck et Schlegel), were immunized by the intraperitoneal (i. p.) injection route with formalin‐killed whole cells of Vibrio anguillarum that originated from a diseased fish. Fifty days later, a booster vaccination was given by the same route. Control fish were similarly treated with sterile phosphate‐buffered saline. The efficacy of vaccination was evaluated based on protection against two bacterial challenges and immune responses (both specific and non‐specific). The challenges were performed by i. p. injection with V. anguillarum or V. parahaemolyticus. The results indicated that the vaccinated fish showed higher non‐specific immune activity than the unvaccinated fish. The effects of vaccinations on the phagocytic activity of phagocyte, bactericidal and lysozyme activities were notable, especially on bactericidal activity. Determined by ELISA, antiserum of vaccinated fish displayed high antibody titres. The vaccination conferred protection against V. anguillarum challenge (81.25–93.75% relative percentage survival (RPS)). The RPS was 46.15–53.85% against V. parahaemolyticus challenge, indicating some degree of cross‐protective immunity.  相似文献   

19.
Passive immunization of rainbow trout, Oncorhynchus mykiss (Walbaum), was carried out to determine the persistence of anti-Streptococcus sp. antibodies (ASA) raised in sheep, rabbits or rainbow trout. The protection afforded by passive immunization was compared with the protection obtained from active immunization by immersion in or intraperitoneal (i.p.) injection with formalin-killed cells. Assessments were undertaken concurrently for up to 3 months post-immunization (PI) to evaluate the practical potential of passive immunization. Passively administered sheep and rabbit antibodies were detected in fish sera by enzyme-linked immunosorbcnt assay for more than 60 days after i.p. injection. Fish responded immunologically to these antibodies and the highest humoral responses to sheep and rabbit ASA occurred at 2 months PI. The relative per cent survival (RPS) of rainbow trout challenged with virulent Streptococcus sp. after an i.p. injection (0.1 ml 100 g?1 fish body weight) of sheep, rabbit or fish ASA was: 88.8, 50 and 0% after 1 month; 33.3, 6.8 and 6.8% after 2 months; and 13.3, 0 and 6.6% after 3 months PI, respectively. Fish immunized actively had an RPS of 88.8 and 11.1% after 1 month, 38.1 and 4.7% after 2 months, and 36 and 0% after 3 months PI for the i.p. injection and immersion routes, respectively.  相似文献   

20.
罗非鱼海豚链球菌16S rRNA基因的序列测定和系统进化分析   总被引：18，自引：2，他引：16  
甘西 《水产学报》2007,31(5):618-623
为了从分子水平上对1株致病性罗非鱼链球菌进行分类学鉴定,利用原核生物16SrRNA基因通用引物对分离纯化的罗非鱼致病性链球菌进行16S rRNA基因的克隆及序列分析。结果扩增出长约1.5 kp目的片段,测序得到1条长度为1 447 bp核苷酸序列。核苷酸相似性分析表明,序列与NCB I公布的海豚链球菌(Streptococcus iniae,S.iniae)SCCF5L菌株16SrRNA基因核苷酸序列相似性最高(99.4%),暂称为中国广西株(S.iniae-CGX)。同时,亲源关系较近的S.iniae、S.difficilis和S.agalactiae代表菌株构建的系统发育进化树显示,所得菌株与S.iniae代表菌株组成同一进化分支,与S.agalactiae代表菌株组成的另一进化分支距离较近(95.5%),而与S.difficilis代表菌株组成的进化分支距离较远(92.3%)。上述研究证实,本试验从发病罗非鱼脑组织分离到的致病性链球菌为海豚链球菌。  相似文献   

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