为了调查广东省惠州、肇庆、珠海、湛江4个吉富罗非鱼主养区链球菌病的流行情况和耐药性,并进一步分析β-内酰胺酶基因与青霉素类药物的耐药性关系。本实验通过传统的方法对菌株进行分离纯化,扩增特异性基因cfb以及16s r DNA对各菌株进...为了调查广东省惠州、肇庆、珠海、湛江4个吉富罗非鱼主养区链球菌病的流行情况和耐药性,并进一步分析β-内酰胺酶基因与青霉素类药物的耐药性关系。本实验通过传统的方法对菌株进行分离纯化,扩增特异性基因cfb以及16s r DNA对各菌株进行鉴定;采用k-b法测定分离菌株的药物敏感性;通过PCR检测β-内酰胺酶类基因在分离菌株中的分布情况,并用Statistic6.0统计分析各β-内酰胺酶基因与青霉素类药物的耐药关系。实验结果表明,吉富罗非鱼无乳链球菌的阳性率从高到低的顺序为惠州(46.46%)>湛江(43.24%)>肇庆(17.30%)>珠海(4.17%);药敏结果显示各地区无乳链球菌分离株对青霉素(耐药率为94.29%)和磺胺二甲基嘧啶(耐药率为86.40%)普遍耐药,对恩诺沙星最为敏感(耐药率为3.99%);β-内酰胺酶基因分布与细菌耐药性统计结果显示,无乳链球菌基因组中的9个β-内酰胺酶基因在各分离菌株中的分布呈高度多态性,其中SAG0658基因与氨苄青霉素抗性显著相关,提示SAG0658基因在无乳链球菌耐氨苄青霉素过程中发挥主要作用;此外,9个β-内酰胺酶基因与青霉素抗性无相关性,说明其在菌株对青霉素耐药过程中并未发挥明显作用,提示分离菌株对青霉素的耐药性可能依赖其他途径。展开更多
Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present s...Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.展开更多
In recent years,piscine nodaviruses have emerged as major pathogens of a wide range of larval and juvenile marine finfish resulting in high mortality in aquaculture worldwide.Affected fish exhibit a range of neurologi...In recent years,piscine nodaviruses have emerged as major pathogens of a wide range of larval and juvenile marine finfish resulting in high mortality in aquaculture worldwide.Affected fish exhibit a range of neurological signs,such as erratic swimming behaviour with the associated microscopic lesions of necrosis and vacuolation of the central nervous tissues and retina.Numerous round-shaped,unenveloped and 25-30 nm in diameter virus particles were found in the cytoplasm of affected retinal and nerve cells.Nodaviruses have a bipartite genome of positive-sense RNA,with RNA1 encoding the RNA-dependent RNA polymerase and RNA2 encoding the capsid protein.Both RNA are capped,but not polyadenylated.The family Nodaviridae comprises two genera: Alphanodavirus and Betanodavirus,members of which primarily infect insects and fish,respectively.Therefore,betanodavirus is also named piscine nodavirus.At present,piscine nodaviruses are divided into four genotypes based on partial sequences of the coat protein gene.ELISA and RT-PCR amplification have been developed as specific diagnostic methods for the detection of the virus.Antibodies to striped jack(Pseudocaranx dentex) nervous necrosis(SJNNV) were found in 65% of plasma samples collected from wild and domestic brood stocks of striped jack,suggesting that the virus is very prevalent.Viral antigens were detected in eggs,larvae,and ovaries of hatchery-reared and wild spawner fish,suggesting both horizontal and vertical modes of transmission of the virus.Selection of nodavirus-free spawners using ELISA for detection of antigens and RT-PCR techniques have successfully reduced incidences of the virus infections in juvenile sea bass(Dicentrarchus labrax),striped jack and barfin flounder(Verasper moseri).The SSN-1 and GF cell lines have been successfully used in isolating piscine nodaviruses.Although there are many papers describing the molecular characteristics of betanodavirus,our knowledge of the genomic attributes of these viruses is still limited.Vaccination studies are being undertaken by a number of researchers and need to be fostered.In particular,the use of passive immunization of broodfish with homologous and heterologous,high-titre antisera are worthy of investigation.展开更多
文摘对斜带石斑鱼（Epinephelus coioides）的胚胎及仔稚幼鱼形态发育进行了观察与研究，详细描述从受精卵到初孵仔鱼的28个具体发育时期的形态特征和发育时间。根据研究观察结果，将斜带石斑鱼胚胎发育划分为卵裂期、囊胚期、原肠胚期、神经胚期和器官形成期。在水温（25±0.5）℃、盐度31.0、pH7.8的海水中，斜带石斑鱼胚胎历时28 h 30 min完成整个胚胎发育孵化出膜。胚后发育主要根据卵黄囊、腹鳍棘及第二背鳍棘、鳞片、体色的变化分为仔鱼期、稚鱼期、幼鱼期。仔鱼期根据卵黄囊的有无又分为早期仔鱼和晚期仔鱼。在水温24.5～29.2℃，盐度28.8～33.5，pH7.5～8.5的海水中，培育至36d，发育最快的斜带石斑鱼结束仔鱼期进入稚鱼期；培育至42d，发育最快的稚鱼完成变态，成为幼鱼。斜带石斑鱼胚后发育过程中最明显的变化是腹鳍棘与第二背鳍棘以及鳍棘上小刺的长出与收回。
文摘为了调查广东省惠州、肇庆、珠海、湛江4个吉富罗非鱼主养区链球菌病的流行情况和耐药性,并进一步分析β-内酰胺酶基因与青霉素类药物的耐药性关系。本实验通过传统的方法对菌株进行分离纯化,扩增特异性基因cfb以及16s r DNA对各菌株进行鉴定;采用k-b法测定分离菌株的药物敏感性;通过PCR检测β-内酰胺酶类基因在分离菌株中的分布情况,并用Statistic6.0统计分析各β-内酰胺酶基因与青霉素类药物的耐药关系。实验结果表明,吉富罗非鱼无乳链球菌的阳性率从高到低的顺序为惠州(46.46%)>湛江(43.24%)>肇庆(17.30%)>珠海(4.17%);药敏结果显示各地区无乳链球菌分离株对青霉素(耐药率为94.29%)和磺胺二甲基嘧啶(耐药率为86.40%)普遍耐药,对恩诺沙星最为敏感(耐药率为3.99%);β-内酰胺酶基因分布与细菌耐药性统计结果显示,无乳链球菌基因组中的9个β-内酰胺酶基因在各分离菌株中的分布呈高度多态性,其中SAG0658基因与氨苄青霉素抗性显著相关,提示SAG0658基因在无乳链球菌耐氨苄青霉素过程中发挥主要作用;此外,9个β-内酰胺酶基因与青霉素抗性无相关性,说明其在菌株对青霉素耐药过程中并未发挥明显作用,提示分离菌株对青霉素的耐药性可能依赖其他途径。
文摘Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.
文摘In recent years,piscine nodaviruses have emerged as major pathogens of a wide range of larval and juvenile marine finfish resulting in high mortality in aquaculture worldwide.Affected fish exhibit a range of neurological signs,such as erratic swimming behaviour with the associated microscopic lesions of necrosis and vacuolation of the central nervous tissues and retina.Numerous round-shaped,unenveloped and 25-30 nm in diameter virus particles were found in the cytoplasm of affected retinal and nerve cells.Nodaviruses have a bipartite genome of positive-sense RNA,with RNA1 encoding the RNA-dependent RNA polymerase and RNA2 encoding the capsid protein.Both RNA are capped,but not polyadenylated.The family Nodaviridae comprises two genera: Alphanodavirus and Betanodavirus,members of which primarily infect insects and fish,respectively.Therefore,betanodavirus is also named piscine nodavirus.At present,piscine nodaviruses are divided into four genotypes based on partial sequences of the coat protein gene.ELISA and RT-PCR amplification have been developed as specific diagnostic methods for the detection of the virus.Antibodies to striped jack(Pseudocaranx dentex) nervous necrosis(SJNNV) were found in 65% of plasma samples collected from wild and domestic brood stocks of striped jack,suggesting that the virus is very prevalent.Viral antigens were detected in eggs,larvae,and ovaries of hatchery-reared and wild spawner fish,suggesting both horizontal and vertical modes of transmission of the virus.Selection of nodavirus-free spawners using ELISA for detection of antigens and RT-PCR techniques have successfully reduced incidences of the virus infections in juvenile sea bass(Dicentrarchus labrax),striped jack and barfin flounder(Verasper moseri).The SSN-1 and GF cell lines have been successfully used in isolating piscine nodaviruses.Although there are many papers describing the molecular characteristics of betanodavirus,our knowledge of the genomic attributes of these viruses is still limited.Vaccination studies are being undertaken by a number of researchers and need to be fostered.In particular,the use of passive immunization of broodfish with homologous and heterologous,high-titre antisera are worthy of investigation.
