中国南方地区罗非鱼无乳链球菌的分子流行病学研究

从广东省以及海南省等地区养殖的患病罗非鱼体内分离、收集到多株致病菌,经生化分析和分子生物学鉴定,均为无乳链球菌。对这些菌株分别进行了耐药谱测定、分子分型试验以及分子血清型分析。药敏试验结果表明,2007—2010年分离到的无乳链球菌耐药谱基本相似;多位点可变数目串联重复序列分析(MLVA)试验中,选择5个高变异指数的可变数目重复位点(VNTR)进行分子分型,结果表明,所有鱼源无乳链球菌菌株为同一MLVA型,而作为对照的牛源无乳链球菌则明显不同;为了对这些菌株进一步分型,分别进行了分子血清型和表面蛋白抗原基因的检测,结果表明,鱼源无乳链球菌的分子血清型均为Ⅰa型,表面蛋白抗原均为alpha-C蛋白。这进一步说明了不同年份和不同地区的鱼源无乳链球菌在基因水平上为同一分子类型,具有相同的起源或传染源。同时也说明,我国南方地区罗非鱼无乳链球菌在这几年中未发生明显的遗传变异。这些结果为罗非鱼无乳链球菌病疫苗研制,疫病监测及药物防治的研究提供理论依据。     

Molecular typing by pulsed-field gel electrophoresis of Flavobacterium psychrophilum isolates derived from Japanese fish

Sixty-four isolates of Flavobacterium psychrophilum from ayu, Plecoglossus altivelis altivelis (Temminck & Schlegel), and other fish (n=16) in Japan and the type strain (NCIMB 1947(T)) were typed using pulsed-field gel electrophoresis (PFGE) with endonuclease BlnI and XhoI. These isolates were classified into 20 clusters and 42 genotypes by PFGE analysis. The most predominant cluster of isolates from ayu was cluster XII (n=20), followed by clusters XVII, XVI, XX, XI, IX, X, XIII and XV; the remaining 17 isolates from other fish were divided into clusters I, II, III, IV, V, VI, VII, VIII, XIV, XVIII and XIX. The PFGE genotype of isolates from ayu clearly differed from those of other fish. The isolates from ayu in Gunma Prefecture belonged to clusters XII, XVI, XVII and XX, and the strains of three of these clusters (XII, XVII and XX) were isolated from ayu in 15 of 19 prefectures. PFGE typing enabled more accurate classification of isolates into clusters than previously achieved by analysing the restriction fragment length polymorphism of PCR products. These results suggest that F. psychrophilum isolated from ayu and other fish are genetically different and strains with several PFGE types have spread within Japan.     

Clonal structure in Ichthyobacterium seriolicida,the causative agent of bacterial haemolytic jaundice in yellowtail,Seriola quinqueradiata,inferred from molecular epidemiological analysis

Bacterial haemolytic jaundice caused by Ichthyobacterium seriolicida has been responsible for mortality in farmed yellowtail, Seriola quinqueradiata, in western Japan since the 1980s. In this study, polymorphic analysis of I. seriolicida was performed using three molecular methods: amplified fragment length polymorphism (AFLP) analysis, multilocus sequence typing (MLST) and multiple‐locus variable‐number tandem repeat analysis (MLVA). Twenty‐eight isolates were analysed using AFLP, while 31 isolates were examined by MLST and MLVA. No polymorphisms were identified by AFLP analysis using EcoRI and MseI, or by MLST of internal fragments of eight housekeeping genes. However, MLVA revealed variation in repeat numbers of three elements, allowing separation of the isolates into 16 sequence types. The unweighted pair group method using arithmetic averages cluster analysis of the MLVA data identified four major clusters, and all isolates belonged to clonal complexes. It is likely that I. seriolicida populations share a common ancestor, which may be a recently introduced strain.     

Multilocus sequence analysis revealed a high genotypic diversity of Aeromonas hydrophila infecting fish in Uganda

A multilocus sequence analysis (MLSA) was carried out to delineate Aeromonas hydrophila from fish in Uganda. Five housekeeping genes including recA, gyrB, metG, gltA and pps; and the 16S rRNA gene were amplified and sequenced from a total of nine A. hydrophila isolates. The obtained sequences were edited, and consensus sequences generated for each gene locus. The housekeeping gene sequences were concatenated and phylogenetic analysis performed in MEGA version 7.0.2. Pairwise distances ranged from 0.000 to 0.118, highest within the gltA gene locus and lowest within the 16S rRNA gene. The average evolutionary diversity within isolates from the same source ranged between 0.002 and 0.037, and it was 0.033 between the different sources. Similar tree topologies were obtained from the different gene loci with recA, metG and gyrB being more consistent in discriminating isolates according to sources while the 16S rRNA gene had the lowest resolution. The concatenated tree had the highest discriminatory power. This study revealed that A. hydrophila strains infecting fish in Uganda are of diverse genotypes suggesting different sources of infection in a given outbreak. Efforts to minimize spread of the bacteria across sources should be emphasized to control infections of mixed genotypes.     

无乳链球菌LIP蛋白的截短表达及免疫保护性研究

为评价罗非鱼源无乳链球菌(Streptococcus agalactiae)BX2012株脂蛋白(lipoprotein)的免疫原性及其对奥利亚罗非鱼(Oreochromis aureus)和大菱鲆(Scophthalmus maximus)的保护效果,以无乳链球菌脂蛋白基因序列(GenBank序列号:CP000114.1,SAK_0321)的B细胞线性抗原表位区设计特异性引物进行扩增,构建重组表达载体,将截短表达的脂蛋白制备成亚单位疫苗,同时制备灭活疫苗进行免疫比对。结果显示:重组表达载体p ET-32a-LIP342在BL21(DE3)中获得了良好的可溶性表达,分子质量约为30 kDa,纯化使LIP342纯度由41.75%提至87.23%;Western blotting分析显示,LIP342可被兔抗无乳链球菌高免血清特异性识别;LIP342在目前已知不同种属来源或不同血清型的无乳链球菌分离株中同源性为90.35%~100%,在罗非鱼源分离株中同源性为100%;无乳链球菌LIP342蛋白和灭活疫苗均可显著提升奥利亚罗非鱼和大菱鲆血清抗体水平,而LIP342诱导的血清抗体水平均显著高于灭活疫苗;经0.1 mL 1×10~9cfu/mL的无乳链球菌攻毒后,LIP342蛋白和灭活疫苗对供试鱼的累积存活率均显著高于PBS对照组。结果表明,高度保守的LIP342具有较好的免疫原性,可作为无乳链球菌亚单位疫苗候选因子。     

Identification of growth‐related nucleotide polymorphism in cultured Pacific bluefin tuna,Thunnus orientalis

Pacific bluefin tuna (PBF), Thunnus orientalis, is commercially one of the most important species of tuna. In this study, amplified fragment length polymorphism (AFLP) screening was conducted to find the growth‐related polymorphic DNA in cultured PBF. Fish hatched in 2007 were harvested at an age of 818–1994 days. They were categorized into superior, average and inferior growth groups, depending on their growth score at the time of harvest. On AFLP screening of 24 fish, with eight fish from each group, 215 polymorphic DNA fragments were observed. A second amplification, with EcoRI + ACC and MseI + CCC primers, generated a polymorphic fragment of 630 bp at a rate of 80.0% (n = 15) in the superior, 56.3% (n = 16) in the average and 20.0% (n = 15) in the inferior growth groups. Polymerase chain reaction (PCR) primers, which could amplify both AFLP‐positive and AFLP‐negative loci, were developed using the consensus sequence outside the AFLP target fragment. Eleven haplotypes were obtained by sequence analysis of the PCR product at the AFLP target loci. Among those, haplotype 1 was statistically significant in the superior and average growth groups and could be used as a molecular marker for distinguishing the individuals with superior and average growth from those with inferior growth.     

Antigenicity of Streptococcus agalactiae extracellular products and vaccine efficacy

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.     

PCR-RFLP genotypes associated with quinolone resistance in isolates of Flavobacterium psychrophilum

A novel genotyping method for epizootiological studies of bacterial cold-water disease caused by Flavobacterium psychrophilum and associated with quinolone resistance was developed. Polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP) was performed on 244 F. psychrophilum isolates from various fish species. PCR was performed with primer pair GYRA-FP1F and GYRA-FP1R amplifying the A subunit of the DNA gyrase (GyrA) gene, which contained the quinolone resistance determining region. Digestion of PCR products with the restriction enzyme Mph1103I showed two genotypes, QR and QS. The difference between these genotypes was amino acid substitutions at position 83 of GyrA (Escherichia coli numbering). The genotype QR indicated an alanine residue at this position associated with quinolone resistance in F. psychrophilum isolates. Of the 244 isolates tested in this study, the number of QR genotype isolates was 153 (62.7%). In isolates from ayu (n=177), 146 (82.5%) were genotype QR. With combination of this technique and previously reported PCR-RFLP genotyping, eight genotypes were observed in F. psychrophilum isolates. Using this genotyping system, the relationships between genotype and host fish species, or locality of isolation, were analysed and are discussed.     

中国罗非鱼主养区无乳链球菌的分子流行特征及其传播方式

为分析2007—2015年中国罗非鱼主养区无乳链球菌(Streptococcus agalactiae)的分子特征和流行情况,分离并收集了248株罗非鱼源无乳链球菌。通过分子血清型、MLST、毒力基因和前噬菌体等分型方法对248株无乳链球菌进行了分子遗传特征分析。结果表明,229株无乳链球菌(92.3%)的分子血清型是Ⅰa型,其余19株均是Ⅰb型(7.7%)。MLST分析结果表明,所有Ⅰa型无乳链球菌都是ST7型,所有Ⅰb型无乳链球菌都是ST261型。毒力基因检测结果发现,229株Ⅰa-ST7型无乳链球菌的毒力基因型相同,即V1型;19株Ⅰb-ST261型无乳链球菌的毒力基因型相同,即V2型。前噬菌体检测结果表明,Ⅰa-ST7型无乳链球菌可分为两种前噬菌体基因型,分别是P1型(36株)和P2型(193株);Ⅰb-ST261型无乳链球菌的10个前噬菌体基因都是阴性,即P3型。根据以上4种分子分型方法可将248株无乳链球菌分为3种基因型,即Ⅰa-ST7-V1-P1型、Ⅰa-ST7-V1-P2型和Ⅰb-ST261-V2-P3型。2010—2011年主要流行菌株由Ⅰa-ST7-V1-P1型转变为Ⅰa-ST7-V1-P2型,其中Ⅰa-ST7-V1-P1型是2011年之前的主要流行菌株,Ⅰa-ST7-V1-P2型在2011年及之后成为主要流行菌株。研究表明,近年来我国罗非鱼源无乳链球菌发生了明显的遗传变异,同时,根据我国罗非鱼主养区无乳链球菌的流行特点,推测我国罗非鱼源无乳链球菌是通过苗种或水体等介质进行传播的,属于输入性传播方式。     

Homogeneity of <Emphasis Type="Italic">Streptococcus dysgalactiae</Emphasis> from farmed amberjack <Emphasis Type="Italic">Seriola dumerili</Emphasis> in Japan

Streptococcus dysgalactiae strains have been isolated from cultured amberjack Seriola dumerili and yellowtail Seriola quinqueradiata in Japan. To characterize the fish isolates, we performed genetic analysis and compared the biochemical properties of these isolates with those of the S. dysgalactiae subsp. dysgalactiae and S. dysgalactiae subsp. equisimilis strains isolated from mammals. The genetic analysis revealed that the fish isolates were genetically very similar to each other with high DNA–DNA relatedness (>95.4%) and sequence homology. Meanwhile, the DNA relatedness between mammalian isolates and the fish isolates was 73.4–82.6%. In biased sinusoidal gel electrophoresis (BSFGE) analysis, the restriction patterns of mammalian isolates were different from those of fish isolates. The fish isolates did not show streptokinase activity in plasminogen obtained from mammals. These characteristics enabled us to distinguish between the fish isolates and the Sdd and Sde strains isolated from mammals. In order to obtain epidemiological information on the fish isolates, BSFGE patterns from 284 S. dysgalactiae strains from fish in Japan were examined. Based on the results of BSFGE analysis, the fish isolates were classified into 16 groups (AP1–AP16) with restriction enzyme ApaI. The dendrogram based on BSFGE analysis indicated that all fish isolates using in this study were closely related.     

广东地区吉富罗非鱼无乳链球菌病的流行情况与耐药性

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。     

MHC基因在鱼类遗传育种中的研究与应用

主要组织相容性复合体（MHC,Major histoeompatibility complex）是最具多态性的免疫分子。其多态性主要取决于群体内MHC基因的大量等位基因及等位基因间高度的序列变异。等位基因的序列变异主要发生在抗原肽结合区（peptide binding region,PBR）,这决定了不同基因型个体的免疫力差异。鱼类受多倍化影响,MHC基因的多态性还表现在同一基因存在多个基因座位（多拷贝）。MHC的高度多态性及与机体免疫力的相关性,使其在鱼类遗传进化、抗病育种等方面倍受关注。本文介绍了有关鱼类MHC基因的结构、功能及其在遗传育种中的应用。     

Streptococcus agalactiae Vaccination and Infection Stress in Nile Tilapia,Oreochromis niloticus

Abstract

The stress response following intraperitoneal (IP) injection of a non-adjuvant Streptococcus agalactiae vaccine in cultured warmwater Nile tilapia, Oreochromis niloticus, has not been investigated. Further, little or no information is available on stress following S. agalactiae infection and what effect, if any, vaccination has on susceptibility to infection. The objective of this study was to develop preliminary information on the associations between vaccination, stress, and infection. Blood glucose levels were used to evaluate stress in the fish at different time intervals following vaccination and challenge with S. agalactiae. Blood glucose levels were measured in vaccinates and controls at 0, 2, 6, 24 hours, and 28 days post-immunization (0 hours pre-challenge), and at 2,6, 24,48,72, and 312 hours following challenge with 1.5 × 10⁴ colony forming units (CFU) of S. agalactiae/fish. Significant increases in blood glucose levels were observed only in association with the injection of the vaccine and at 2 hours after injection. After S. agalactiae challenge, both controls and vaccinates had significantly (P < 0.05) higher blood glucose values at 2, 24,48, and 72 hours than at 0 hours. However, blood glucose levels in vaccinates were significantly (P < 0.05) lower than the controls at 24, 48, and 312 hours. Blood glucose levels and mortality of the infected controls were significantly correlated (r² = 0.9236, P = 0.0134). The cumulative mortality of the vaccinates and controls was 10% and 60% after 13 days post-challenge, respectively. The relative percent survival (RPS) was 83.4. Our results indicate that the vaccine was efficacious against S. agalactiae and induced short-term stress in tilapia. These preliminary results also suggested, for the first time, that vaccination may significantly reduce the infection stress associated with S. agalactiae infection in tilapia.     

红尾皇冠鱼(Aequidens rivulatus)病原无乳链球菌的分离、鉴定与特性分析

为查明天津地区养殖红尾皇冠鱼(Aequidens rivulatus)大量死亡的原因,取患病红尾皇冠鱼进行细菌分离,从病鱼肾脏中分离到优势菌株071901,人工感染试验证实其对红尾皇冠鱼有较强的致病性;采用形态学观察、生理生化特性、16S r RNA基因序列系统发育树分析及cfb(GBS-specific gene cfb,CAMP factor)基因检测对菌株071901进行鉴定。结果显示,菌株071901为革兰氏阳性球菌,生化特性与链球菌属的无乳链球菌(Streptococcus agalactiae)相近,基于16S r RNA基因序列的系统发育分析将菌株071901与无乳链球菌聚为一支,以071901为模板特异性扩增无乳链球菌的cfb基因,也得到了预期的900 bp的核酸片段,进一步证实菌株071901为无乳链球菌。对30种抗生素的药敏结果显示,菌株071901对红霉素、阿奇霉素等19种药物敏感,对多粘霉素、罗红霉素、呋喃唑酮等11种药物耐药。     

Common pearl oysters in China, Japan, and Australia are conspecific: evidence from ITS sequences and AFLP

ABSTRACT:   To elucidate the species status of Pinctada fucata in China, P. fucata martensii in Japan and P. imbricata in Australia, one population of each taxon was studied using internal transcribed spacer 1 and 2 (ITS1, and ITS2) and amplified fragment length polymorphism (AFLP) markers. ITS1 and ITS2 were 401–405 and 229–237 bp long, respectively. Twenty-nine ITS1 and 15 ITS2 unique genotypes were obtained from 44 and 34 individuals, respectively, with some genotypes shared by two or three populations. In AFLP analysis, each individual exhibited a distinct phenotype. No population had diagnostic markers. Mean genetic divergences within and among the three populations were very low and overlapped (between-population: 0.7–0.9% for ITS1, 0.9–1.3% for ITS2, and 53.3–55.6% for AFLP; within-population: 0.5–0.9% for ITS1, 0.8–1.2% for ITS2, and 50.4–53.6% for AFLP). Low levels of genetic differentiation were observed among the three populations while the Australian population is partially genetically isolated. Under an infinite allele model, genetic differentiation among populations was not significant based on a permutation test. Under an infinite site model, most F _ST values were not significant for ITS data although they were significant for AFLP data. Network analysis using ITS data indicated that individuals from the same population did not cluster together. Analysis of molecular variance ( amova ) demonstrated that > 94% variation was contributed by within-population variation. These findings suggest that the three taxa are conspecific and Pinctada fucata is the correct name.     

Streptococcosis on a red tilapia, Oreochromis sp., farm: a case study

A commercial red tilapia farm was diagnosed with Streptococcus agalactiae infection using histopathology, microbiology and immunohistochemistry. One hundred fish were randomly taken from different weight/age groups including broodstock, market fish (larger than 150 g), on-growing fish between 20 and 150 g, juveniles and larvae. Fish were clinically examined, anaesthetised and necropsied. Samples were taken from brain, liver, spleen, eyes and kidney for microbiology. All organs were processed for histopathology and an indirect immunoperoxidase test (IIP). Organs from wild fish and birds found in close proximity to the farm were also sampled for microbiology and IIP. The prevalence of lesions or infection found by IIP, histopathology and microbiology was 16%, 29% and 7% respectively. Clinical disease, lesions or infection were not seen in larvae or juveniles. By contrast, infection and disease were found in fish larger than 20 g, suggesting that the condition was linked to the intensive culture conditions of broodstock, on-growing and market fish. S. agalactiae was not found in wild fish, or in birds, by microbiology and IIP.     

Epidemiological study on <Emphasis Type="Italic">Lactococcus garvieae</Emphasis> isolates from fish in Japan

In Japan, Lactococcus garvieae infection has been the main fish disease in aquaculture. Although commercial oral and injectable vaccines have been used to prevent L. garvieae infection in Japan, L. garvieae has been isolated not only from unvaccinated fish but also from vaccinated fish in which immunity induced by vaccination had diminished. In order to obtain epidemiological information on this fish pathogen, we conducted biased sinusoidal field gel electrophoresis (BSFGE) pattern analysis and phage typing of L. garvieae isolates (n = 427) from fish in Japan. These isolates were obtained from 13 different fish species between 1980 and 2007. In the BSFGE analysis, L. garvieae isolates were classified into 17 groups (S1–S17) based on the SmaI digestion patterns and into four groups (A1–A4) based on the ApaI digestion patterns. Phage typing revealed five different phage susceptibility profiles (A–E) in L. garvieae isolates. Since 2005, comparisons of the results of phage typing and BSFGE have indicated the presence of a novel genotype (S16/A4) with phage type E. All the strains belonging to this type showed lincomycin sensitivity.     

中国七种水生动物源无乳链球菌的分子特征及其对斑马鱼的致病性

为了解中国水生动物源无乳链球菌的分子流行特征,揭示其传播和流行规律,本实验对分离得到并鉴定的10株7种水生动物源无乳链球菌通过分子血清型、多位点序列分型(MLST)分型、毒力基因型和前噬菌体分型等方法进行分子分型;其次,通过斑马鱼评价7种水生动物源无乳链球菌的致病性。分子血清型分析结果表明,10株无乳链球菌可分为3种血清型,即Ⅰa、Ⅰb和Ⅲ型;MLST分型结果表明,Ⅰa型无乳链球菌均为ST7型,Ⅰb无乳链球菌均是ST261型,只有Ⅲ型无乳链球菌是ST739型。进一步分型结果表明,10株无乳链球菌可分为3种毒力基因型和4种前噬菌体基因型。根据4种分型结果可知,10株水生动物源无乳链球菌可分为4种类型,其中虎纹蛙源无乳链球菌具有独立的分子血清型、MLST型、毒力基因型和前噬菌体基因型,即Ⅲ-ST739-V1-P3;罗非鱼源无乳链球菌的基因型有3种,即Ⅰa-ST7-V2-P1、Ⅰa-ST7-V2-P2和Ⅰa-ST7-V3-P4;红尾皇冠鱼、鳙和罗非鱼源无乳链球菌的基因型相同:Ⅰa-ST7-V2-P2;卵形鲳鲹、宝石鲈和罗非鱼源无乳链球菌具有相同的基因型:Ⅰa-ST7-V2-P1;鲮和罗非鱼源无乳链球菌的基因型相同,即Ⅰb-ST261-V3-P4。致病性研究表明,7种水生动物源无乳链球菌对斑马鱼均有强致病性。研究表明,两栖类虎纹蛙源无乳链球菌和鱼源无乳链球菌的基因型明显不同,它们之间遗传变异较大,因此,无乳链球菌在两栖类和鱼类之间相互传播的可能性较小。鱼源无乳链球菌有3种基因型,且这3种基因型均在罗非鱼中流行,这表明无乳链球菌在鱼类中相互传播的可能性较大,尤其是在罗非鱼与其他鱼类之间。     

Differential pathogenicity of five Streptococcus agalactiae isolates of diverse geographic origin in Nile tilapia (Oreochromis niloticus L.)

Streptococcus agalactiae is an emerging pathogen of fish and has caused significant morbidity and mortality worldwide. The main objective of this study is to assess whether pathogenic differences exist among isolates from different geographic locations. Nile tilapia (Oreochromis niloticus L.) were administered an intraperitoneal injection of suspension containing USA, Brazil, Honduras, Israel, or Kuwait S. agalactiae isolates at concentrations ranging from 10² to 10⁷ cfu mL^?1. The LD₅₀ values 7 days after challenge were as follows: USA (1.0 × 10² cfu mL^?1), Brazil (1.5 × 10³ cfu mL^?1), Honduras (6.8 × 10³ cfu mL^?1), Israel (1.0 × 10⁴ cfu mL^?1) and Kuwait (7.2 × 10⁵ cfu mL^?1). Fish from all groups exhibited lethargy, anorexia, exophthalmia, corneal opacity, erratic swimming, petechiae and mortality. Opercular clearing and ascites were only found after infection with certain geographic isolates. The findings in this study indicate that S. agalactiae isolates of different geographic origin can cause significant mortalities after experimental challenge and can have different pathogenic capacities. Isolates from the Americas (USA, Brazil and Honduras) were more pathogenic to Nile tilapia than isolates from the Middle East/Asia (Israel and Kuwait).