Isolation of viral haemorrhagic septicaemia virus from mummichog, stickleback, striped bass and brown trout in eastern Canada

Viral haemorrhagic septicaemia virus (VHSV) was isolated from mortalities occurring in populations of mummichog, Fundulus heteroclitus, stickleback, Gasterosteus aculeatus aculeatus, brown trout, Salmo trutta, and striped bass, Morone saxatilis, in New Brunswick and Nova Scotia, Canada. The isolated viral strains produced a cytopathic effect on the epithelioma papillosum cyprini cell line. Serum neutralization indicated the virus was VHSV and sequencing identified the rhabdovirus isolates as the North American strain of VHSV. Phylogenetic analysis indicated that the isolates are closely related and form a distinguishable subgroup of North American type VHSV. To our knowledge, this is the first report of VHSV in mummichog and striped bass.     

Temporal genetic structure of an intergrade largemouth bass (Micropterus salmoides) population

Since 1972, numerous impoundments throughout Texas have been stocked with Florida largemouth bass Micropterus salmoides floridanus. In order to examine some of the long-term effects of such efforts, the present study summarizes past studies and complements those with more current observations on the intergrade largemouth bass population in Aquilla Lake, Hill Country, Texas. The reservoir received 4 annual stockings during the month of May from 1982 to 1985. Collections occurred periodically from February 1984 to June 1987, February to August 1988, and monthly from August 1991 to November 1992. Horizontal starch-gel electrophoresis was conducted to determine allele frequencies at loci sAAT-B*, SOD*1. Mean heterozygosity and percent introgression were highest for age-0 fish of the 1986 year class. Assortative mating was not apparent in 1991 or 1992 year classes. Collections of specific year classes over time consistently indicated decreases in the relative abundance of the northern phenotype. Relative to the northern phenotype, the abundance of Florida year classes remained relatively constant over time. Relative abundances of F₁ hybrids and F_x, intergrades (i. e., F₁) from specific year glasses were observed to generally increase with age.     

大口黑鲈苗种致病性类志贺邻单胞菌的分离和鉴定

对江苏南京某养殖场温室养殖的患病大口黑鲈鱼种进行病原分离,得到1株优势菌株,命名为Q-1.对其进行生化检测以及回归感染试验,结合16S rRNA基因序列测定,确定菌株Q-1为类志贺邻单胞菌(Plesiomonas shigelloides).药敏试验结果表明,菌株Q-1对恩诺沙星、硫酸新霉素、氟苯尼考、盐酸多西环素和氟...     

FishPathogens.eu/vhsv: a user-friendly viral haemorrhagic septicaemia virus isolate and sequence database

A database has been created, http://www.FishPathogens.eu , with the aim of providing a single repository for collating important information on significant pathogens of aquaculture, relevant to their control and management. This database will be developed, maintained and managed as part of the European Community Reference Laboratory for Fish Diseases function. This concept has been initially developed for viral haemorrhagic septicaemia virus and will be extended in future to include information on other significant aquaculture pathogens. Information included for each isolate comprises sequence, geographical origin, host origin and useful key literature. Various search mechanisms make it easy to find specific groups of isolates. Search results can be presented in several different ways including table-based, map-based and graph-based outputs. When retrieving sequences, the user is given freedom to obtain data from any selected part of the genome of interest. The output of the sequence search can be readily retrieved as a FASTA file ready to be imported into a sequence alignment tool of choice, facilitating further molecular epidemiological study.     

Trade practices are main factors involved in the transmission of viral haemorrhagic septicaemia

Viral haemorrhagic septicaemia (VHS), caused by the novirhabdovirus viral haemorrhagic septicaemia virus (VHSV), causes significant economic problems to European rainbow trout, Oncorhynchus mykiss (Walbaum), production. The virus isolates can be divided into four distinct genotypes with additional subgroups. The main source of outbreaks in European rainbow trout farming is sublineage Ia isolates. Recently, this group of isolates has been further subdivided in to two subclades of which the Ia‐2 consists of isolates occurring mainly in Continental Europe outside of Denmark. In this study, we sequenced the full‐length G‐gene sequences of 24 VHSV isolates that caused VHS outbreaks in Polish trout farms between 2005 and 2009. All these isolates were identified as genotype Ia‐2; they divided however into two genetically distinct subgroups, that we name Pol I and Pol II. The Pol I isolates mainly caused outbreaks in the southern part of Poland, while Pol II isolates predominantly were sampled in the north of Poland, although it seems that they have been transmitted to other parts of the country. Molecular epidemiology was used for characterization of transmission pathways. This study shows that a main cause of virus transmission appears to be movement of fish. At least in Polish circumstances trading practices appear to have significant impact on spreading of VHSV infection.     

Global spread and evolution of viral haemorrhagic septicaemia virus

Viral haemorrhagic septicaemia virus (VHSV) is a rhabdovirus that infects over 48 species of teleosts and is lethal in many. VHSV threatens marine and aquatic fisheries. VHSV was first discovered outside Europe in 1988 in fish from the Pacific coast of North America. In 1994, VHSV was discovered in Newfoundland. In 2003, VHSV was isolated from fish in Lake St. Clair (Michigan and Ontario). In this study, we used 46 nucleotide sequences for the glycoprotein gene from 12 studies and 150 nucleotide sequences for the nucleoprotein gene from nine studies. We combined phylogenetics and a geographic information system to visualize the transmission paths of VHSV lineages. We also reconstructed the spread of VHSV lineages through optimization of geographic data for viral isolates on phylogenetic trees. We demonstrate that VHSV was transmitted from the North Atlantic Ocean and/or Baltic Sea to the Atlantic coast of North America and Japan in independent events. From the Atlantic coast, the virus was transmitted independently to the Laurentian Great Lakes and the Pacific coast of Canada and the contiguous United States. From the Pacific Northwest, the virus was transmitted to Asia and Alaska in independent events. These results clarify the debate ongoing in the literature on the geographic spread of VHSV.     

Persistence of Florida largemouth bass alleles in a northern Arkansas population of largemouth bass, Micropterus salmoides Lacèpéde

Abstract– Lake Ashbaugh, located in northeast Arkansas, was constructed in 1981, and initially stocked with Florida largemouth bass followed by supplemental stockings of northern largemouth bass. Allele frequencies of three discriminant allozyme loci (sAAT-B, sIDH-B, sMDH-B) between Florida and northern largemouth bass were determined for 414 largemouth bass collected between 1994 and 1996. F_x bass dominated our sample, with 62.3% possessing Florida largemouth bass alleles. A high incidence of Hardy-Weinberg disequilibrium was observed, indicative of genetic change within the population. No significant differences were identified for frequency of age classes, relative weight, and length at age between the northern, F₁ and F_x phenotypes. Despite being located north of what is generally considered suitable for stocking Florida largemouth bass, it was demonstrated that temperature is not selective at present against bass possessing Florida largemouth bass alleles. However, caution should prevail when introducing non-native stock into native gene complexes, as introduced genes persist through many generations.     

A mortality event in wrasse species (Labridae) associated with the presence of viral haemorrhagic septicaemia virus

Viral haemorrhagic septicaemia (VHS) is an infectious disease of farmed and wild fish and has an extensive host range in both freshwater and marine environments. In December 2012, a wrasse population consisting of ballan, Labrus bergylta (Ascanius), corkwing, Symphodus melops (L.), cuckoo, Labrus mixtus L., goldsinny, Ctenolabrus rupestris (L.), and rock cook, Centrolabrus exoletus (L.), held at a marine hatchery in the Shetland Isles, Scotland, experienced a mortality event. Approximately 10 000 wrasse were being held at the facility on behalf of an Atlantic salmon, Salmo salar L., aquaculture company prior to being deployed for the biological control of parasites on marine pen Atlantic salmon, aquaculture sites. Fish Health Inspectors from Marine Scotland Science initiated a diagnostic investigation, and subsequent diagnostic testing confirmed the site to be VHSV positive by qRT-PCR and virus isolation followed by ELISA. A VHSV genotype-specific qRT-PCR assay revealed that the isolates belonged to genotype III, the European marine strain of the virus. The virus genotype was further confirmed by nucleic acid sequencing of the partial nucleoprotein (N) and glycoprotein (G) genes followed by BLAST nucleotide searches. This study reports for the first time the detection of VHSV within multiple wrasse species and highlights the need for a comprehensive risk-based approach to the use of wrasse and other finfish species as biological controls within the aquaculture industry.     

Whole-genome next-generation sequencing and phylogenetic characterization of viral haemorrhagic septicaemia virus in Korea

Whole-genome next-generation sequencing was used to investigate the local evolution of viral haemorrhagic septicaemia virus, a serious pathogen affecting economically important fish such as rainbow trout and turbot in Europe and olive flounder in Asia. Sequence analysis showed that all isolates were genotype IVa, but could be classified further into four subgroups (K1–K4). In addition, genomic regions encompassing the nucleoprotein, phosphoprotein, matrix protein and non-virion protein genes, as well as the seven non-coding regions, were relatively conserved, whereas glycoprotein and RNA-dependent RNA polymerase genes were variable in the coding region. Taken together, the data demonstrate that whole-genome next-generation sequencing may be useful for future surveillance, prevention and control strategies against viral haemorrhagic septicaemia.     

Development and validation of a novel Taqman‐based real‐time RT‐PCR assay suitable for demonstrating freedom from viral haemorrhagic septicaemia virus

Viral haemorrhagic septicaemia (VHS) is a serious disease in several fish species. VHS is caused by the rhabdovirus viral haemorrhagic septicaemia virus (VHSV). To prevent spreading of the pathogen, it is important to use a fast, robust, sensitive and specific diagnostic tool to identify the infected fish. Traditional diagnosis based on isolation in cell culture followed by identification using, for example, ELISA is sensitive and specific but slow. By switching to RT‐PCR for surveillance and diagnosis of VHS the time needed before a correct diagnosis can be given will be considerably shortened and the need for maintaining expensive cell culture facilities reduced. Here we present the validation, according to OIE guidelines, of a sensitive and specific Taqman‐based real‐time RT‐PCR. The assay detects all isolates in a panel of 79 VHSV isolates covering all known genotypes and subtypes, with amplification efficiencies of approximately 100%. The analytical and diagnostic specificity of the real‐time RT‐PCR is close to 1, and the analytical and diagnostic sensitivity is comparable with traditional cell‐based methods. In conclusion, the presented real‐time RT‐PCR assay has the necessary qualities to be used as a VHSV surveillance tool on par with cell culture assays.     

Viral replication in excised fin tissues (VREFT) corresponds with prior exposure of Pacific herring, Clupea pallasii (Valenciennes), to viral haemorrhagic septicaemia virus (VHSV)

Procedures for a viral replication in excised fin tissue (VREFT) assay were adapted to Pacific herring, Clupea pallasii, and optimized both to reduce processing time and to provide the greatest resolution between naïve herring and those previously exposed to viral haemorrhagic septicaemia virus (VHSV), Genogroup IVa. The optimized procedures included removal of the left pectoral fin from a euthanized fish, inoculation of the fin with >10⁵ plaque‐forming units (PFU) mL^?1 VHSV for 1 h, rinsing the fin in fresh medium six times to remove unadsorbed virions, incubation of the fin in fresh medium for 4 days and enumeration of the viral titre in a sample of the incubation medium by plaque assay. The optimized VREFT assay was effective at identifying the prior exposure history of laboratory‐reared Pacific herring to VHSV. The geometric mean VREFT value was significantly greater (P < 0.01) among naïve herring (1.2 × 10³ PFU mL^?1) than among groups that survived exposure to VHSV (1.0–2.9 × 10² PFU mL^?1); additionally, the proportion of cultures with no detectable virus was significantly greater (P = 0.0002) among fish that survived exposure to VHSV (39–47%) than among naïve fish (3.3%). The optimized VREFT assay demonstrates promise for identifying VHSV exposure history and forecasting disease potential in populations of wild Pacific herring.     

Effects of season, maturity, and sex on lipid class dynamics in largemouth bass (Micropterus salmoides Lacepede)

Abstract Factors that can influence changes in lipid class concentrations in specific tissues have not been reported for largemouth bass Micropterus salmoides. We investigated patterns of lipid composition by measuring seasonal levels of total lipid and major lipid classes in the muscle and liver tissues of juvenile and adult largemouth bass and gonad tissue of adults. Fish were collected in December, February, April and July to correspond with key stages of the sexual cycle. Total lipid concentrations were substantially greater in hepatic (32.8 [female] to 56.6 [male] mg. g^?1) and gonadal tissues (34.4 [F] to 204.0 [M] mg. g^?1) than in muscle (7. 5 [F] to 11.4 [M] mg. g^?1). Muscle lipid concentrations in juveniles were significantly higher (10.9 [F] and 11.4 [M] mg. g^?1) in summer than in winter in adult muscle tissue (8.6 [F] and 9.4 [MI mg. g^?1). A 44% increase in ovary lipid was observed from preparatory to prespawn stages, followed by a 78% decline after the spawn. Additionally, seasonal hepatic lipid concentrations were substantially higher in juveniles. Concentrations of lipid classes (free cholesterol [C], triacylglycerol [TAG], free-fatty acids [FFA], esterified cholesterol [CE] and phospholipid [PL]) of all tissue types varied significantly among season, sex, and maturity categories. Changes with all lipid classes in the ovary coincided with those occurring in total ovarian lipid. Mobilization and transport of TAG and PL from the liver to the ovary appeared to occur during gametogenesis. All class concentrations in juveniles were invariably higher in liver than in muscle, but seasonally variable among hepatic and gonad tissues of adults. Juvenile males showed slightly greater hepatic C concentrations than other groups. Muscle and hepatic C were generally higher in juveniles.     

Experimental infection of turbot, Psetta maxima (L.), with strains of viral haemorrhagic septicaemia virus isolated from wild and farmed marine fish

The susceptibility of turbot, Psetta maxima, to infection with two strains of viral haemorrhagic septicaemia virus (VHSV) obtained from wild Greenland halibut, Reinhardtius hippoglossoides, and from farmed turbot was examined. A marine VHSV strain known to be highly pathogenic for turbot was also utilized for comparative purposes. Fish were infected by intra-peritoneal (i.p.), immersion or cohabitation, and maintained at two different temperatures (8 and 15 degrees C). Infection trials showed that the three VHSV isolates were pathogenic for turbot fingerlings by i.p. injection at both temperatures, with high levels of mortality. Virus was recovered from most pools of dead fish i.p. challenged, but not from surviving fish. Although clinical signs were not induced following waterborne exposure, viral growth was obtained from some pools of surviving fish challenged by immersion with strain GH40 from Greenland halibut, which indicates that the virus can survive in sea water and infect other fish via horizontal transmission. Furthermore, although low, the clinical signs and mortality observed in fish cohabitating with turbot challenged with strain GH40 confirms horizontal transmission and indicates that the passage through fish increases the virulence of this strain for turbot. These findings indicate that Greenland halibut, as other wild fish, may play an important role in the epizootiology of VHSV and suggest a potential risk for the turbot farming industry.     

Investigation of round goby viral haemorrhagic septicaemia outbreak in New York

Eleven viral haemorrhagic septicaemia virus (VHSV) genotype IVb isolates were sequenced, and their genetic variation explored to determine the source of a VHS outbreak on the eastern shore of Cayuga Lake. An active fish kill of round gobies (Neogobius melanostomus, Pallas) was intensively sampled at King Ferry, NY and nearby Long Point State Park in May 2017. Gross lesions observed on 67 moribund round gobies and two rock bass (Ambloplites rupestris, Rafinesque) included moderately haemorrhagic internal organs and erythematous areas on the head, flank, and fins. RT‐qPCR tests for VHSV were positive for all 69 fish. Viral isolation on epithelioma papulosum cyprinid cells showed cytopathic effect characteristic of VHSV for six round goby samples from King Ferry. The complete nucleotide sequence of the VHSV IVb genomes of five Cayuga Lake round goby isolates were derived on an Illumina platform along with 2017 VHSV IVb isolates from round gobies collected from the following: Lake Erie near Dunkirk, NY; the St. Lawrence River near Clayton and Cape Vincent, NY; and Lake St. Lawrence near Massena, NY. The phylogenetic tree created from these aligned sequences and four other complete VHSV IVb genomes shows Cayuga Lake isolates are closely related to the Lake Erie isolates.     

Diet of the largemouth bass, Micropterus salmoides (Lacepède), in Lake Naivasha, Kenya

Abstract The major food items of adult largemouth bass, Micropterus salmoides (Lacepède), in Lake Naivasha, Kenya are not markedly different from those in its native range. Although insects and their larvae are major components of the diet for both temperate and equatorial populations of juvenile bass, the equatorial population also eats Procambarus clarkii (Girard), juvenile fish and aquatic weeds. In temperate populations they also eat crustaceans, rotifers and oligochaetes. The frequency of occurrence of the major prey organisms in the stomachs of M. salmoides varies seasonally with population peaks in these organisms. Similar to North American populations, the Naivasha population feeds during the day between dawn and dusk. In contrast to temperate populations, the population of M. salmoides in this equatorial lake feeds throughout the year, with feeding intensity correlated with water temperature.