Infection dynamics and tissue tropism of Parvicapsula pseudobranchicola (Myxozoa: Myxosporea) in farmed Atlantic salmon (Salmo salar).

BACKGROUND: The myxosporean parasite Parvicapsula pseudobranchicola commonly infects farmed Atlantic salmon in northern Norway. Heavy infections are associated with pseudobranch lesions, runting and mortality in the salmon populations. The life-cycle of the parasite is unknown, preventing controlled challenge experiments. The infection dynamics, duration of sporogony, tissue tropism and ability to develop immunity to the parasite in farmed Atlantic salmon is poorly known. We conducted a field experiment, aiming at examining these aspects. METHODS: Infections in a group of Atlantic salmon were followed from before sea-transfer to the end of the production (604 days). Samples from a range of tissues/sites were analysed using real-time RT-PCR and histology, including in situ hybridization. RESULTS: All salmon in the studied population rapidly became infected with P. pseudobranchicola after sea-transfer medio August. Parasite densities in the pseudobranchs peaked in winter (November-January), and decreased markedly to March. Densities thereafter decreased further. Parasite densities in other tissues were low. Parasite stages were initially found to be intravascular in the pseudobranch, but occurred extravascular in the pseudobranch tissue at 3 months post-sea-transfer. Mature spores appeared in the pseudobranchs in the period with high parasite densities in the winter (late November-January), and were released (i.e. disappeared from the fish) in the period January-March. Clinical signs of parvicapsulosis (December-early February) were associated with high parasite densities and inflammation in the pseudobranchs. No evidence for reinfection was seen the second autumn in sea. CONCLUSIONS: The main site of the parasite in Atlantic salmon is the pseudobranchs. Blood stages occur, but parasite proliferation is primarily associated with extravascular stages in the pseudobranchs. Disease and mortality (parvicapsulosis) coincide with the completion of sporogony. Atlantic salmon appears to develop immunity to P. pseudobranchicola. Further studies should focus on the unknown life-cycle of the parasite, and the pathophysiological effects of the pseudobranch infection that also could affect the eyes and vision.

Phenotypic and genetic characterization of Piscirickettsia salmonis from Chilean and Canadian salmonids.

BACKGROUND: The study presents the phenotypic and genetic characterization of selected P. salmonis isolates from Atlantic salmon and rainbow trout suffering from SRS (salmonid rickettsial septicemia) in Chile and in Canada. The phenotypic characterization of the P. salmonis isolates were based on growth on different agar media (including a newly developed medium), different growth temperatures, antibiotics susceptibility and biochemical tests. RESULTS: This is the first study differentiating Chilean P. salmonis isolates into two separate genetic groups. Genotyping, based on 16S rRNA-ITS and concatenated housekeeping genes grouped the selected isolates into two clades, constituted by the Chilean strains, while the Canadian isolates form a branch in the phylogenetic tree. The latter consisted of two isolates that were different in both genetic and phenotypic characteristics. The phylogenies and the MLST do not reflect the origin of the isolates with respect to host species. The isolates included were heterogeneous in phenotypic tests. CONCLUSIONS: The genotyping methods developed in this study provided a tool for separation of P. salmonis isolates into distinct clades. The SRS outbreaks in Chile are caused by minimum two different genetic groups of P. salmonis. This heterogeneity should be considered in future development of vaccines against this bacterium in Chile. Two different strains of P. salmonis, in regards to genetic and phenotypic characteristics, can occur in the same contemporary outbreak of SRS.

Variable Number of Tandem Repeats (VNTR) analysis of Flavobacterium psychrophilum from salmonids in Chile and Norway.

BACKGROUND: Flavobacterium psychrophilum causes serious fish diseases such RTFS and BCWD, affecting the aquaculture industry worldwide. Commercial vaccines are not available and control of the disease depends on the use of antibiotics. Reliable methods for detection and identification of different isolates of this bacterium could play an important role in the development of good management strategies. The aim of this study was to identify genetic markers for discrimination between isolates. A selection of eight VNTRs from 53 F. psychrophilum isolates from Norway, Chile, Denmark and Scotland were analyzed. The results were compared with previous work on the same pathogen using MLST for genetic differentiation. RESULTS: The VNTR analysis gave a separation between the F. psychrophilum isolates supporting the results of previous MLST work. A higher diversity was found among the Chilean isolates compared to those from Norway, which suggests a more homogenous reservoir in Norway. Transgenerational transmission of F. psychrophilum from other countries, exporting salmon embryos to Chile, may explain the differences in diversity. The same transmission mechanisms could also explain the wide geographical distribution of identical isolates in Norway. But, this could also be a result of movement of smolts and embryos. The selected VNTRs are stable genetic markers and no variation was observed after several passages on agar plates at different temperatures. CONCLUSIONS: These VNTRs are important additions for genotyping of F. psychrophilum isolates. Future studies on VNTRs of F. psychrophilum should include isolates from more host species from a wider geographical area. To get a more robust genotyping the VNTRs should be used in concert with MLST. Future studies of isolates with high and low virulence should focus on identifying virulence markers using VTNRs and MLST.

Multiple-locus, variable number of tandem repeat analysis (MLVA) of the fish-pathogen Francisella noatunensis.

BACKGROUND: Since Francisella noatunensis was first isolated from cultured Atlantic cod in 2004, it has emerged as a global fish pathogen causing disease in both warm and cold water species. Outbreaks of francisellosis occur in several important cultured fish species making a correct management of this disease a matter of major importance. Currently there are no vaccines or treatments available. A strain typing system for use in studies of F. noatunensis epizootics would be an important tool for disease management. However, the high genetic similarity within the Francisella spp. makes strain typing difficult, but such typing of the related human pathogen Francisella tullarensis has been performed successfully by targeting loci with higher genetic variation than the traditional signature sequences. These loci are known as Variable Numbers of Tandem Repeat (VNTR). The aim of this study is to identify possible useful VNTRs in the genome of F. noatunensis. RESULTS: Seven polymorphic VNTR loci were identified in the preliminary genome sequence of F. noatunensis ssp. noatunensis GM2212 isolate. These VNTR-loci were sequenced in F. noatunensis isolates collected from Atlantic cod (Gadus morhua) from Norway (n = 21), Three-line grunt (Parapristipoma trilineatum) from Japan (n = 1), Tilapia (Oreochromis spp.) from Indonesia (n = 3) and Atlantic salmon (Salmo salar) from Chile (n = 1). The Norwegian isolates presented in this study show both nine allelic profiles and clades, and that the majority of the farmed isolates belong in two clades only, while the allelic profiles from wild cod are unique. CONCLUSIONS: VNTRs can be used to separate isolates belonging to both subspecies of F. noatunensis. Low allelic diversity in F. noatunensis isolates from outbreaks in cod culture compared to isolates wild cod, indicate that transmission of these isolates may be a result of human activity. The sequence based MLVA system presented in this study should provide a good starting point for further development of a genotyping system that can be used in studies of epizootics and disease management of francisellosis.