Current vaccination strategy against Piscirickettsia salmonis in Chile based only on the EM-90 genogroup shows incomplete cross-protection for the LF-89 genogroup.

Piscirickettsia salmonis, the primary bacterial disease in Chilean salmon farming, necessitates a constant refinement of control strategies. This study hypothesized that the current vaccination strategy for SRS control in the Chilean Atlantic salmon aquaculture industry, which has been in place since 2017 (ALPHA JECT® 5.1 plus LiVac®), solely relies on vaccines formulated with the EM-90 genogroup of P. salmonis (PS-EM-90), triggering a partial cross-immunity response in fish infected with the LF-89 genogroup (PS-LF-89). Relative Percent Survival (RPS) and cell-mediated immune (CMI) response were evaluated in Atlantic salmon post-smolts vaccinated with the standard vaccination strategy but challenged with both PS-EM-90 and PS-LF-89, in addition to other vaccination strategies considering primo vaccination and booster with other commercial vaccines and the possible enhancing effects of the combination with a natural immunomodulator (PAQ-Xtract®) administered orally. The intraperitoneal (I.P.) challenge was performed after 2395°-days (DD) after the start of the immunostimulant delivery, 1905 DD after the primo vaccination, and 1455 DD after the booster vaccination. Unvaccinated fish showed 73.6 and 41.7 % mortality when challenged with PS-EM-90 and PS-LF-89, respectively. Fish infected with PS-LF-89 died significantly faster (21 days post-infection, dpi) than fish challenged with PS-EM-90 (28 dpi) (p = 0.0043) and had a higher probability of death (0.4626) than fish challenged with PS-EM-90. RPS had a significant positive correlation with the PS-EM-90 load of the P. salmonis genogroup (r = 0.540, p < 0.01) but not with the PS-LF-89 load (r = 0.155, p > 0.05). This demonstrated that the immunization strategies were more effective in lowering PS-EM-90 loads, resulting in higher survival rates in fish challenged with PS-EM-90. The current industry vaccination strategy recorded a 100 % RPS when fish were challenged with PS-EM-90, but the RPS dropped significantly to 77 % when fish were challenged with PS-LF-89, meaning that the strategy did not show complete cross-protection. But after adding PAQ-Xtract®, the RPS improved from 77 % to 92 % in fish that were vaccinated with the standard method but then challenged with PS-LF-89. The most effective vaccination strategy was based on LiVac® as primo vaccination and ALPHA JECT® 5.1 plus LiVac® as booster vaccination, with or without PAQ-Xtract®, in both PS-EM-90 (100 %) and PS-LF-89 (96 %) challenged fish. The serum concentration of anti-P. salmonis IgM did not show a correlation with the protection of immunization strategies expressed in survival. Low serum IL-12 and high serum IFNγ concentrations showed a correlation with higher bacterial loads and lower survival. Aggregate analysis showed a significant correlation between higher numbers of CD8+ cells in the head-kidney, higher fish survival, and a lower bacterial load. The immunization strategies were safe for fish and induced only mild microscopic lesions in the gut. Taken together, our results help to better understand the biological interaction between P. salmonis and post-smolt vaccinated Atlantic salmon to deepen the knowledge on vaccine-induced protection, CMI immune response, and cross-immunity applied to improve the current immunization strategy for SRS control in the Chilean salmon industry.

First Report of Aeromonas veronii as an Emerging Bacterial Pathogen of Farmed Nile Tilapia (Oreochromis niloticus) in Brazil.

Brazil is one of the world's leading producers of Nile tilapia, Oreochromis niloticus. However, the industry faces a major challenge in terms of infectious diseases, as at least five new pathogens have been formally described in the last five years. Aeromonas species are Gram-negative anaerobic bacteria that are often described as fish pathogens causing Motile Aeromonas Septicemia (MAS). In late December 2022, an epidemic outbreak was reported in farmed Nile tilapia in the state of São Paulo, Brazil, characterized by clinical signs and gross pathology suggestive of MAS. The objective of this study was to isolate, identify, and characterize in vitro and in vivo the causative agent of this epidemic outbreak. The bacterial isolates were identified as Aeromonas veronii based on the homology of 16S rRNA (99.9%), gyrB (98.9%), and the rpoB gene (99.1%). A. veronii showed susceptibility only to florfenicol, while it was resistant to the other three antimicrobials tested, oxytetracycline, enrofloxacin, and amoxicillin. The lowest florfenicol concentration capable of inhibiting bacterial growth was ≤0.5 µg/mL. The phenotypic resistance of the A. veronii isolate observed for quinolones and tetracycline was genetically confirmed by the presence of the qnrS2 (colE plasmid) and tetA antibiotic-resistant genes, respectively. A. veronii isolate was highly pathogenic in juvenile Nile tilapia tested in vivo, showing a mortality rate ranging from 3 to 100% in the lowest (1.2 × 104) and highest (1.2 × 108) bacterial dose groups, respectively. To our knowledge, this study would constitute the first report of highly pathogenic and multidrug-resistant A. veronii associated with outbreaks and high mortality rates in tilapia farmed in commercial net cages in Brazil.

Co-Infection by LF-89-Like and EM-90-Like Genogroups of Piscirickettsia Salmonis in Farmed Atlantic Salmon in Chile: Implications for Surveillance and Control of Piscirickettsiosis.

Piscirickettsiosis (SRS), caused by Piscirickettsia salmonis, is the main infectious disease that affects farmed Atlantic salmon in Chile. Currently, the official surveillance and control plan for SRS in Chile is based only on the detection of P. salmonis, but neither of its genogroups (LF-89-like and EM-90-like) are included. Surveillance at the genogroup level is essential not only for defining and evaluating the vaccination strategy against SRS, but it is also of utmost importance for early diagnosis, clinical prognosis in the field, treatment, and control of the disease. The objectives of this study were to characterize the spatio-temporal distribution of P. salmonis genogroups using genogroup-specific real-time probe-based polymerase chain reaction (qPCR) to discriminate between LF-89-like and EM-90-like within and between seawater farms, individual fish, and tissues/organs during early infection in Atlantic salmon under field conditions. The spatio-temporal distribution of LF-89-like and EM-90-like was shown to be highly variable within and between seawater farms. P. salmonis infection was also proven to be caused by both genogroups at farm, fish, and tissue levels. Our study demonstrated for the first time a complex co-infection by P. salmonis LF-89-like and EM-90-like in Atlantic salmon. Liver nodules (moderate and severe) were strongly associated with EM-90-like infection, but this phenotype was not detected by infection with LF-89-like or co-infection of both genogroups. The detection rate of P. salmonis LF-89-like increased significantly between 2017 and 2021 and was the most prevalent genogroup in Chilean salmon aquaculture during this period. Lastly, a novel strategy to identify P. salmonis genogroups based on novel genogroup-specific qPCR for LF-89-like and EM-90-like genogroups is suggested.

Reference Intervals for Blood Biomarkers in Farmed Atlantic Salmon, Coho Salmon and Rainbow Trout in Chile: Promoting a Preventive Approach in Aquamedicine.

The mission of veterinary clinical pathology is to support the diagnostic process by using tests to measure different blood biomarkers to support decision making about farmed fish health and welfare. The objective of this study is to provide reference intervals (RIs) for 44 key hematological, blood biochemistry, blood gasometry and hormones biomarkers for the three most economically important farmed salmonid species in Chile (Atlantic salmon, coho salmon and rainbow trout) during the freshwater (presmolt and smolt age range) and seawater stages (post-smolt and adult age range). Our results confirmed that the concentration or activity of most blood biomarkers depend on the salmonid species, age range and/or the interaction between them, and they are often biologically related to each other. Erythogram and leukogram profiles revealed a similar distribution in rainbow trout and coho salmon, but those in Atlantic salmon were significantly different. While the activity of the most clinically important plasma enzymes demonstrated a similar profile in Atlantic salmon and rainbow trout, coho salmon demonstrated a significantly different distribution. Plasma electrolyte and mineral profiles showed significant differences between salmonid species, especially for rainbow trout, while Atlantic salmon and coho salmon demonstrated a high degree of similarity. Furthermore, electrolytes, minerals and blood gasometry biomarkers were significantly different between age ranges, suggesting a considerably different distribution between freshwater and seawater-farmed fish. The RIs of clinically healthy fish described in this study take into account the high biological variation of farmed fish in Chile, as the 3.059 individuals came from 78 different fish farms, both freshwater and seawater, and blood samples were collected using the same pre-analytical protocol. Likewise, our study provides the Chilean salmon farming industry with standardized protocols that can be used routinely and provides valuable information to improve the preventive vision of aquamedicine through the application of blood biomarkers to support and optimize health, welfare and husbandry management in the salmon farming industry.

Atlantic Salmon Pre-smolt Survivors of Renibacterium salmoninarum Infection Show Inhibited Cell-Mediated Adaptive Immune Response and a Higher Risk of Death During the Late Stage of Infection at Lower Water Temperatures.

Bacterial kidney disease (BKD) is widespread in many areas of the world and can cause substantial economic losses for the salmon aquaculture industry. The objective of this study was to investigate the pathophysiological response and gene expression profiles related to the immune response at different water temperatures and to identify the best immunopathological biomarkers to define a phenotype of resistance to BKD. The abundance of msa transcripts of R. salmoninarum in the head kidney was significantly higher in infected fish at 11°C. R. salmoninarum induced significantly more severe kidney lesions, anemia and impaired renal function at 11°C. In addition, the expression pattern of the genes related to humoral and cell-mediated immune responses in infected fish at 11 and 15°C was very similar, although R. salmoninarum induced a significantly greater downregulation of the adaptive immune response genes at the lower water temperature. These results could be due to a suppressed host response directly related to the lowest water temperature and/or associated with a delayed host response related to the lowest water temperature. Although no significant differences in survival rate were observed, fish infected at the lowest temperature showed a higher probability of death and delayed the mortality curve during the late stage of infection (35 days after infection). Thirty-three immunopathological biomarkers were identified for potential use in the search for a resistance phenotype for BKD, and eight were genes related specifically to the adaptive cell-mediated immune response.