Insights on immune and inflammatory responses of rainbow trout (Oncorhynchus mykiss) affected by Lactococcus garvieae or submitted to vaccination against lactococcosis

Lactococcosis, caused by the Gram-positive bacterium Lactococcus garvieae, is a major concern in Italian rainbow trout (Oncorhynchus mykiss) aquaculture. Basins are periodically affected by outbreaks, especially during the summer/fall months, and farmers resort to strenuous vaccination campaigns in order to limit the infection. The present PhD project aimed at summarizing and improving the knowledge on the response of rainbow trout to L. garvieae by evaluating the immune/inflammatory reactivity after natural or experimental infections, as well as upon “in lab” or “in field” vaccinations. The understanding of host-pathogen or host-vaccine interactions implied the analysis of humoral/cellular immune parameters and the study of immune-genes expression. Firstly, a bibliographic revision was drafted as introduction of the doctoral thesis (Chapter 1), which describes the extant knowledge on the immune response of rainbow trout affected by lactococcosis or submitted to vaccination and identified areas that require further investigations and dedicated studies. Particular emphasis has been placed to describe the current understanding on how immune parameters and inflammatory mechanisms are affected in rainbow trout individuals when they are clinically infected or examined after the vaccination. We performed the immune profiling of trout (vaccinated and unvaccinated) (Chapter 2) naturally exposed to L. garvieae by assessing classical hematological/serological immune parameters like leukogram, serum lysozyme activity, peroxidase activity, antiprotease activity, bactericidal activity, total proteins, total IgM, and specific antibodies to L. garvieae. This study provided insights on the outcome of the vaccination campaign as vaccinated fish showed higher level of specific anti-L. garvieae IgM, total protein and IgM, and a lower level of peroxidase and antiprotease activity, compared to both unvaccinated asymptomatic and symptomatic fish. Another study (Chapter 3) was dedicated to the expression of immune related genes in head kidney of infected versus healthy fish, as well as to explore the Near Infrared Spectroscopy (NIR) (SCiO molecular sensor) as a novel approach to discriminate between them. SCiO readings allowed detection of two different spectral populations potentially ascribable to asymptomatic and symptomatic. The immune genes modulation highlighted how the progression of a L. garvieae infection is characterised by a specific gene expression pattern, which is crucial in the mode of action of trout immune system against the bacterium. Then we evaluated the potential of the portable SCiO sensor to assess the health status of rainbow trout (Chapter 4). In this study the tool was found to discriminate between healthy and sick fish [sensibility 0.95 (0.75-1.00 I.C. 95%), specificity 0.80 (0.56-0.94 I.C. 95%)]. By virtue of rapid acquisition, low cost and use on site, the SCiO might represent a promising prognostic tool for monitoring rainbow trout health in aquaculture. Finally, a multi-disciplinary study has been performed (Chapter 5) throughout a spontaneous episode of lactococcosis in farmed rainbow trout, including histology, classical and molecular-based bacteriology, blood analysis and serology, immune genes expression, specifically addressing the better comprehension of bacterial spreading within the fish body and the fish inflammatory reactivity against this harmful biological stimulus. Bacteria were found in blood smears of 8/20 (40%) clinical individuals and no bacteria were detected in asymptomatic fish while L. garvieae DNA was present in 18/20 (90%) symptomatic fish and in 2/20 (10%) asymptomatic fish. Symptomatic fish showed a higher peroxidase activity and a lower lysozyme activity and specific antibodies (IgM to L. garvieae) compared to non-clinical fish (P≤0.05). Among several immune genes, IL-8 showed the most relevant up-regulation in symptomatic group compared to the asymptomatic one (P<0.001).