Transcriptomic analysis reveals a Piscirickettsia salmonis-induced early inflammatory response in rainbow trout skeletal muscle.

Skeletal muscle is the most abundant tissue in teleosts and is essential for movement and metabolism. Recently, it has been described that skeletal muscle can express and secrete immune-related molecules during pathogen infection. However, the role of this tissue during infection is poorly understood. To determine the immunocompetence of fish skeletal muscle, juvenile rainbow trout (Oncorhynchus mykiss) were challenged with Piscirickettsia salmonis strain LF-89. P. salmonis is the etiological agent of piscirickettsiosis, a severe disease that has caused major economic losses in the aquaculture industry. This gram-negative bacterium produces a chronic systemic infection that involves several organs and tissues in salmonids. Using high-throughput RNA-seq, we found that 60 transcripts were upregulated in skeletal muscle, mostly associated with inflammatory response and positive regulation of interleukin-8 production. Conversely, 141 transcripts were downregulated in association with muscle filament sliding and actin filament-based movement. To validate these results, we performed in vitro experiments using rainbow trout myotubes. In myotubes coincubated with P. salmonis strain LF-89 at an MOI of 50, we found increased expression of the proinflammatory cytokine il1b and the pattern recognition receptor tlr5s 8 and 12 h after infection. These results demonstrated that fish skeletal muscle is an immunologically active organ that can implement an early immunological response against P. salmonis.

Effect of cortisol on the immune-like response of rainbow trout (Oncorhynchus mykiss) myotubes challenged with Piscirickettsia salmonis.

Salmonids are a species of high commercial value in Chilean aquaculture, where muscle is the final product of the industry. Fish can be affected by stress during intensive cultures, increasing susceptibility to infections. Recently, we reported that muscle is an important focus of immune reactions. However, studies have shown the immunosuppressive effect of stress only in lymphoid organs, and few studies have been conducted on muscle and immunity. Hence, we determine the effects of cortisol on the immune-like response of fish myotubes challenged with Piscirickettsia salmonis by three trials. First, rainbow trout primary culture of muscle was cultured and treated with cortisol (100 ng/mL) for 3 and 4 h. Second, myotubes were challenged with P. salmonis (MOI 50) for 4, 6 and 8 h. And third, muscle cell cultures were pretreated with cortisol and then challenged with P. salmonis. The mRNA levels of glucocorticoid pathway and innate immunity were evaluated by qPCR. Cortisol increased the klf15 levels and downregulated the innate immune-related tlr5m gene and antimicrobial peptides. P. salmonis challenge upregulated several immune-related genes. Finally, cortisol pretreatment followed by P. salmonis challenge differentially modulated stress- and immune-related genes. These data suggest that fish muscle cells possess an intrinsic immune response and are differentially regulated by cortisol, which could lead to bacterial outbreaks in muscle under stress conditions.

Genome-scale metabolic reconstruction for the insidious bacterium in aquaculture Piscirickettsia salmonis.

Piscirickettsia salmonis is a fish bacterium that causes the disease piscirickettsiosis in salmonids. This pathology is partially controlled by vaccines. The lack of knowledge has hindered its culture on laboratory and industrial scale. The study describes the metabolic phenotype of P. salmonis in culture. This study presents the first genome-scale model (iPF215) of the LF-89 strain of P. salmonis, describing the central metabolic pathway, biosynthesis and molecule degradation and transport mechanisms. The model was adjusted with experiment data, allowing the identification of the capacities that were not predicted by the automatic annotation of the genome sequences. The iPF215 model is comprised of 417 metabolites, 445 reactions and 215 genes, was used to reproduce the growth of P. salmonis (µmax 0.052±0.005h-1). The metabolic reconstruction of the P. salmonis LF-89 strain obtained in this research provides a baseline that describes the metabolic capacities of the bacterium and is the basis for developing improvements to its cultivation for vaccine formulation.