Salmonid rickettsial septicemia (SRS) is one of the major diseases affecting the salmon industry in Chile . The intracellular bacterium Piscirickettsia salmonis is the responsible pathogen of SRS that can infect multiple tissues in its host. New approaches for its control are considered a significant challenge in the scientific community. Advances in high-throughput sequencing technologies allow a better understanding of the transcriptomic responses of organisms in several biological scenarios, such as pathogen-host interaction. In this sense, miRNAs play an essential role in the transcriptomic response of Salmo salar during infection with P. salmonis , promoting a change in the diversity of miRNA families. Also, miRNAs co-modulate the transcriptional activity of their target genes, suggesting a putative function of non-coding RNAs in the immune response of salmon infected with an intracellular pathogen . This study aimed to identify candidates for small non-coding RNA (sRNA) involved in the pathogenesis process of P. salmonis during infection in S. salar and validate at a functional level the genomic modulation of these sRNA at in vitro model of P. salmonis infection .
Transcriptome of experimental infections with P. salmonis EM-90 wild and attenuated in Atlantic salmon was used for sRNA candidate selection . First, putative sRNAs P. salmonis binding sites in up and down-regulated S. salar genes during infection were predicted using RNA22 version 2.0 software. Then, two sRNA were selected based on RNAseq analysis expression, synthesized as mimics (mir-222, mir 143-37), and co-transfected with the GFP reporter gene in the salmon head kidney SHK-1 cell line . After 48h, the modified SHK-1 cells were infected with 1x106 CFU/mL P. salmonis. Cytotoxicity and cytopathic effects were monitored 24 hours after infection.
The experimental group transfected with mir-143- 37 showed lower cytotoxicity against infection with P. salmonis compared to the other groups tested (Table 1). Moreover , quantitative PCR analysis indicated regulation of transcription of immune-related genes in SHK-1 groups transfected with mimics. In addition , an increase in the relative expression of the E3 ubiquitin-protein ligase CBL-like gene was observed, suggesting that mir-143-37 could be intervening in the regulation of ubiquitination processes in salmon cells.
Further studies will be necessary to validate these results . Nevertheless, our study suggests that the pathogenicity process of P. salmonis could be regulated through s RNA. T his study highlights the potential use of s RNA as a bacterial attenuation technique.
Funding: ANID-Chile through and FONDAP (#15110027) and Ph.D. Scholarship, No. 21191482