Iron deprivation is a nutritional immunity mechanism through which fish can limit the amount of iron available to invading bacteria. The aim of this study was to evaluate the modulation of iron metabolism genes in the liver and brain of sub-Antarctic Notothenioid Eleginops maclovinus challenged with Piscirickettsia salmonis. The specimens were inoculated with two P. salmonis strains: LF-89 (ATCC® VR-1361™) and Austral-005 (antibiotic resistant). Hepatic and brain samples were collected at intervals over a period of 35 days. Gene expression (by RT-qPCR) of proteins involved in iron storage, transport, and binding were statistically modulated in infected fish as compared to control counterparts. Specifically, the expression profiles of the transferrin and hemopexin genes in the liver, as well as the expression profiles of ferritin-M, ferritin-L, and transferrin in the brain, were similar for both experimental groups. Nevertheless, the remaining genes such as ferritin-H, ceruloplasmin, hepcidin, and haptoglobin presented tissue-specific expression profiles that varied in relation to the injected bacterial strain and sampling time-point. These results suggest that nutritional immunity could be an important immune defense mechanism for E. maclovinus against P. salmonis injection. This study provides relevant information for understanding iron metabolism of a sub-Antarctic Notothenioid fish.

Fluctuations in ambient temperature along with the presence of pathogenic microorganisms can induce important cellular changes that alter the homeostasis of ectothermic fish. The aim of this study was to evaluate how sudden or gradual changes in environmental temperature together with the administration of Piscirickettsia salmonis modulate the transcription of genes involved in cellular stress response in the liver of Eleginops maclovinus. Fish were subjected to the following experimental conditions in duplicate: C− 12 °C: Injection only with culture medium, C+ 12 °C: Injection with P. salmonis, AM 18 °C: Injection only with culture medium under acclimation at 18 °C, AB 18 °C: Injection with P. salmonis under acclimation at 18 °C, SM 18 °C: Injection only with culture medium and thermal shock at 18 °C and SB 18 °C: Injection with P. salmonis and thermal shock at 18 °C and sampling at 4-, 8-, 12-, 16- and 20-day post injection (dpi). The genes implied in the heat shock response (HSP70, HSC70, HSP90, and GRP78), apoptosis pathway (BAX and SMAC/Diablo), ubiquitination (E2, E3, ubiquitin, and CHIP), and 26 proteasome complex (PSMB7, PSMC1, and PSMA2) showed expression profiles dependent on time and type of injection applied. All the genes greatly increased their expression levels at day 16 and showed moderate increases at day 20, except for PSMA2 which showed a higher increase between 4- and 12-day post challenges. Our results suggest that the changes observed at the final days of the experiment are due to temperature more than P. salmonis.

The innate immune system can limit the growth of invading pathogens by depleting micronutrients at a cellular and tissue level. However, it is not known whether nutrient depletion mechanisms discriminate between living pathogens (which require nutrients) and pathogen-associated molecular patterns (PAMPs) (which do not). We stimulated SHK-1 cells with different PAMPs (outer membrane vesicles of Piscirickettsia salmonis “OMVs”, protein extract of P. salmonis “TP” and lipopolysaccharides of P. salmonis “LPS”) isolated from P. salmonis and evaluated transcriptional changes in nutritional immunity associated genes. Our experimental treatments were: Control (SHK-1 stimulated with bacterial culture medium), OMVs (SHK-1 stimulated with 1μg of outer membrane vesicles), TP (SHK-1 stimulated with 1μg of total protein extract) and LPS (SHK-1 stimulated with 1μg of lipopolysaccharides). Cells were sampled at 15-, 30-, 60- and 120-minutes post-stimulation. We detected increased transcription of zip8, zip14, irp1, irp2 and tfr1 in all three experimental conditions and increased transcription of dmt1 in cells stimulated with OMVs and TP, but not LPS. Additionally, we observed generally increased transcription of ireg-1, il-6, hamp, irp1, ft-h and ft-m in all three experimental conditions, but we also detected decreased transcription of these markers in cells stimulated with TP and LPS at specific time points. Our results demonstrate that SHK-1 cells stimulated with P. salmonis PAMPs increase transcription of markers involved in the transport, uptake, storage and regulation of micronutrients such as iron, manganese and zinc.