In this work, we described the first isolation of Streptococcus iniae in red porgy, (Pagrus pagrus), as well as, the first isolation of this pathogen in Europe affecting gilthead seabream (Sparus aurata). In both farmed fish species, the disease showed lethargy, anorexia, abnormal swimming, exophthalmia and sudden death, with mortality rates over 25% in red porgy, and 10% in gilthead seabream. β-hemolytic grampositive cocci, catalase negative and oxidase negative, was isolated in pure culture from internal organs. Conventional and rapid identification systems, and 16S rRNA gene partial sequencing were used to identify the causative agent of the natural disease death. LD50 trials were carried out to show the virulence of this isolated strain in these species, red porgy and gilthead seabream, showing values from 1.7x104 cfu/fish and 1.32x105 cfu/fish respectively, as well as chronic infection and cohabitation trials. Most prominent lesions were meningoencephalitis and multi focal infiltration of macrophages cells in kidney and spleen. Gentamicin protection assays and fluorescence microscopy were used to investigate the ability of Streptococcus iniae strains to adhere and invade the fish fibroblastic cell line SAF-1 derived from Sparus aurata. All strains tested were detected intracellularly, though internalisation levels varied among strains. Experiments carried out at 4ºC demonstrated that active cell metabolism are necessary for bacterial internalisation. Intracellular bacteria were detected up to 3 days indicating the possibility that some bacterial cells remain viable inside SAF-1 cells. Our in vitro findings indicate that Streptococcus iniae are capable of adhering, entering and surviving within the nonphagocytic fibroblastic cell line SAF-1 wich may be important for persistence and establishment of a carrier state in marine fish species.

Máster Oficial en Cultivos Marinos. VI Máster Internacional en Acuicultura. Trabajo presentado como requisito parcial para la obtención del Título de Máster Oficial en Cultivos Marinos, otorgado por la Universidad de Las Palmas de Gran Canaria (ULPGC), el Instituto Canario de Ciencias Marinas (ICCM), y el Centro Internacional de Altos Estudios Agronómicos Mediterráneos de Zaragoza (CIHEAM). Premio al mejor trabajo de investigación del CIHEAM 2011