The European sea bass Dicentrarchus labrax is the main species reared in Mediterranean aquaculture. Its larval stage, which is very sensitive and highly affected by sanitary and environmental conditions, is particularly scrutinized in hatcheries. Recently, a Mediterranean sea bass farm had to deal with an abnormal increase in mortality, especially between 20 and 35 days post-hatching (dph). Biological investigations led to the observation of cytopathic effects on three different fish cell lines after almost 3 weeks of culture at 14 • C in contact with homogenized affected larvae, suggesting the presence of a viral agent. High-throughput sequencing revealed a 6818-nucleotide-long RNA genome with six putative ORFs, corresponding to the organization of viruses belonging to the Totiviridae family. This genome clustered with the newly described and suggested Pistolvirus genus, sharing 45.5% to 37.2% nucleotide identity with other piscine toti-like viruses such as Cyclopterus lumpus toti-like virus (CLuTLV) or piscine myocarditis virus (PMCV), respectively. Therefore, we propose to name this new viral agent sea bass toti-like virus (SBTLV). Specific real-time RT-PCR confirmed the presence of the viral genome in the affected larval homogenate from different production batches and the corresponding cell culture supernatant. Experimental infections performed on sea bass fingerlings did not induce mortality, although the virus could be detected in various organs and a specific immune response was developed. Additional studies are needed to understand the exact involvement of this virus in the mortality observed in hatcheries and the potential associated cofactors.