Kuruma shrimp (Penaeus japonicus) is a commercially important Penaeid species widely cultured, and highly valued seafood in Japan. However, diseases are limiting factors for P. japonicus resulting in massive mortalities and low production output. In August and September 2022, two disease outbreaks were observed in culture farms in Okinawa, Japan. Diseased animals exhibited muscle whitening extending from the abdominal to the distal region. This study reports the first disease occurrence in Kuruma shrimp associated with Photobacterium damselae subsp. damselae (Pdd) having whitened musculature as a major clinical sign.
After obtaining the RNA and DNA shotgun sequences, we identified nonhost contigs showing significant taxonomic alignment to bacterial species. This led to the investigation of a bacterium as a possible causative agent, which later we characterized its population in the diseased shrimp muscle. The majority of the 16S rRNA sequence recombinant clones had their highest homology with Photobacterium sp. (> 99%) which isolation of any of its species was pursued. The three bacterial isolates (WMD-P1, WMD-P2, and WMD-P3) from the whitened muscle tissue identified as Photobacterium damselae subsp. damselae (Pdd) were further characterized.
Their genomes consisted of two circular chromosomes with varying numbers of plasmid. Its size ranges from 4.47 Mb to 4.60 Mb with an average GC content of 40.8%, with the predicted number of coding sequences (CDs) ranging from 3816 to 4031. From the genomes, two reported virulence genes encoding for leukocidin pore-forming toxin (hlyA) and phospholipase (pldA) were identified.
Putative virulence factors are involved in adherence, antiphagocytosis, chemotaxis and motility, iron uptake, quorum sensing, secretion system, and immune evasion. Also, the presence of prophages, genomic islands, and antimicrobial-resistant genes suggests episodes of horizontal gene transfer. Average nucleotide identity (ANI) and pangenome analyses revealed a high genetic relationship of the isolates (>98%) with Pdd from other sources. Moreover, intramuscular injection at 1 × 108 CFU/ml and 1 × 103 CFU/ml produced pathological signs similar to those in naturally infected shrimp after 24 hpi and 10dpi respectively.