ABSTRACT
Vibrio parahaemolyticus causes acute hepatopancreatic necrosis disease (AHPND) in farmed shrimp. Due to its damage potential, which could be as high as a 100% mortality rate, bacteriophages have emerged as a promising natural control intervention other than antibiotics, yet multiple roadblocks need to be overcome. In this study, six bacteriophages isolated from seafood samples, seawater, and estuary water in Sinaloa, Mexico, demonstrated a narrow host range among Mexican AHPND-causing V. parahaemolyticus. All bacteriophages are composed of a double-stranded DNA genome with lengths ranging between 43,268 and 57,805 bp. All six phages exhibited latency periods of 10-30 min and burst sizes of 34-168 viral particles per infected cell. The optimal MOI for bacteriophage propagation was 0.01-1. No transfer RNA (tRNA), virulence, or resistance genes were found in either genome, and the life cycle of these phages was classified as virulent by the PhageAI platform. Phylogenetic and comparative genomics analyzes assigned phages M3, C2, M9, and M83 as new species not yet reported within the genus Maculvirus, Autographiviridae family. ALK and CHI phages were assigned as new members of a new genus not yet classified within the subfamily Queuovirinae. The findings highlight the potential of CHI, ALK, M3, C2, M9, and M83 as promising alternatives against AHPND-causing V. parahaemolyticus from Mexico.
ABSTRACT
Acute hepatopancreatic necrosis disease (AHPND) is a life-threatening disease to recently stocked shrimp. This disease is mainly caused by Vibrio parahaemolyticus and, to date, it has not been effectively controlled. Bacteriophages are a promising method to control bacterial diseases in aquaculture and multiple phages that infect Asian strains of V. parahaemolyticus have been described. However, few studies have characterized the bacteriophages that infect Latin American strains. Here, two lytic Vibrio phages (vB_VpaP_AL-1 and vB_VpaS_AL-2) were isolated from estuary water in Sinaloa, Mexico. The host ranges were tested using ten AHPND-causing strains isolated from Mexico and phage AL-1 was able to infect two strains while AL-2 infected four. One-step growth curve showed that AL-1 produced 85 PFU/cell and AL-2 produced 68 PFU/cell in 30 and 40 min, respectively. Both phages were able to tolerate temperatures ranging from 20 to 50 °C and pH values ranging from 4 to 10. Phages AL-1 and AL-2 have double-stranded DNA genomes of 42,854 bp and 58,457 bp, respectively. In total, 53 putative ORFs associated with the phage structure, packing, host lysis, DNA metabolism, and additional functions were predicted in the AL-1 genome, while 92 ORFs associated with the same functions as the AL-1 and 1 tRNA were predicted in the AL-2 genome. The lifecycle was classified as virulent for both phages. Morphology, phylogeny, and comparative genomic analyses assigned phage AL-1 as a new member of the genus Maculvirus in the Autographiviridae family, and phage AL-2 as a new member of the Siphoviridae family. These findings suggest that vB_VpaP_AL-1 and vB_VpaS_AL-2 are potential biocontrol agents against AHPND-causing V. parahaemolyticus from Mexico.
