Aeromonas salmonicida is an important pathogen that causes furunculosis in trout and salmon with high morbidity and mortality, resulting in significant economic losses in aquaculture. Overuse of antibiotics has led to the continuous emergence of drug-resistant strains. Hence, there is an urgent need to find an alternative environmentally friendly antimicrobial agent. In this study, we isolated a virulent phage of A. salmonicida, named ASG01, which belongs to the Myoviridae family and maintains lytic activity at a pH value range from 4 to 12 and in the temperature range from 30 °C to 60 °C. The whole genomic sequence of ASG01 showed 82% similarity to Aeromonas phage pAh6-C. The cell wall hydrolase (Cwh)-encoding gene from the genome of ASG01 was predicted and heterologously expressed. Notably, in the absence of additional phage genes, endogenous expression of Cwh could lyse E. coli cells and greatly inhibit the growth of tested fish pathogenic bacteria. The lytic activity of Cwh was eliminated when the predicted active site was mutated. These results indicate that Cwh of ASG01 possessed excellent lytic activity and a wide antibacterial spectrum, suggesting its potential as an effective enzybiotic.

Currently, it is widely accepted that the type III secretion system (T3SS) serves as the transport platform for bacterial virulence factors, while flagella act as propulsion motors. However, there remains a noticeable dearth of comparative studies elucidating the functional disparities between these two mechanisms. Entomopathogenic nematode symbiotic bacteria (ENS), including Xenorhabdus and Photorhabdus, are Gram-negative bacteria transported into insect hosts by Steinernema or Heterorhabdus. Flagella are conserved in ENS, but the T3SS is only encoded in Photorhabdus. There are few reports on the function of flagella and the T3SS in ENS, and it is not known what role they play in the infection of ENS. Here, we clarified the function of the T3SS and flagella in ENS infection based on flagellar inactivation in X. stockiae (flhDC deletion), T3SS inactivation in P. luminescens (sctV deletion), and the heterologous synthesis of the T3SS of P. luminescens in X. stockiae. Consistent with the previous results, the swarming movement of the ENS and the formation of biofilms are dominated by the flagella. Both the T3SS and flagella facilitate ENS invasion and colonization within host cells, with minimal impact on secondary metabolite formation and secretion. Unexpectedly, a proteomic analysis reveals a negative feedback loop between the flagella/T3SS assembly and the type VI secretion system (T6SS). RT-PCR testing demonstrates the T3SS’s inhibition of flagellar assembly, while flagellin expression promotes T3SS assembly. Furthermore, T3SS expression stimulates ribosome-associated protein expression.

Bacteriophages are increasingly considered viable alternatives to antibiotics for controlling aquaculture pathogens, yet the structural basis of siphophage infection in aquatic bacteria remains largely unexplored. Here, we present the near-atomic-resolution cryo-electron microscopy structure of Avs-1, a siphophage infecting the freshwater aquaculture pathogen Aeromonas veronii. The capsid reconstruction at 2.98 Å and spike structure at 3.58 Å reveal a virion architecture broadly conserved among lambda-like siphophages, including an HK97-fold capsid, a canonical portal–neck complex, and a tail tube composed of approximately 26 stacked hexameric rings. However, the distal tail tip exhibits pronounced specialization, assembling into a C3-symmetric tripod-like core formed by the distal tail protein, hub protein, insertion protein, and the C-terminus of the tape measure protein. The trimeric spike protein gp43 extends from this platform and adopts a β-helical scaffold with a structurally divergent distal domain (D4), highlighting the structural diversity of phage tail-tip architectures. Together, these results reveal how a largely conserved virion framework is combined with a diversified distal tail apparatus, expanding the structural repertoire of characterized siphophages and providing a framework for investigating the molecular basis of phage–host interactions.