In the present study, Larimichthys crocea and Pseudomonas plecoglossicida were selected as a host-pathogen interaction model for teleosts and prokaryotic pathogens. Five shRNAs were designed and synthesized to silence the fliA gene, all of which resulted in pronounced reductions in fliA mRNA; the mutant strain with the best silencing efficiency of 92.16% was chosen for subsequent analysis. A significant decrease in motility, intracellular survival and escape was observed for the fliA-RNAi strain of P. plecoglossicida, whereby silencing of the fliA gene led to a 30% decrease in mortality and a four-day delay in the onset of infection in L. crocea. Moreover, silencing of P. plecoglossicida fliA resulted in a significant change in both the pathogen and host transcriptome in the spleens of infected L. crocea. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of pathogen transcriptome data showed that silencing fliA resulted in downregulation of 18 flagellum-related genes; KEGG analysis of host transcriptome data revealed that infection with the fliA-RNAi strain caused upregulation of 47 and downregulation of 106 immune-related genes. These pathogen-host interactions might facilitate clearance of P. plecoglossicida by L. crocea, with a significant decrease in fliA-RNAi P. plecoglossicida strain virulence in L. crocea.

This study aimed to investigate the effect of reduced expression of the luxR gene on the virulence of Pseudomonas plecoglossicida and the immune response of Epinephelus coioides. To achieve this, RNA interference (RNAi) was used to silence the luxR gene, and the pathogenicity of wild-type and luxR-RNAi strains of P. plecoglossicida, as well as the immune response of Epinephelus coioides to the infection of these two strains, were compared. The mutant strain with the highest silencing efficiency of 70.1% was selected for subsequent analysis. Silencing the luxR gene in the mutant strain resulted in a significant 30% reduction in mortality rates in artificially infected Epinephelus coioides compared to the wild-type strain. Transcriptome analysis revealed that the host transcriptome, particularly in the spleens of infected Epinephelus coioides, was markedly altered by the silencing of the luxR gene in the mutant strain. Tilapia infected with the luxR-RNAi strain exhibited altered immune defenses, with changes in gene expression primarily in the NOD-like receptor (NLR) signaling pathway. These results suggest that the luxR gene plays a crucial role in the host’s resistance to pathogen invasion, and reducing its expression could decrease quorum sensing (QS) signals while increasing the expression of the IL-1β gene in the host’s NLR pathway. This effect may lead to a pro-inflammatory response that enhances the immune response to infection. Further investigation of these mechanisms may lead to innovative approaches to treating bacterial infections.

Vibrio harveyi is a normal flora in natural marine habitats and a significant opportunistic pathogen in marine animals. This bacterium can cause a series of lesions after infecting marine animals, in which muscle necrosis and ulcers are the most common symptoms. This study explored the adaptation mechanisms of V. harveyi from the seawater environment to host fish muscle environment. The comprehensive transcriptome analysis revealed dramatic changes in the transcriptome of V. harveyi during its adaptation to the host fish muscle environment. Based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, flagellar assembly, oxidative phosphorylation, bacterial chemotaxis, and two-component systems play crucial roles in V. harveyi’s adaptation to host fish muscle. A comparison of biological phenotypes revealed that V. harveyi displayed a significant increase in flagellar length, swimming, twitching, chemotaxis, adhesion, and biofilm formation after induction by host fish muscle, and its dominant amino acids, especially bacterial chemotaxis induced by host muscle, Ala and Arg. It could be speculated that the enhancement of bacterial chemotaxis induced by amino acids plays a key role in the adaptation of V. harveyi from seawater to the muscle of the host fish.

Pseudomonas plecoglossicida is the pathogen of visceral white spot disease in marine fish, which usually occurs at 16–19 °C and has resulted in heavy economic losses. Our previous RNA sequencing revealed that the expression of the fliL gene in P. plecoglossicida was significantly up-regulated during infection of the host. In order to study the influence of the fliL gene on the virulence of P. plecoglossicida, the fliL gene of the NZBD9 strain was knocked out by the homologous recombination method, the fliL gene-deleted strain (ΔfliL strain) constructed, and complemented the fliL gene to the ΔfliL strain to obtain the C-ΔfliL strain. The growth curves of the NZBD9 strain, ΔfliL strain, and C-ΔfliL strain did not show significant differences. Compared with the NZBD9 strain, the motility, adhesion, and biofilm formation ability were tendered in the ΔfliL strain (p < 0.05); the complement of the fliL gene enhanced these abilities to the level of the NZBD9 strain. The results of artificial infection experiments showed that the LD₅₀ of NZBD9 strain, ΔfliL strain, and C-ΔfliL strain in hybrid grouper (Epinephelus fuscoguttatus ♀ × E. lanceolatus ♂) were 5.0 × 10³ CFU/fish, 6.3 × 10⁴ CFU/fish, and 1.3 × 10³ CFU/fish, respectively. RNA sequencing was performed on wild-type strains and ΔfliL strains. A total of 126 differentially expressed genes (DEGs) were screened (p < 0.05), of which 114 were downregulated and 12 were upcontrolled, among which several genes related to the six-type secretion system and transport activity were significantly downregulated. The DEGs were aligned to the GO and KEGG databases and enriched to 44 GO pathways and 39 KEGG pathways, respectively. The active pathways of ABC transporters were significantly enriched in both databases. These results indicate that the fliL gene is related to the movement, biofilm formation, and adhesion ability of P. plecoglossicida, and may reduce virulence by affecting substance transport and bacterial secretion.

Visceral white spot disease caused by Pseudomonas plecoglossicida severely threatens marine aquaculture, highlighting the need for effective control strategies. To clarify the role of a novel small RNA, sRNA0024, in bacterial pathogenicity, we constructed an sRNA0024 deletion mutant (ΔsRNA0024) and compared its phenotype and virulence with those of the wild-type strain NZBD9. In vitro assays showed that deletion of sRNA0024 did not affect bacterial growth but significantly reduced biofilm formation and adhesion. In vivo infection experiments in orange-spotted grouper (Epinephelus coioides) demonstrated that the ΔsRNA0024 mutant had a 3.8-fold higher 50% lethal dose (LD₅₀), improved host survival, and milder splenic lesions than the wild type. Histopathology and host transcriptome analyses revealed weakened activation of complement–coagulation cascades, neutrophil extracellular traps, leukocyte migration, and inflammatory signaling pathways, indicating a lower-intensity immune response. Bacterial transcriptomics showed that deletion of sRNA0024 was associated with reduced luxR expression and attenuated quorum-sensing–associated virulence traits, supporting a possible role for this small RNA in modulating luxR expression and QS-related host immunopathology. These findings identify sRNA0024 as an important contributor to the virulence of P. plecoglossicida and highlight the sRNA0024–luxR module as a potential antivirulence target for controlling visceral white spot disease in aquaculture.