Edwardsiella piscicidais a severe fish pathogen. Haem utilization systems play an important role in bacterial adversity adaptation and pathogenicity. In this study, a speculative haem utilization protein, HutZ_Ep, was characterized inE. piscicida.hutZ_Epis encoded with two other genes,hutWandhutX, in an operon that is similar to the haem utilization operonhutWXZidentified inV. cholerae. However, protein activity analysis showed that HutZ_Epis probably not related to hemin utilization. To explore the biological role of HutZ_Ep, a markerlesshutZ_Epin-frame mutant strain, TX01ΔhutZ, was constructed. Deletion ofhutZ_Epdid not significantly affect bacterial growth in normal medium, in iron-deficient conditions, or in the presence of haem but significantly retarded bacterial biofilm growth. The expression of known genes related to biofilm growth was not affected byhutZ_Epdeletion, which indicated that HutZ_Epwas probably a novel factor promoting biofilm formation inE. piscicida. Compared to the wild-type TX01, TX01ΔhutZexhibited markedly compromised tolerance to acid stress and host serum stress. Pathogenicity analysis showed that inactivation ofhutZ_Epsignificantly impaired the ability ofE. piscicidato invade and reproduce in host cells and to infect host tissue. In contrast to TX01, TX01ΔhutZwas defective in blocking host macrophage activation. The expression ofhutZ_Epwas directly regulated by the ferric uptake regulator Fur. This study is the first functional characterization of HutZ in a fish pathogen, and these findings suggested that HutZ_Epis essential forE. piscicidabiofilm formation and contributes to host infection.

Introduction: Vibriosis causes enormous economic losses of marine fish. The present study investigated the intestinal microbial response to acute infection of half-smooth tongue sole with different-dose Vibrio alginolyticus within 72 h by metagenomic sequencing.Methods: The inoculation amount of V. alginolyticus for the control, low-dose, moderate-dose, and high-dose groups were 0, 8.5 × 101, 8.5 × 104, and 8.5 × 107 cells/g respectively, the infected fish were farmed in an automatic seawater circulation system under a relatively stable temperature, dissolved oxygen and photoperiod, and 3 ~ 6 intestinal samples per group with high-quality DNA assay were used for metagenomics analysis.Results: The acute infections with V. alginolyticus at high, medium, and low doses caused the change of different-type leukocytes at 24 h, whereas the joint action of monocytes and neutrophils to cope with the pathogen infection only occurred in the high-dose group at 72 h. The metagenomic results suggest that a high-dose V. alginolyticus infection can significantly alter the intestinal microbiota, decrease the microbial α-diversity, and increase the bacteria from Vibrio and Shewanella, including various potential pathogens at 24 h. High-abundance species of potential pathogens such as V. harveyii, V. parahaemolyticus, V. cholerae, V. vulnificus, and V. scophthalmi exhibited significant positive correlations with V. alginolyticus. The function analysis revealed that the high-dose inflection group could increase the genes closely related to pathogen infection, involved in cell motility, cell wall/ membrane/envelope biogenesis, material transport and metabolism, and the pathways of quorum sensing, biofilm formation, flagellar assembly, bacterial chemotaxis, virulence factors and antibiotic resistances mainly from Vibrios within 72 h.Discussion: It indicates that the half-smooth tongue sole is highly likely to be a secondary infection with intestinal potential pathogens, especially species from Vibrio and that the disease could become even more complicated because of the accumulation and transfer of antibiotic-resistance genes in intestinal bacteria during the process of V. alginolyticus intensified infection.

A novel nidovirus, CSBV Bces-Po19, was isolated from the marine fish, Japanese flounder (Paralichthys olivaceus). The viral genome was 26,597 nucleotides long and shared 98.62% nucleotide identity with CSBV WHQSR4345. PacBio Sequel and Illumina sequencing were used to perform full-length transcriptome sequencing on CSBV Bces-Po19-sensitive (S) and -resistant (R) Japanese flounder. The results of negative staining revealed bacilliform and spherical virions. There were in total 1444 different genes between CSBV Bces-Po19 S and R groups, with 935 being up-regulated and 513 being down-regulated. Metabolism-, immune-, and RNA-related pathways were significantly enriched. Furthermore, CSBV Bces-Po19 infection induced alternative splicing (AS) events in Japanese flounder; the S group had a higher numbers of AS events (12,352) than the R group (11,452). The number of long non-coding RNA (lncRNA) in the S group, on the other hand, was significantly lower than in the R group. In addition to providing valuable information that sheds more light on CSBV Bces-Po19 infection, these research findings provide further clues for CSBV Bces-Po19 prevention and treatment.

Tributyrin (TB), as a novel feed additive, holds broad market prospects and is crucial for promoting fish growth and maintaining intestinal health. We first identified the fatty acid metabolism-related gene cpt1b in the intestines of mandarin fish (Siniperca chuatsi) from the TB-supplemented group. A total of 600 mandarin fish (200.0 ± 5.0 g) were evenly allocated into three groups. The control group (C) received only the standard extruded feed, while the experimental groups were supplemented with tributyrin (TB) at concentrations of 500 mg/kg (T1 group) and 1000 mg/kg (T2 group), respectively. Cloning yielded a 2364 bp open reading frame (ORF) encoding 787 amino acids, with the gene possessing two conserved transmembrane domains. Phylogenetic analysis further indicated a close phylogenetic relationship between largemouth blackbass (Micropterus salmoides) and mandarin fish. Tissue distribution and intestinal enzyme activity analyses revealed that supplementation with varying concentrations of TB upregulates cpt1b gene expression in different tissues, while modulating intestinal digestive enzyme and antioxidant enzyme activities. Our findings suggest a potential mechanism involving enhanced intestinal enzyme activity, reduced fat accumulation, increased expression of lipid oxidation-related genes, and accelerated TB degradation in the intestine.