Identification, characterization and complete genome analysis of a Vibrio anguillarum isolated from Sebastes schlegelii.

Vibrio anguillarum is an important fish pathogen in mariculture, which can infect fish with great economic losses. In this study, a Vibrio anguillarum isolated from Sebastes schlegelii was named VA1 and was identified and characterized from aspects of morphology, physiological and biochemical characteristics, 16SRNA, virulence genes, drug sensitivity, and extracellular enzyme activity. At the same time, The VA1 was investigated at the genomic level. The results showed that a Gram-negative was isolated from the diseased fish. The VA1 was characterized with uneven surface and visible flagella wrapped in a sheath and microbubble structures. The VA1 was identified as Vibrio anguillarum based on the 16S RNA sequence and physiological and biochemical characteristics. The VA1 carried most of the virulence genes (24/29) and was resistant to penicillin, oxacillin, ampicillin, cefradine, neomycin, pipemidic acid, ofloxacin, and norfloxacin. The pathogenicity of the isolated strain was confirmed by an experimental analysis, and its LD50 was 6.43 × 106 CFU/ml. The VA1 had the ability to secrete gelatinase, protease, and amylase, and it had α-hemolysis. The whole genome size of the VA1 was 4232328bp and the G + C content was 44.95 %, consisting of two circular chromosomes, Chromosome1 and Chromosome2, with no plasmid. There were 1006 predicted protein coding sequences (CDSs). A total of 526 genes were predicted as virulence-related genes which could be classified as type IV pili, flagella, hemolysin, siderophore, and type VI secretion system. Virulence genes and correlation data were supported with the histopathological examination of the affected organs and tissues. 194 genes were predicted as antibiotic resistance genes, including fluoroquinolone antibiotic, aminoglycoside antibiotic, and beta-lactam resistant genes, which agreed with the results of the above drug sensitivity, indicating VA1 to be a multidrug-resistant bacterium. This study provided a theoretical basis for a better understanding of pathogenicity and antibiotic resistance, which might contribute to the prevention of V. anguillarum in the future.

The effects of Bacillus subtilis on the immunity, mucosal tissue morphology, immune-related gene transcriptions, and intestinal microbiota in flounder (Paralichthys olivaceus) with two feeding methods: Continuous versus discontinuous feeding.

Probiotics as dietary additives can improve weight gain, feed efficiency, and disease resistance in cultured fish. In this research, we evaluated and compared the effects of Bacillus subtilis on immunity, mucosal tissue morphology, immune-related gene transcriptions, and intestinal microbiota in flounder (Paralichthys olivaceus) by a 30-day feeding experiment based on a continuous feeding schedule (E1) and a discontinuous feeding schedule (E2). As a result, the use of B. subtilis exerted the best positive effects on survival rate, enzyme activity, mucosal tissue morphology, immune-related gene transcriptions, and intestinal microbiota in flounders. Alkaline phosphatase (AKP), lysozyme (LZM), and superoxide dismutase (SOD) activities in the liver of E2 were higher than those of E1 (P < 0.05). Furthermore, the villi length in the intestinal tract and the fold length in the stomach of E2 were also higher than in E1 (P < 0.05). The il-1 expression levels in the spleen were significantly increased in E2 (P < 0.05) compared to E1. We performed 16 S rRNA sequencing analysis to find that Bacillus in E1 (1.06%) and E2 (1.01%) had higher relative abundances than in E0 (0.053%) at the end of the experiments, indicating that short-term application of B. subtilis with the continuous or discontinuous feeding method can allow both the adaptation of the ecosystem to the presence of probiotics by the establishment of new species in the gut microbiota and the ability these new probiotic species to perform corresponding functions. No significant differences in the ability of probiotic establishment were observed between E1 and E2. Our findings provided a unique perspective to explore the mechanism of immune enhancement with probiotics and to screen the optimal administration strategy in aquaculture application for probiotic use. Together, these results point to some level of enhancement in immune status by continuous and discontinuous feeding after a short-term feeding period, which could be used as a prophylactic strategy for flounder health management.

Effects of the water-soluble fraction of diesel oil on the sera biochemical indicators, histological changes, and immune responses of black rockfish Sebastes schlegelii.

In order to understand the physiological and immune responses of Sebastes schlegelii to the water-soluble fraction of diesel oil (WSD), S. schlegelii were used as the experimental objects to study the effect of WSD on the sera biochemical indicators, histological changes, and immune responses. Significant differences in sera biochemical indicators were observed in S. schlegelii after WSD exposure. The alkaline phosphatase (ALP), glucose (GLU), and globulin (GLB) were reduced by 3.51-fold, 3.12- fold, and 1.58-fold, respectively; however, K+ was increased by 3.55-fold. The results of HE staining showed that interstitial congestion was observed in the liver; the secondary lamellae deformity and hyperplasia, epithelial lifting, the primary lamellae hyperplasia, and aneurism were observed in the gill. Epidermis thickness increased, and epidermal hyperplasia in the skin was shown. The length of the secondary lamellae shortened significantly after WSD exposure. The results of AB-PAS staining showed that three different types of mucous cells were observed in the gill, and a significant increase in the number of all three types of mucous cells was observed after WSD exposure (P < 0.05). In addition, the results of the relative mRNA expressions in the liver of eleven immune-related genes showed that the relative expression levels of IL-1ß, IL-8, TNF receptor, BAFF, C1s, C1r, and MyD88 in the WSD group were substantially higher than those in the LPS group (P < 0.05), and the relative expression of caspase 10 was significantly lower than that in the LPS group (P < 0.05). At the same time, no significant differences were observed in the relative expression levels of IL-1, TNFα, and C1inh between the two groups (P > 0.05). This study was expected to provide essential data for health assessments of S. schlegelii and establish the foundation for the immune-related researches of S. schlegelii after WSD exposure.