Solea senegalensis Bacterial Intestinal Microbiota Is Affected by Low Dietary Inclusion of Ulva ohnoi.

The inclusion of macroalgae in the diets of farmed fish offers the opportunity for an added-value dietary ingredient to the nutraceutical feed. The composition of algae varies greatly among species. Several Ulva species have been considered in aquafeed formulations for different farmed fish, and Ulva ohnoi is being applied recently. However, the effects of seaweed dietary inclusion on the host must be evaluated. Considering the important role of the host intestinal microbiota, the potential effects of U. ohnoi dietary inclusion need to be studied. In this study, the characterization of the intestinal microbiome of Solea senegalensis, a flatfish with high potential for aquaculture in South Europe, receiving U. ohnoi (5%)-supplemented diet for 90 days has been carried out. In addition, the functional profiles of bacterial communities have been determined by using PICRUSt, a computational approach to predict the functional composition of a metagenome by using marker gene data and a database of reference genomes. The results show that long-term dietary administration of U. ohnoi (5%)-supplemented feed modulates S. senegalensis intestinal microbiota, especially in the posterior intestinal section. Increased relative abundance of Vibrio jointly with decreased Stenotrophomonas genus has been detected in fish receiving Ulva diet compared to control-fed fish. The influence of the diet on the intestinal functionality of S. senegalensis has been studied for the first time. Changes in bacterial composition were accompanied by differences in predicted microbiota functionality. Increased abundance of predicted genes involved in xenobiotic biodegradation and metabolism were observed in the microbiota when U. ohnoi diet was used. On the contrary, predicted percentages of genes associated to penicillin and cephalosporin biosynthesis as well as beta-lactam resistance were reduced after feeding with Ulva diet.

Inclusion of dietary Ulva ohnoi 5% modulates Solea senegalensis immune response during Photobacterium damselae subsp. piscicida infection.

Macroalgae represent valuable sources of functional ingredients for fish diets, and the influence of supplemented aquafeeds on growth performance has been studied for some fish and seaweed species. In the present work, the potential immunomodulation exerted by U. ohnoi (5%) as dietary ingredient was investigated in Senegalese sole. After feeding with the experimental diets for 90 d, fish immune response before and after challenge with Photobacterium damselae subsp. piscicida (Phdp) was assessed. In absence of infection, systemic immune response was not modified by 5% U. ohnoi dietary inclusion for 90 d. Thus, no differences in liver and head kidney immune gene transcription or serum lysozyme, peroxidase, antiprotease and complement activities were observed based on the diet received by Senegalese sole specimens. Regarding mucosal immune parameters, no changes in gene transcription were detected in the skin and gills, whilst only tnf, cd4 and cd8 were significantly up-regulated in the intestine of fish fed with U. ohnoi, compared to the values obtained with control diet. On the contrary, when S. senegalensis specimens were challenged with Phdp, modulation of the immune response consisting in increased transcription of genes encoding complement (c1q4, c3, c9), lysozyme g (lysg), tumor necrosis factor alpha (tnfα) as well as those involved in the antioxidant response (gpx, sodmn) and iron metabolism (ferrm, hamp-1) was observed in the liver of fish fed with U. ohnoi. In parallel, decreased inflammatory cytokine and complement encoding gene transcription was displayed by the spleen of fish receiving the algal diet. Though mortality rates due to Phdp challenge were not affected by the diet received, lower pathogen loads were detected in the liver of soles receiving U. ohnoi diet. Further research to investigate the effects of higher inclusion levels of this seaweed in fish diets, feeding during short periods as wells as to assess the response against other pathogens needs to be carried out.

Modulation of Intestinal Microbiota in Solea senegalensis Fed Low Dietary Level of Ulva ohnoi.

Gastrointestinal (GI) microbiota has a relevant role in animal nutrition, modulation of the immune system and protection against pathogen invasion. Interest in algae as source of nutrients and functional ingredients for aquafeeds is increasing in order to substitute conventional feedstuffs by more sustainable resources. The diet is an important factor in the modulation of the microbiota composition, and functional ingredients have been proposed to shape the microbiota and contribute benefits to the host. However, fish microbiome research is still limited compared to other hosts. Solea senegalensis is a flat fish with high potential for aquaculture in South Europe. In this study, a characterization of the microbiome of S. senegalensis (GI) tract and the effects of feeding Ulva ohnoi supplemented diet has been carried out. Differences in the composition of the microbiota of anterior and posterior sections of S. senegalensis GI tract have been observed, Pseudomonas being more abundant in the anterior sections and Mycoplasmataceae the dominant taxa in the posterior GI tract sections. In addition, modulation of the GI microbiota of juvenile Senegalese sole fed for 45 days a diet containing low percentage of U. ohnoi has been observed in the present study. Microbiota of the anterior regions of the intestinal tract was mainly modulated, with higher abundance of Vibrio spp. in the GI tract of fish fed dietary U. ohnoi.

Transcription of IVIAT and Virulence Genes in Photobacterium damselae Subsp. piscicida Infecting Solea senegalensis.

Photobacterium damselae subsp. piscicida (Phdp) is responsible for disease outbreaks in marine aquaculture worldwide. Solea senegalensis, a valuable fish species for aquaculture in the south of Europe, is frequently affected by this pathogen. It is well established that bacteria respond to environmental signals and, in the case of pathogens, this ability may determine the outcome of their interaction with the host. Determination of gene expression under in vivo conditions constitutes a valuable tool in the assessment of microbial pathogenesis. Considering that different hosts may represent different environments for the pathogen, expression of Phdp virulence and in vivo induced antigen (IVIAT) genes during S. senegalensis infection has been determined in the present work. Increased transcription of genes encoding proteins involved in iron acquisition (Irp1, Irp2, HutB and HutD), oxidative stress defence (AhpC and Sod), adhesion (PDP_0080), toxins (AIP56) and metabolism (Impdh, Shmt and AlaRS) were detected in Phdp infecting S. senegalensis head kidney or liver. The highest increases corresponded to genes involved in survival under iron limiting conditions and oxidative stress, indicating their essential role during infection of sole. Results obtained give insight into Phdp virulence strategies and contribute to the identification of promising targets for the control of photobacteriosis.

Changes in intestinal morphology and microbiota caused by dietary administration of inulin and Bacillus subtilis in gilthead sea bream (Sparus aurata L.) specimens.

Changes produced in gilthead sea bream (Sparus aurata L.) intestinal morphology and microbiota caused by dietary administration of inulin and Bacillus subtilis have been studied. Gilthead sea bream specimens were fed diets containing 0 (control), inulin (10 g kg(-1)), B. subtilis (10(7) cfu g(-1)), or B. subtilis + inulin (10(7) cfu g(-1) + 10 g kg(-1)) for four weeks. Curiously, fish fed the experimental diets (inulin, B. subtilis, or B. subtilis + inulin) showed the same morphological alterations when studied by light and electron microscopy, while significant differences in the signs of intestinal damage were detected by the morphometric study. All of the observed alterations were present only in the gut mucosa, and intestinal morphometric study revealed no effect of inulin or B. subtilis on the intestinal absorptive area. Furthermore, experimental diets cause important alterations in the intestinal microbiota by significantly decreasing bacterial diversity, as demonstrated by the specific richness, Shannon, and range-weighted richness indices. The observed alterations demonstrate that fish fed experimental diets had different signs of gut oedema and inflammation that could compromise their body homeostasis, which is mainly maintained by the epithelial lining of the gastrointestinal tract. To our knowledge, this is the first in vivo study regarding the implications of the use of synbiotics (conjunction of probiotics and prebiotics) on fish gut morphology and microbiota.

Histological alterations and microbial ecology of the intestine in gilthead seabream (Sparus aurata L.) fed dietary probiotics and microalgae.

The effects on histology and microbial ecology in gilthead seabream (Sparus aurata) intestine caused by dietary probiotic and microalgae were studied. Fish were fed non-supplemented (C, control) or supplemented diets with Tetraselmis chuii, Phaeodactylum tricornutum and Bacillus subtilis single or combined (diets T, P, B, BT and BP) for 4 weeks. Curiously, fish fed the experimental diets showed similar morphological alterations when studied by light and electron microscopy and significant signs of intestinal damage were detected. No effect of microalgae or B. subtilis on the intestinal absorptive area was observed, whereas the number of goblet cells and IELs were significantly lower in fish fed the T, P, B and BT diets and T, BT and BP diets, respectively. Interestingly, only the diets containing B. subtilis resulted in a significant reduction of microvilli height. Alterations such as wide intercellular spaces and large vacuoles in enterocytes were observed in fish fed T, B, BT, BT and P in lesser degrees. These observations demonstrate that fish fed experimental diets presented different signs of oedema and inflammation that could compromise their body homeostasis. Moreover, the experimental diets cause important alterations in the intestinal microbiota by a significant decrease in bacterial diversity, as demonstrated by the fall in specific richness, Shannon and range-weighted richness indices. To our knowledge, this is the first in vivo study regarding the implications of the use of probiotics in combination with immunostimulants on fish intestinal morphology and microbiota. More morphofunctional studies are needed in order to correlate the nutritional and immune aspects of fish gut.