Isabel M. Cerezo’s research while affiliated with University of Malaga and other places

Simple Summary The use of functional feeds in aquaculture is currently increasing. In this study, we investigated the impact of a diet containing Chlorella fusca and ethanol-inactivated Vibrio proteolyticus in Chelon labrosus. After 90 days of feeding, we assessed how this diet affected the fish’s gut microbiota and gene expression related to metabolism, stress, and the immune system. We also tested the immune response after submitting fish to challenges with Aeromonas hydrophila and polyinosinic–polycytidylic acid (poly I:C). Results showed that the combined dietary administration influenced the microbial community in the fish’s intestines, but it did not change the way these microorganisms functioned. In terms of gene expression, we observed significant variations in several genes in different fish organs from fish fed the combination of microalgae and probiotics. Notably, the combined diet seemed to enhance the fish’s ability to regulate stress and immune-related genes, suggesting that it could improve their resistance to stress and infections. Overall, the present study sheds light on how this diet affects both the gut microbiota and gene expression in C. labrosus, potentially benefiting their health and immune response. Abstract The use of functional feeds in aquaculture is currently increasing. This study aimed to assess the combined impact of dietary green microalgae Chlorella fusca and ethanol-inactivated Vibrio proteolyticus DCF12.2 (CVP diet) on thick-lipped grey mullet (Chelon labrosus) juvenile fish. The effects on intestinal microbiota and the transcription of genes related to metabolism, stress, and the immune system were investigated after 90 days of feeding. Additionally, the fish were challenged with Aeromonas hydrophila and polyinosinic–polycytidylic acid (poly I:C) to evaluate the immune response. Microbiota analysis revealed no significant differences in alpha and beta diversity between the anterior and posterior intestinal sections of fish fed the control (CT) and CVP diets. The dominant genera varied between the groups; Pseudomonas and Brevinema were most abundant in the CVP group, whereas Brevinema, Cetobacterium, and Pseudomonas were predominant in the CT group. However, microbial functionality remained unaltered. Gene expression analysis indicated notable changes in hif3α, mhcII, abcb1, mx, and tnfα genes in different fish organs on the CVP diet. In the head kidney, gene expression variations were observed following challenges with A. hydrophila or poly I:C, with higher peak values seen in fish injected with poly I:C. Moreover, c3 mRNA levels were significantly up-regulated in the CVP group 72 h post-A. hydrophila challenge. To conclude, incorporating C. fusca with V. proteolyticus in C. labrosus diet affected the microbial species composition in the intestine while preserving its functionality. In terms of gene expression, the combined diet effectively regulated the transcription of stress and immune-related genes, suggesting potential enhancement of fish resistance against stress and infections.

This work aimed to assess the suitability of a microalgal blend as a dietary ingredient for feeding juveniles of marine carnivorous and herbivorous teleost, as is the case of Sparus aurata and Mugil cephalus, respectively, and to isolate microorganisms from different media and characterize them on the base of their enzymatic activities and their antagonism against important fish pathogens. Thirty juveniles of each species (70 ± 3.2 g S. aurata mean weight and 47 ± 2.8 g M. cephalus mean weight) were distributed in four tanks (15 individuals each) corresponding to four independent dietary treatments (control and microalgae diets designed for each species). Fish were fed their corresponding diets ad libitum for 108 days. At the end of the trial, fish were weighed, and plasma, liver, perivisceral fat, and the entire intestines were obtained for the evaluation of growth performance and metabolic assessment. Furthermore, 117 bacterial strains were isolated in different culture media from the gastrointestinal tract of S. aurata fed the microalgae blend and further characterized for their potential use as probiotics in aquaculture. S. aurata fed the microalgae-supplemented diet (25% dietary inclusion) showed a significant increase in weight gain, specific growth rate, feed efficiency, hepatosomatic, and intestine length indices. However, growth performance and somatic indices in M. cephalus were not affected by the experimental diets. Plasma samples from S. aurata fed the microalgal diet revealed higher levels of glucose and triglycerides and a decrease in cortisol levels. No significant differences were found in any biochemical parameters among the experimental diets in M. cephalus. In conclusion, both species demonstrated a favorable adaptation to the nutritional formulation employed in this study, and bacterial strains UMA-169 and UMA-216 (both identified as Bacillus pumilus) could be considered for use in aquaculture as they might benefit host health by improving digestion and absorption of different energy sources and by minimizing the colonization of pathogenic species.

Background: The development of a sustainable business model with social acceptance, makes necessary to develop new strategies to guarantee the growth, health, and well-being of farmed animals. Debaryomyces hansenii is a yeast species that can be used as a probiotic in aquaculture due to its capacity to i) promote cell proliferation and differentiation, ii) have immunostimulatory effects, iii) modulate gut microbiota, and/or iv) enhance the digestive function. To provide inside into the effects of D. hansenii on juveniles of gilthead seabream (Sparus aurata) condition, we integrated the evaluation of the main key performance indicators coupled with the integrative analysis of the intestine condition, through histological and microbiota state, and its transcriptomic profiling. Results: After 70 days of a nutritional trial in which a diet with low levels of fishmeal (7%) was supplemented with 1.1% of D. hansenii (17.2 × 105 CFU), an increase of ca. 12% in somatic growth was observed together with an improvement in feed conversion in fish fed a yeast-supplemented diet. In terms of intestinal condition, this probiotic modulated gut microbiota without affecting the intestine cell organization, whereas an increase in the staining intensity of mucins rich in carboxylated and weakly sulphated glycoconjugates coupled with changes in the affinity for certain lectins were noted in goblet cells. Changes in microbiota were characterized by the reduction in abundance of several groups of Proteobacteria, especially those characterized as opportunistic groups. The microarrays-based transcriptomic analysis found 232 differential expressed genes in the anterior-mid intestine of S. aurata, that were mostly related to metabolic, antioxidant, immune, and symbiotic processes. Conclusions: Dietary administration of D. hansenii enhanced somatic growth and improved feed efficiency parameters, results that were coupled to an improvement of intestinal condition as histochemical and transcriptomic tools indicated. This probiotic yeast stimulated host-microbiota interactions without altering the intestinal cell organization nor generating dysbiosis, which demonstrated its safety as a feed additive. At the transcriptomic level, D. hansenii promoted metabolic pathways, mainly protein-related, sphingolipid, and thymidylate pathways, in addition to enhance antioxidant-related intestinal mechanisms, and to regulate sentinel immune processes, potentiating the defensive capacity meanwhile maintaining the homeostatic status of the intestine.

Simple Summary This study evaluated using NGS techniques and bioinformatic analysis the influence on the microbiota and transcriptomic response of skin mechanically wounded gilthead seabream (S. aurata) specimens fed with a probiotic SpPdp11 supplemented diet. Four group of fish were established: non-wounded and wounded fed control diet, and non-wounded and wounded fed a probiotic enriched diet. The wounded group that received the probiotic diet showed a decrease in the abundance of taxa related to bacterial biofilm formation, and transcriptomic results suggested that specimens of this same group had a group of genes up-regulated and down-regulated related differently to those expressed in control group (non-wounded). Then, five genera that presented significant differences between these groups showed positive correlations with genes related to cell migration and negative correlations with inflammation and cell proliferation. These results are promising, and they open new perspectives and possibilities in the use of probiotic SpPdp11 to improve the skin after an injury, which can happen frequently in farmed specimens. Abstract Skin lesions are a frequent fact associated with intensive conditions affecting farmed fish. Knowing that the use of probiotics can improve fish skin health, SpPdp11 dietary administration has demonstrated beneficial effects for farmed fish, so its potential on the skin needs to be studied more deeply. The wounded specimens that received the diet with SpPdp11 showed a decrease in the abundance of Enterobacteriaceae, Photobacterium and Achromobacter related to bacterial biofilm formation, as well as the overexpression of genes involved in signaling mechanisms (itpr3), cell migration and differentiation (panxa, ttbk1a, smpd3, vamp5); and repression of genes related to cell proliferation (vstm4a, areg), consistent with a more efficient skin healing processes than that observed in the wounded control group. In addition, among the groups of damaged skin with different diets, Achromobacter, f_Ruminococcaceae, p_Bacteroidetes, Fluviicola and Flavobacterium genera with significant differences showed positive correlations with genes related to cell migration and negative correlations with inflammation and cell proliferation and may be the target of future studies.

Streptococcus agalactiae is an important bacterial pathogen in intensive Nile tilapia production, causing high mortality rates and great economic losses. This work aimed to evaluate the Nile tilapia vaccination against S. agalactiae and fed with ration containing probiotic AQUA PHOTO® composed of Bacillus subtilis and Lactobacillus plantarum, on the immune response action and gut microbiota. The experimental design was completely randomized with five treatments (CON = control; ADJ = adjuvant; PRO = probiotic; VAC = vaccine; PRO + VAC = probiotic + vaccine) and five replicates. The vaccine (bacterin + adjuvant) was injected after 21 days (21d) of probiotic feeding and the vaccine was booster 14 days post-vaccination (35d). After 14 days of the booster (49d), the fish were challenged with S. agalactiae and observed for more than 14 days, completing 63 days. The immunized group showed a better survival rate (CON 40%; ADJ 57%; PRO 67%; VAC 87%; PRO + VAC 97%). The treatments VAC and PRO + VAC, after booster produced higher levels of IgM antibodies compared with the control from the same time. The combination of probiotic and vaccination provided better protection against S. agalactiae infection, directly affecting the gut microbiological profile. These results indicated the contribution of probiotic to the adaptive immune response through the modulation of the intestinal microbiota, improving the effect of the vaccination. In conclusion, AQUA PHOTO®, composed of B. subtilis and L. plantarum, orally administered to Nile tilapia vaccinated against and challenged with S. agalactiae increases protection from infection and modifies the intestinal microbiota profile of the host, promoting the microbiota balance and improving adaptive immune response.

This study aimed to characterize the intestinal microbiota of wild thick-lipped grey mullets (Chelon labrosus) and explore its potential functionality on the host. Intestinal contents of anterior and posterior sections from wild fish were collected and DNA was extracted. Subsequently, the V3–V4 regions of 16S rRNA were sequenced using the Illumina technology and results were analyzed by bioinformatics pipeline. The functional profile of the microbial community was analyzed using PICRUSt software. Shannon and Simpson diversity indices were significantly higher in the posterior section of wild specimens. The overall taxonomic composition suggests a certain homogeneity in the anterior section of the intestine and heterogeneity in the posterior section. Due to this, no statistical differences were detected at any level among both intestinal sections. Predicted functions of intestinal microbiota showed the most abundant were those related to amino acid metabolism, carbohydrate metabolism, energy metabolism, membrane transport, and cell replication and repair. Furthermore, the analysis revealed microbial functional genes related to the elimination of environmental toxins.

The success of using probiotics is conditioned not only by the species of microorganism, but also by the dose administered and the method of administration. In aquaculture, adding probiotics to water makes them easier to administer, but the dilution effect may reduce the number of microorganisms that animals ingest. Probiotics may also be used to improve the chemical and microbiological characteristics of water. When administered in food, the microorganisms may be inactivated as they pass through the digestive tract. To avoid their inactivation, probiotics may be encapsulated in several materials. The durationAdministration, frequency, duration of treatment with probiotics as well as whether they are administered in one dose or in several pulses can also determine their effectiveness. Likewise, some strategies involve the use of several probiotics at the same time, having a synergistic effectSynergistic effect on the host. Finally, the effectiveness of probiotics should be evaluated if they are administered to feed as inactivated cells or if subcellular componentsSubcellular component of them are used.

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.

Fish stress is a major concern in the aquaculture industry. Many stressors coming from routine practices can predispose fish to compromised growth, immunity and overall health. This study focuses on the characterization of the skin microbiota using next generation sequencing (NGS) platform by targeting a genomic marker 16S and to determine growth performance and immune status of gilthead seabream (Sparus aurata) during an episode of chronic stress. Two groups were established: control group and chronically stressed group. Stressed fish were subjected to 1 min air exposure twice a week for 4 weeks. Results showed that stress negatively affected fish growth performance. Cellular and humoral systemic immunity remained unaffected while local immunity in skin was positively stimulated (total IgM and peroxidase). Skin mucus microbial composition showed significant differences especially after 14 days. Stressed fish showed a decrease in the abundance of the genera Acinetobacter, NS3a_marine_group and Pseudomonas, while Pseudoalteromonas and Marinagarivorans increased significantly. In conclusion, air exposure stress was associated with alterations in skin mucosal immunity and microbial composition that may have been beneficial to the host favoring adaptation to stress.