Gut Microbiota Of Rainbow Trout Research Articles

Alteration of the Intestinal Bacterial Community in Rainbow Trout (Oncorhynchus mykiss): The Role of Animal, Plant, and Microbial Diets

Intestinal bacterial community in rainbow trout (Oncorhynchus mykiss) is increasingly recognized as important for aquaculture and fish health. This review summarizes the current knowledge on how various feed components, including animal- and plant-derived ingredients, as well as other feed additives, influence the gut microbiota of rainbow trout. Studies using 16S rRNA gene profiling and metagenomics demonstrate how dietary changes affect the composition, diversity, and functionality of bacterial communities. The findings suggest that there is no single optimal alternative ingredient for fish feeds, but emphasize that the ratio of natural components plays a more significant role in shaping the intestinal bacterial community of trout. This balance is crucial for maintaining diversity, functionality, and overall fish health, thereby promoting growth and disease resistance, while enhancing aquaculture efficiency. The use of popular animal protein derived from the larva of Hermetia illucens increases the amount of Pseudomonadota, but the chitin they contain can prevent the absorption of nutrients. Thus, the development of optimal feed formulas requires careful selection and balancing of various natural ingredients to sustain a healthy microbiota and overall fish well-being.

259 Genome-Wide Insights into Whole Gut Microbiota of Rainbow Trout,Oncorhynchus Mykiss Associated with Changes in Dietary Composition and Temperature Regimens

Abstract Gut microbiomes play a significant role in the growth, metabolism, and health of fish. However, we know very little about interactive effects of variations of dietary composition and temperature on rainbow trout gut microbiota. Exactly 288 rainbow trout weighing 45.6g ± 0.05 (average ± SD) were fed four isocaloric, isolipidic, and isonitrogenous diets comprised 40% crude protein and 20% crude lipid and formulated as 100 % animal-based protein (AP) and blend of 50 fish oil (FO)/50 camelina oil (CO), 100 % AP and100 % CO, 100 % plant-based protein (PP) and blend of 50FO/50CO or 100 % PP and 100 % CO in 14 or 18°C for 150 days. Gut content was analyzed using 16S rRNA gene and shotgun sequencing. The most abundant phyla identified regardless of diet were Tenericutes, Firmicutes, Proteobacteria, Spirochaetes, Bacteroidetes, and Actinobacteria while Aeromonadaceae and Enterobacteriaceae were dominant families in 18°C. Moreover, gut microbes were dominated by genes relating to amino acid, carbohydrate, fat, and energy metabolisms and influenced by temperature. The shared functional profiles for all the diets suggest that plant protein sources in combination with CO could be as good as the fish meal with 50/50 FO and CO in rainbow trout farming.

Performance, immune response, disease resistance, and gut microbiota of rainbow trout, Oncorhynchus mykiss (Walbaum, 1792) juveniles fed ground leonardite with a high humic substance content

In the present study, the effects of feeding ground leonardite (LE), which contained 68% of humic substances, to juvenile Oncorhynchus mykiss were examined on overall growth and production performance, feed nutrient digestibility, gut microbiota, health condition, antioxidant status, immune response, and disease resistance. During two consecutive experiments, fish were fed four experimental diets with different LE supplementary levels: 0, 0.5, 1, and 3% of feed mass (LE0, LE0.5, LE1, and LE3, respectively). Firstly, the fish were tested in a growing experiment (GE, n = 300 fish/group, body weight 15.4 ± 2.5 g) for 56 days. Thereafter, these fish (n = 75 fish/group, body weight 51.9 ± 8.7 g) were exposed to an intraperitoneal infection challenge experiment (IC) with Aeromonas salmonicida subsp. salmonicida for 28 days.In the GE, dietary LE did not affect the growth, production, or health performance of O. mykiss juveniles, although the fish more than tripled their initial body weight. The LE dose had an impact on the digestibility of feed nutrients (dry matter, gross energy, and ash), but not on the protein and lipid digestibility or solid waste outputs. The gut microbial communities were highly diverse across the experimental groups and remained distinct between the control and LE groups. The LE had a dose-dependent effect on the cell composition of the head kidney and peripheral blood leukocytes. The highest LE addition exerted the immune suppressive effect, and a decreased count of immune cells was observed in the LE3.The peritoneal cavity infection in the IC trial induced the rapid recruitment of myeloid cells in all groups shortly after the infection, except for the LE3, which had an apparent delayed response. The cumulative mortality and specific trout antibody levels against A. salmonicida did not significantly vary among tested groups, although their average initial level dropped by half at 3 days post-infection (dpi), and subsequently doubled at 28 dpi. The plasma biochemical parameters (TP, TAG, and CHOL) showed an overall decrease at 0–3 dpi, while increasing back to the initial levels at 3–6 dpi. Only LDH negatively correlated with LE inclusion, this can reflect the oxidative stress in the highest LE inclusion.Overall, our results support that there is a dose-dependent effect that should be optimised for ground LE dietary use in aquaculture.

Genome‐wide insights into whole gut microbiota of rainbow trout, Oncorhynchus mykiss, fed plant proteins and camelina oil at different temperature regimens

AbstractGut microbiota impacts fish metabolism, nutrient utilization, and health. We know little about how temperature and diet interact with rainbow trout gut microbes. A total of 288 fish (average body weight: 45.6 g) fed four iso‐caloric, ‐lipidic, and ‐nitrogenous diets comprised crude protein (40%) and lipid (20%) manufactured as 100% animal‐based protein (AP) and blend of 50 fish oil (FO)/50 camelina oil (CO), 100% AP and 100% CO, 100% plant‐based protein (PP) and blend of 50 FO/50 CO or 100% PP and 100% CO at 14 or 18°C water temperature. Gut content was analyzed using 16S rRNA gene and shotgun sequencing. Alpha‐diversity did not change significantly. Regardless of diet, Tenericutes, Firmicutes, Proteobacteria, Spirochaetes, Bacteroidetes, and Actinobacteria dominated. At family level, Aeromonadaceae and Enterobacteriaceae dominated at 18°C, while Mycoplasmataceae dominated at 14°C. Moreover, genes relating to amino acid, carbohydrate, fat, and energy metabolisms and fatty acid biosynthesis significantly increased at 18°C. Functional profiles did not vary significantly among diets, except for a higher methionine and cysteine metabolism in fish fed plant ingredient compared animal diet, suggesting trout fed plant‐derived protein mixed with CO could be as effective as those fed fish meals mixed with 50/50 FO and CO.

Potential of shrimp waste meal and insect exuviae as sustainable sources of chitin for fish feeds

Aquaculture is one of the world's fastest growing food-producing sectors, providing more than half of all fish consumed globally for human nutrition. However, to maintain such growth and meet the increasing demand for aquatic food, sustainable raw materials for fish feeds are needed. In this regard, insects represent one of the most promising alternatives to fish meal (FM) protein source for use in aquafeeds. In addition to protein, insects contain bioactive compounds, such as chitin, which is a natural polysaccharide abundantly present in the pupal exuviae of some insects. Studies have shown that dietary chitin or its derivate chitosan acts as a prebiotic thus modulating the gut microbial communities of fish. Accordingly, the present study aimed to evaluate the effect of two waste products rich in chitin, i.e., shrimp head meal (SHM), and insect (Hermetia illucens) pupal exuviae on the gut microbiota of rainbow trout (Oncorhynchus mykiss). Three isoproteic, isolipidic, and isoenergetic diets containing either FM, SHD, or a combination of FM and 1.6% of pupal exuviae meal (PEM) were tested through a 91-day feeding trial. At the end of the experiment, no differences in final mean body weight, specific growth rate, and feed conversion ratio values were observed between fish experimental groups. Mortality was <1% and it did not correlate with diet for the entire duration of the trial. However, a modulatory effect of dietary pupal exuviae on fish gut microbiota was detected. Indeed, gut bacterial species richness improved by including insect exuviae. In particular, Firmicutes and Actinobacteria phyla, mainly represented by Bacillus, Facklamia, Brevibacterium, and Corynebacterium genera, were enriched in trout receiving pupal exuviae. These genera are chitinolytic and short-chain fatty acids (SCFAs)-producing bacteria. SCFAs production was confirmed by gas chromatography analysis, which detected the highest amount of butyrate in feces of trout fed with pupal exuviae meal. Functional inference analysis of intestinal microbiota using PICRUST metagenome prediction tool, showed differences in response to diet. In particular, eleven pathways were significantly different between control fish (FM) and fish fed the PEM diet, whereas twenty functional traits were significantly different between the FM and SHM fish groups. Overall, our data confirmed that chitin from insect's pupal exuviae represents a promising functional ingredient, better than SHM, for positively modulating gut microbiota communities of rainbow trout.

Distinct microbial assemblages associated with genetic selection for high- and low- muscle yield in rainbow trout

BackgroundFish gut microbial assemblages play a crucial role in the growth rate, metabolism, and immunity of the host. We hypothesized that the gut microbiota of rainbow trout was correlated with breeding program based genetic selection for muscle yield. To test this hypothesis, fecal samples from 19 fish representing an F2 high-muscle genetic line (ARS-FY-H) and 20 fish representing an F1 low-muscle yield genetic line (ARS-FY-L) were chosen for microbiota profiling using the 16S rRNA gene. Significant differences in microbial assemblages between these two genetic lines might represent the effect of host genetic selection in structuring the gut microbiota of the host.ResultsTukey’s transformed inverse Simpson indices indicated that high muscle yield genetic line (ARS-FY-H) samples have higher microbial diversity compared to those of the low muscle yield genetic line (ARS-FY-L) (LMM, χ2(1) =14.11, p < 0.05). The fecal samples showed statistically distinct structure in microbial assemblages between the genetic lines (F1,36 = 4.7, p < 0.05, R2 = 11.9%). Functional profiling of bacterial operational taxonomic units predicted characteristic functional capabilities of the microbial communities in the high (ARS-FY-H) and low (ARS-FY-L) muscle yield genetic line samples.ConclusionThe significant differences of the microbial assemblages between high (ARS-FY-H) and low (ARS-FY-L) muscle yield genetic lines indicate a possible effect of genetic selection on the microbial diversity of the host. The functional composition of taxa demonstrates a correlation between bacteria and improving the muscle accretion in the host, probably, by producing various metabolites and enzymes that might aid in digestion. Further research is required to elucidate the mechanisms involved in shaping the microbial community through host genetic selection.

Effects of increasing dietary level of organic acids and nature-identical compounds on growth, intestinal cytokine gene expression and gut microbiota of rainbow trout (Oncorhynchus mykiss) reared at normal and high temperature

Organic acids (OA) and nature-identical compounds (NIC) such as monoterpenes and aldehydes are well-known growth and health promoters in terrestrial livestock while their application for fish production is recent and their mechanisms of action require further study. Hence, this study tested the increasing dietary level (D0, D250, D500, D1000; 0, 250, 500 and 1000 mg kg feed−1 respectively) of a microencapsulated blend containing citric and sorbic acid, thymol and vanillin over 82 days on rainbow trout to assess the effects on growth, feed utilization, intestine cytokine gene expression and gut microbiota (GM). Furthermore, the effects on intestinal cytokine gene expression and GM were also explored after one week at high water temperature (23 °C). OA and NIC improved specific growth rate (SGR) and feed conversion rate (FCR) during the second half (day 40–82) of the feeding trial, while at the end of the trial protein (PER) and lipid efficiency (LER) increased with increasing dietary level. GM diversity and composition and cytokine gene expression analysis showed no significant differences in fish fed with increasing doses of OA and NIC (82 days) demonstrating the absence of inflammatory activity in the intestinal mucosa. Although there were no statistical differences, GM structure showed a tendency in clustering D0 group separately from the other dietary groups and a trend towards reduction of Streptococcus spp. was observed in the D250 and D1000 groups. After exposure to high water temperature, lower GM diversity and increased gene expression of inflammatory intestinal cytokines were observed for both inclusions (D0 vs. D1000) compared to groups in standard condition. However, the gene up-regulation involved a limited number of cytokines showing the absence of a substantial inflammation process able to compromise the functional activity of the intestine. Despite further study should be conducted to fully clarify this mechanism, cytokines up-regulation seems to be concomitant to the reduction of the GM diversity and, particularly, to the reduction of specific lactic acid bacteria such as Leuconostoc. The application of the microencapsulate blend tested can be a useful strategy to improve growth and feed utilization in rainbow trout under normal temperature conditions. According to the results organic acids and nature-identical compounds did not revert the effects triggered by the increased temperature of water.

Analysis of the fecal microbiota of fast- and slow-growing rainbow trout (Oncorhynchus mykiss)

BackgroundDiverse microbial communities colonizing the intestine of fish contribute to their growth, digestion, nutrition, and immune function. We hypothesized that fecal samples representing the gut microbiota of rainbow trout could be associated with differential growth rates observed in fish breeding programs. If true, harnessing the functionality of this microbiota can improve the profitability of aquaculture. The first objective of this study was to test this hypothesis if gut microbiota is associated with fish growth rate (body weight). Four full-sibling families were stocked in the same tank and fed an identical diet. Two fast-growing and two slow-growing fish were selected from each family for 16S rRNA microbiota profiling.Microbiota diversity varies with different DNA extraction methods. The second objective of this study was to compare the effects of five commonly used DNA extraction methods on the microbiota profiling and to determine the most appropriate extraction method for this study. These methods were Promega-Maxwell, Phenol-chloroform, MO-BIO, Qiagen-Blood/Tissue, and Qiagen-Stool. Methods were compared according to DNA integrity, cost, feasibility and inter-sample variation based on non-metric multidimensional scaling ordination (nMDS) clusters.ResultsDifferences in DNA extraction methods resulted in significant variation in the identification of bacteria that compose the gut microbiota. Promega-Maxwell had the lowest inter-sample variation and was therefore used for the subsequent analyses. Beta diversity of the bacterial communities showed significant variation between breeding families but not between the fast- and slow-growing fish. However, an indicator analysis determined that cellulose, amylose degrading and amino acid fermenting bacteria (Clostridium, Leptotrichia, and Peptostreptococcus) are indicator taxa of the fast-growing fish. In contrary, pathogenic bacteria (Corynebacterium and Paeniclostridium) were identified as indicator taxa for the slow-growing fish.ConclusionDNA extraction methodology should be carefully considered for accurate profiling of the gut microbiota. Although the microbiota was not significantly different between the fast- and slow-growing fish groups, some bacterial taxa with functional implications were indicative of fish growth rate. Further studies are warranted to explore how bacteria are transmitted and potential usage of the indicator bacteria of fast-growing fish for development of probiotics that may improve fish health and growth.

The Effects of Dietary Insect Meal from Hermetia illucens Prepupae on Autochthonous Gut Microbiota of Rainbow Trout (Oncorhynchus mykiss).

Simple SummaryThe composition of fish-gut microbial communities has been demonstrated to adapt when the host is fed different ingredients. Fishmeal and soy are the conventional protein sources used in aquafeeds. However, these feed options are not sustainable anymore due to the progressive depletion of wild marine fish stocks and the considerable environmental cost of protein-rich terrestrial plant cultivation. In this perspective, insects could be a promising source of protein and may help aquaculture to cope with the increasing global demand for new protein sources, representing the idea of “waste into feed” bioconversion. In this perspective, we evaluated the effects of dietary insect meal from Hermetia illucens (Hi) larvae on autochthonous gut microbiota of rainbow trout (Oncorhynchus mykiss). Hi larvae were grown on leftover fruit and vegetables provided by a local wholesale market. Three diets, with increasing levels of insect meal inclusion (10%, 20%, and 30%) and a control diet without insect meal were tested in a 3-month fish feeding trial. The data showed that feeding insects influences the intestinal bacterial communities, thus improving fish gut health. In our opinion, these findings represent a precious tool for future research on salmonid’s microbial communities and their interaction with diet and the host.This study evaluated the effects of dietary insect meal from Hermetia illucens larvae on autochthonous gut microbiota of rainbow trout (Oncorhynchus mykiss). Three diets, with increasing levels of insect meal inclusion (10%, 20%, and 30%) and a control diet without insect meal were tested in a 12-week feeding trial. To analyze the resident intestinal microbial communities, the Illumina MiSeq platform for sequencing of 16S rRNA gene and QIIME pipeline were used. The number of reads taxonomically classified according to the Greengenes database was 1,514,155. Seventy-four Operational Taxonomic Units (OTUs) at 97% identity were identified. The core of adhered intestinal microbiota, i.e., OTUs present in at least 80% of mucosal samples and shared regardless of the diet, was constituted by three OTUs assigned to Propiobacterinae, Shewanella, and Mycoplasma genera, respectively. Fish fed the insect-based diets showed higher bacterial diversity with a reduction in Proteobacteria in comparison to fish fed the fishmeal diet. Insect-meal inclusion in the diet increased the gut abundance of Mycoplasma, which was attributed the ability to produce lactic and acetic acid as final products of its fermentation. We believe that the observed variations on the autochthonous intestinal microbiota composition of trout are principally due to the prebiotic properties of fermentable chitin.

Dietary live yeast and increased water temperature influence the gut microbiota of rainbow trout.

The objective was to determine the effects of dietary substitution of fishmeal (FM) with live yeast and increasing water temperature on the diversity and composition of gut microbiota in rainbow trout. Fish were fed either FM or yeast (Saccharomyces cerevisiae) and reared in water temperatures of either 11°C (cold) or 18°C (warm) for 6weeks. Luminal content and mucosa were collected from the distal gut and the load, diversity and species abundance of yeast and bacteria were analysed using agar plating, MALDI-TOF and rRNA gene amplicon sequencing. Yeast in the gut of fish fed FM were represented by S. cerevisiae, Rhodotorula spp. and Debaryomyces hansenii, while fish fed yeast contained 4-5 log higher CFU per g of yeast that were entirely represented by S. cerevisiae. For gut bacteria, sequencing of 16S rRNA gene amplicons using Illumina MiSeq showed lower bacterial diversity and abundance of lactic acid bacteria, especially Lactobacillus, in fish reared in warm rather than cold water. Fish fed yeast had similar bacterial diversity and lower abundance of Leuconostocaceae and Photobacterium compared with fish fed FM. Feeding live yeast mainly increased yeast load in the gut, while increased water temperature significantly altered the gut microbiota of rainbow trout in terms of bacterial diversity and abundance. Live yeast can replace 40% of FM without disrupting bacteria communities in the gut of rainbow trout, while increased water temperature due to seasonal fluctuations and/or climate change may result in a gut dysbiosis that may jeopardize the health of farmed fish.

Effects of dietary inclusion of the yeasts Saccharomyces cerevisiae and Wickerhamomyces anomalus on gut microbiota of rainbow trout

Rainbow trout (Oncorhynchus mykiss) were fed for 10weeks on a diet containing either 30% fish meal (FM) or with 20, 40 and 60% replacement of fish meal protein with Saccharomyces cerevisiae (SC) or a mixture of Wickerhamomyces anomalus and S. cerevisiae (WA). Luminal contents and mucosal tissue from the distal intestine were collected and analysed for yeast and bacterial loads by agar plating. Diversity and abundance were determined by sequencing of amplicons generated from the 26S rRNA (yeast) and 16S rRNA (bacteria) genes. In addition, the diets were analysed before and after feed extrusion to determine the viability and composition of the yeasts ingested by the fish. After extrusion, 9–10logcellsg−1 of yeast were still intact in the SC and WA diets, but culturable yeast showed log-reductions of 5–7CFUg−1. For yeasts isolated from the gut contents, 81–96% of colonies consisted of Debaryomyces hansenii, with few or no colonies of S. cerevisiae or W. anomalus despite their high inclusion rate in the diets. Characterisation of gut bacteria using Illumina MiSeq showed that 70 and 19% of sequences were classified to the phyla Firmicutes and Proteobacteria, specifically sequences identified as Leuconostocaceae, Lactobacillaceae and Photobacterium. Compared with the FM diet, the WA40 diet reduced bacterial diversity, whereas the WA60 diet increased the abundance of the pathogenic yeast Candida albicans and reduced lactic acid bacteria in the gut. Overall, 40 and 60% replacement of fish meal protein with a mixture of W. anomalus and S. cerevisiae significantly altered the gut microbiota of rainbow trout, while 20% replacement and diets with only S. cerevisiae had little or no effect.

Dietary probiotic supplementation modulated gut microbiota and improved growth of juvenile rainbow trout (Oncorhynchus mykiss)

In aquaculture, infectious diseases are the major cause of economic losses. Probiotic supplementation may change the microbiota of the digestive tract and modulate the immune defences and nutritional performance. This study was conducted to evaluate the dietary supplementation of multi-species (A: Bacillus sp., Pediococcus sp., Enterococcus sp., Lactobacillus sp.) and single-species probiotics (B: Pediococcus acidilactici) on growth performance and gut microbiota of rainbow trout (Oncorhynchus mykiss). A basal diet was supplemented with probiotic A or B, at two concentrations each (A1, A2, B1 and B2) or not supplemented (control treatment). Diets were distributed to 30 groups of 20 fish, 3 times a day. The gut microbiota was analysed at the end of the feeding trial (96days) with 16S rDNA denaturing gradient gel electrophoresis (16S-DGGE). Changes in gut microbial community were assessed by Shannon index (H´) and number of operational taxonomic units (OTUs). After 56days of feeding, weight gain was significantly improved in fish fed diet A1 when compared to the control group. Dietary probiotic supplementation changed the gut microbial composition. Number of OTUs (R) was higher in fish fed A1 (multi-species at lower concentration) than in control group, while H´ was higher in fish fed A1, B1 and B2.

PCR-TTGE Analysis of 16S rRNA from Rainbow Trout (Oncorhynchus mykiss) Gut Microbiota Reveals Host-Specific Communities of Active Bacteria

This study assessed the relative contributions of host genetics and diet in shaping the gut microbiota of rainbow trout. Full sibling fish from four unrelated families, each consisting of individuals derived from the mating of one male and one female belonging to a breeding program, were fed diets containing either vegetable proteins or vegetable oils for two months in comparison to a control diet consisting of only fish protein and fish oil. Two parallel approaches were applied on the same samples: transcriptionally active bacterial populations were examined based on RNA analysis and were compared with bacterial populations obtained from DNA analysis. Comparison of temporal temperature gradient gel electrophoresis (TTGE) profiles from DNA and RNA showed important differences, indicating that active bacterial populations were better described by RNA analysis. Results showed that some bacterial groups were significantly (P<0.05) associated with specific families, indicating that microbiota composition may be influenced by the host. In addition, the effect of diet on microbiota composition was dependent on the trout family.

Development of protection in rainbow trout ( Oncorhynchus mykiss, Walbaum) to Vibrio anguillarum following use of the probiotic Kocuria SM1

Probiotic Kocuria SM1, which was part of the allochthonous gut microbiota of rainbow trout, was administered to rainbow trout ( Oncorhynchus mykiss) fingerlings as dietary supplement at ∼10 8 cells g −1 for two weeks. The fish were challenged with Vibrio anguillarum and innate immunity determined over the following 5 weeks. The probiotic-fed fish survived (mortalities = 10–28%; P < 0.05) better than the controls (mortalities = 73–92%) for 5 weeks after stopping administration of Kocuria SM1, although the relative percent survival (RPS) declined steadily from 87 to 36% by the end of the experiment. A significant ( P < 0.05) effect on the immune parameters was observed in fish within 3 weeks after stopping administration of Kocuria SM1 in comparison with the controls, i.e. enhancement of the respiratory burst (OD 550nm) from 0.051 ± 0.014 to 0.067 ± 0.009, lysozyme 605 ± 185 units ml −1 to 872 ± 114 units ml −1, total protein 19.8 ± 2.0 mg ml −1 to 23.8 ± 0.8 mg ml −1 at 2 weeks, and leukocrit 2.9 ± 1.8% to 5.1 ± 1.1%, peroxidase (OD 550nm) 0.23 ± 0.02 to 0.37 ± 0.11 and bacterial killing activities (percentage of surviving bacteria = 82 ± 12% to 57 ± 4%) at 3 weeks. Generally, the levels reduced progressively towards the end of experiment.