Probiotic legacy effects on gut microbial assembly in tilapia larvae.
ABSTRACT: The exposure of fish to environmental free-living microbes and its effect on early colonization in the gut have been studied in recent years. However, little is known regarding how the host and environment interact to shape gut communities during early life. Here, we tested whether the early microbial exposure of tilapia larvae affects the gut microbiota at later life stages. The experimental period was divided into three stages: axenic, probiotic and active suspension. Axenic tilapia larvae were reared either under conventional conditions (active suspension systems) or exposed to a single strain probiotic (Bacillus subtilis) added to the water. Microbial characterization by Illumina HiSeq sequencing of 16S rRNA gene amplicons showed the presence of B. subtilis in the gut during the seven days of probiotic application. Although B. subtilis was no longer detected in the guts of fish exposed to the probiotic after day 7, gut microbiota of the exposed tilapia larvae remained significantly different from that of the control treatment. Compared with the control, fish gut microbiota under probiotic treatment was less affected by spatial differences resulting from tank replication, suggesting that the early probiotic contact contributed to the subsequent observation of low inter-individual variation.
Project description:<h4>Backgrounds and aims</h4>Aluminum contamination of water is becoming increasingly serious and threatens the health status of fish. <i>Lactobacillus plantarum</i> CCFM639 was previously shown to be a potential probiotic for alleviation aluminum toxicity in Nile tilapia. Considering the significant role of the gut microbiota on fish health, it seems appropriate to explore the relationships among aluminum exposure, probiotic supplementation, and the gut microbiota in Nile tilapia and to determine whether regulation of the gut microbiota is related to alleviation of aluminum toxicity by a probiotic in Nile tilapia.<h4>Methods and results</h4>The tilapia were assigned into four groups, control, CCFM639 only, aluminum only, and aluminum + CCFM639 groups for an experimental period of 4 weeks. The tilapia in the aluminum only group were grown in water with an aluminum ion concentration of 2.73 mg/L. The final concentration of CCFM639 in the diet was 10<sup>8</sup> CFU/g. The results show that environmental aluminum exposure reduced the numbers of <i>L. plantarum</i> in tilapia feces and altered the gut microbiota. As the predominant bacterial phyla in the gut, the abundances of Bacteroidetes and Proteobacteria in aluminum-exposed fish were significantly elevated and lowered, respectively. At the genus level, fish exposed to aluminum had a significantly lower abundance of <i>Deefgea</i>, <i>Plesiomonas,</i> and <i>Pseudomonas</i> and a greater abundance of <i>Flavobacterium</i>, <i>Enterovibrio</i>, <i>Porphyromonadaceae uncultured</i>, and <i>Comamonadaceae</i>. When tilapia were exposed to aluminum, the administration of a probiotic promoted aluminum excretion through the feces and led to a decrease in the abundance of <i>Comamonadaceae, Enterovibrio</i> and <i>Porphyromonadaceae</i>. Notably, supplementation with a probiotic only greatly decreased the abundance of <i>Aeromonas</i> and <i>Pseudomonas</i>.<h4>Conclusion</h4>Aluminum exposure altered the diversity of the gut microbiota in Nile tilapia, and probiotic supplementation allowed the recovery of some of the diversity. Therefore, regulation of gut microbiota with a probiotic is a possible mechanism for the alleviation of aluminum toxicity in Nile tilapia.
Project description:This study investigated the effects of the <i>Streptococcus agalactiae</i> antagonizing probiotics <i>Bacillus cereus</i> NY5 and <i>Bacillus subtilis</i> as feed additives for Nile tilapia in terms of growth performance, intestinal health and resistance to <i>S. agalactiae</i>. A total of 720 apparently healthy juvenile Nile tilapia (0.20 ± 0.05 g) were randomly divided into 4 equal groups with 3 replicates for each group. Fish were fed a basal diet (control check group, CK group) supplemented with <i>B. subtilis</i> (1 × 10<sup>8</sup> CFU/g feed, BS group), <i>B. cereus</i> NY5 (1 × 10<sup>8</sup> CFU/g feed, BC group), and <i>B. subtilis</i> + <i>B. cereus</i> NY5 (0.5 × 10<sup>8</sup> CFU/g feed of each probiotic, BS + BC group) for 6 wk, and the probiotic supplementation groups were then fed the basal diet for 1 wk to investigate the gut microbial community. The results of this study showed that BS + BC and BC treatments significantly increased weight gain (WG), feed conversion ratio (FCR) and <i>S</i>. <i>agalactiae</i> resistance in Nile tilapia (<i>P</i> < 0.05). Gut microvilli length and density and c-type lysozyme (<i>lyzc</i>) gene expression were significantly increased by probiotic supplementation (<i>P</i> < 0.05). The results of high-throughput sequencing showed that the <i>B. cereus</i> NY5 and <i>B. subtilis</i> + <i>B. cereus</i> NY5-supplemented feed resulted in a significant improvement in tilapia autochthonous gut bacterial communities and had a stimulation effect on a variety of potential probiotics after 6 wk of feeding. After cessation of probiotic administration for 1 wk, the gut bacteria of the fish in the BS + BC and BC groups had minor changes and maintained a stable state. Consequently, it was inferred that, as a feed supplement, <i>B. cereus</i> NY5 and the mixture of <i>B. subtilis</i> and <i>B. cereus</i> NY5 at 1 × 10<sup>8</sup> CFU/g feed were able to promote growth and disease resistance, which may be associated with the supplement's effects on gut immune status, intestinal morphology, and intestinal microbial community composition.
Project description:<h4>Background</h4>Fish gut microbial colonisation starts during larval stage and plays an important role in host's growth and health. To what extent first colonisation could influence the gut microbiome succession and growth in later life remains unknown. In this study, Nile tilapia embryos were incubated in two different environments, a flow-through system (FTS) and a biofloc system (BFS); hatched larvae were subsequently cultured in the systems for 14 days of feeding (dof). Fish were then transferred to one common recirculating aquaculture system (RAS1, common garden, 15-62 dof), followed by a growth trial in another RAS (RAS2, growth trial, 63-105 dof). In RAS2, fish were fed with two types of diet, differing in non-starch polysaccharide content. Our aim was to test the effect of rearing environment on the gut microbiome development, nutrient digestibility and growth performance of Nile tilapia during post-larvae stages.<h4>Results</h4>Larvae cultured in the BFS showed better growth and different gut microbiome, compared to FTS. After the common garden, the gut microbiome still showed differences in species composition, while body weight was similar. Long-term effects of early life rearing history on fish gut microbiome composition, nutrient digestibility, nitrogen and energy balances were not observed. Still, BFS-reared fish had more gut microbial interactions than FTS-reared fish. A temporal effect was observed in gut microbiome succession during fish development, although a distinct number of core microbiome remained present throughout the experimental period.<h4>Conclusion</h4>Our results indicated that the legacy effect of first microbial colonisation of the fish gut gradually disappeared during host development, with no differences in gut microbiome composition and growth performance observed in later life after culture in a common environment. However, early life exposure of larvae to biofloc consistently increased the microbial interactions in the gut of juvenile Nile tilapia and might possibly benefit gut health.
Project description:We previously determined that several diets used to rear <i>Aedes aegypti</i> and other mosquito species support the development of larvae with a gut microbiota but do not support the development of axenic larvae. In contrast, axenic larvae have been shown to develop when fed other diets. To understand the mechanisms underlying this dichotomy, we developed a defined diet that could be manipulated in concert with microbiota composition and environmental conditions. Initial studies showed that axenic larvae could not grow under standard rearing conditions (27 °C, 16-h light: 8-h dark photoperiod) when fed a defined diet but could develop when maintained in darkness. Downstream assays identified riboflavin decay to lumichrome as the key factor that prevented axenic larvae from growing under standard conditions, while gut community members like <i>Escherichia coli</i> rescued development by being able to synthesize riboflavin. Earlier results showed that conventional and gnotobiotic but not axenic larvae exhibit midgut hypoxia under standard rearing conditions, which correlated with activation of several pathways with essential growth functions. In this study, axenic larvae in darkness also exhibited midgut hypoxia and activation of growth signaling but rapidly shifted to midgut normoxia and arrested growth in light, which indicated that gut hypoxia was not due to aerobic respiration by the gut microbiota but did depend on riboflavin that only resident microbes could provide under standard conditions. Overall, our results identify riboflavin provisioning as an essential function for the gut microbiota under most conditions <i>A. aegypti</i> larvae experience in the laboratory and field.
Project description:The gut microbiota of fish larvae evolves fast towards a complex community. Both host and environment affect the development of the gut microbiota; however, the relative importance of both is poorly understood. Determining specific changes in gut microbial populations in response to a change in an environmental factor is very complicated. Interactions between factors are difficult to separate and any response could be masked due to high inter-individual variation even for individuals that share a common environment. In this study we characterized and quantified the spatio-temporal variation in the gut microbiota of tilapia larvae, reared in recirculating aquaculture systems (RAS) or active suspension tanks (AS). Our results showed that variation in gut microbiota between replicate tanks was not significantly higher than within tank variation, suggesting that there is no tank effect on water and gut microbiota. However, when individuals were reared in replicate RAS, gut microbiota differed significantly. The highest variation was observed between individuals reared in different types of system (RAS vs. AS). Our data suggest that under experimental conditions in which the roles of deterministic and stochastic factors have not been precisely determined, compositional replication of the microbial communities of an ecosystem is not predictable.
Project description:In the present study, we evaluated the effects of administering Enterococcus faecium in food and/or water on the hematological and immunological parameters, intestinal microbiota, resistance to bacterial diseases (streptococcosis and francisellosis) and growth of Nile tilapia. Before the in vivo experiment, probiotic bacteria isolated from Nile tilapia were selected via inhibition tests. Sequencing, annotation, and assembly of the complete genome of the selected bacteria as well as other tests were performed using bioinformatics tools. Three treatments were implemented: G1 (probiotic feeding), G2 (probiotic in water), and G3 (probiotic in food and water); and a negative control (NC) was also employed. Treatment lasted 38 days, and each group consisted of fish and two repetitions. The fish were divided and infected with Streptococcus agalactiae S13 (serotype Ib) and Francisella orientalis. The G1 group had a higher average final weight gain than the G2, G3, and NC groups. Further, a significant increase in the number of thrombocytes was observed in the groups administered probiotics in the diet (G1 and G3). A statistical difference was observed in the mortality of fish infected with S. agalactiae in the NC compared to the treated groups. Cetobacterium was the 43 most abundant genus in the intestinal microbiota of all groups, including the NC group. E. faecium increased the immunity of fish administered the treatment and decreased the mortality caused by S. agalactiae. As an autochtone probiotic, E. faecium does not interfere with the local ecosystem and thus has a great probiotic potential for Nile tilapia in Brazil.
Project description:Establishment of the early-life gut microbiota has a large influence on host development and succession of microbial composition in later life stages. The effect of commensal yeasts - which are known to create a conducive environment for beneficial bacteria - on the structure and diversity of fish gut microbiota still remains unexplored. The present study examined the intestinal bacterial community of zebrafish (Danio rerio) larvae exposed to two fish-derived yeasts by sequencing the V4 hypervariable region of bacterial 16S rRNA. The first stage of the experiment (until 7 days post-fertilization) was performed in cell culture flasks under sterile and conventional conditions for germ-free (GF) and conventionally raised (CR) larvae, respectively. The second phase was carried out under standard rearing conditions, for both groups. Exposure of GF and CR zebrafish larvae to one of the yeast species Debaryomyces or Pseudozyma affected the bacterial composition. Exposure to Debaryomyces resulted in a significantly higher abundance of core bacteria. The difference was mainly due to shifts in relative abundance of taxa belonging to the phylum Proteobacteria. In Debaryomyces-exposed CR larvae, the significantly enriched taxa included beneficial bacteria such as Pediococcus and Lactococcus (Firmicutes). Furthermore, most diversity indices of bacterial communities in yeast-exposed CR zebrafish were significantly altered compared to the control group. Such alterations were not evident in GF zebrafish. The water bacterial community was distinct from the intestinal microbiota of zebrafish larvae. Our findings indicate that early exposure to commensal yeast could cause differential bacterial assemblage, including the establishment of potentially beneficial bacteria.
Project description:Tilapia (Oreochromis mossambicus) is one of the most invasive fish found throughout the World and emerged as a major threat to the indigenous fishes in many countries. Investigating the gut microbial diversity of such fishes is one of the ways to understand its physiology. In the present study, we have explored the gut microbial community structure of tilapia using 16S rRNA gene sequencing on the Illumina Miseq platform. Our study showed significant differences in tilapia gut microbiota collected from different habitats (i.e. river and lakes) suggesting the influence of habitat on the gut microbial diversity of tilapia. This study gives a first insight into the mossambicus tilapia gut microbiota and provides a reference for future studies.
Project description:Estrogenic chemicals are widespread environmental contaminants associated with diverse health and ecological effects. During early vertebrate development, estrogen receptor signaling is critical for many different physiologic responses, including nervous system function. Recently, host-associated microbiota have been shown to influence neurodevelopment. Here, we hypothesized that microbiota may biotransform exogenous 17-?estradiol (E2) and modify E2 effects on swimming behavior. Colonized zebrafish were continuously exposed to non-teratogenic E2 concentrations from 1 to 10 days post-fertilization (dpf). Changes in microbial composition and predicted metagenomic function were evaluated. Locomotor activity was assessed in colonized and axenic (microbe-free) zebrafish exposed to E2 using a standard light/dark behavioral assay. Zebrafish tissue was collected for chemistry analyses. While E2 exposure did not alter microbial composition or putative function, colonized E2-exposed larvae showed reduced locomotor activity in the light, in contrast to axenic E2-exposed larvae, which exhibited normal behavior. Measured E2 concentrations were significantly higher in axenic relative to colonized zebrafish. Integrated peak area for putative sulfonated and glucuronidated E2 metabolites showed a similar trend. These data demonstrate that E2 locomotor effects in the light phase are dependent on the presence of microbiota and suggest that microbiota influence chemical E2 toxicokinetics. More broadly, this work supports the concept that microbial colonization status may influence chemical toxicity.
Project description:The majority of seafood is farmed, with most finfish coming from freshwater ponds. Ponds are often fertilized to promote microbial productivity as a natural feed source to fish. To understand if pond fertilization with livestock manure induces a probiotic or prebiotic effect, we communally reared tilapia (Oreochromis shiranus), and North African catfish (Clarias gariepinus), for 4 weeks under seven manure treatments including layer chicken, broiler chicken, guinea fowl, quail, pig, cow, vs. commercial feed to evaluate microbial community dynamics of the manure, pond water, and fish feces using 16S and 18S rRNA marker genes along with metagenome sequencing. Catfish growth, but not tilapia, was positively associated with plankton abundance (p = 0.0006, R2 = 0.4887) and greatest in ponds fertilized with quail manure (ANOVA, p < 0.05). Manure was unique and influenced the 16S microbiome in pond water, tilapia gut, and catfish gut and 18S community in pond water and catfish guts (PERMANOVA, p = 0.001). On average, 18.5%, 18.6%, and 45.3% of manure bacteria sOTUs, (sub-operational taxonomic units), were present in the water column, catfish feces, and tilapia feces which comprised 3.7%, 12.8%, and 10.9% of the total microbial richness of the communities, respectively. Antibiotic resistance genes were highest in the manure and water samples followed by tilapia feces and lowest in catfish feces (p < 0.0001). In this study, we demonstrate how the bacterial and eukaryotic microbial composition of fish ponds are influenced by specific livestock manure inputs and that the gut microbiome of tilapia is more sensitive and responsive than catfish to these changes. We conclude that animal manure used as fertilizer induces a primarily prebiotic effect on the pond ecosystem rather than a direct probiotic effect on fish.