Assessments of fish gut microbiota during development, and in response to environmental and dietary change

Lead Research Organisation: University of Liverpool
Department Name: Institute of Integrative Biology

Abstract

Background: With human population proliferating at an ever alarming rate, and with billions of individuals presently living on the planet starving or suffering from nutrient deficiencies, producing more fish can make an important contribution. The nutritional benefits of fish consumption have a positive link to increased food security and decreased poverty rates in developing countries. Fish, particularly produced through aquaculture, is commonly cheaper than other animal feed, and can be consumed by lower income individuals. Fish also contains much higher protein levels, as well as other important minerals and vitamins. Increased availability of fish can mean better health and a more diverse diet.
Undoubtedly promoting and maintaining fish health is paramount for enhancing fish production.
Extensive research has shown that microbes that reside in the intestine have important roles in maintaining and promoting the health of the host. They can breakdown some food materials to some essential nutrients and vitamins important to maintaining immunity, health and well-being of the host. The microbes also protect the gut from colonization by pathogenic bacteria.
The identity of fish gut microbial composition and how they can be influenced by environmental, dietary and developmental factors is in its infancy. There are carnivorous, herbivorous and omnivorous fish which naturally select specific diets. We do not know if their digestive physiology can adapt to consume commercial feed. This knowledge is important of formulation of appropriate feed to avoid digestive problems.
In this project, a group of scientists knowledgeable in nutrition, intestinal physiology/microbiology and intestinal adaptation to dietary change aim to assess the impact of i) habitat (farms), ii) fish species of different dietary habits, iii) diet and iv) fish maturation, on the microbial populations that reside in the fish intestine.
The intestinal content of 1) adult carnivorous, herbivorous and omnivorous fish obtained from different locations (farms) and 2) fry of the same species, grown under controlled conditions and maintained on different defined diets, will be used. Using modern technologies, the precise composition of microbial populations, and the nutrients they produce, will be determined. Other bodily parameters such as growth, food conversion ratio, and gut morphology will be assessed.
Anticipated outcomes are:
Proving information on microbial populations that reside in intestines of herbivorous, carnivorous and omnivorous fish.
Identifying contributions of environment, diet and fish species on the composition and activity of gut microbes.
Gaining knowledge on the responses of predominant microbial populations to environment, diet, host and developmental signals.
Impact: Characterization of gut microbiota during fish maturation and in fish with different dietary habits will allow scientifically-based feed (feed supplement) formulation to enhance selective gut microbial populations that promote fish growth, health and productivity.
Benefits from this research are to:
Fish: The correct diet will maintain and promote fish health and welfare, prevent disease and increase productivity.
The public: Increasing availability of fish means better health and more diverse diet.
Society and Economy: Contributing towards wealth creation and economic prosperity. Commercial aquaculture has the potential to stimulate economic growth and create jobs.
R&D investment from global business.
The commercialisation and exploitation of scientific knowledge, leading to interaction with industries.

Technical Summary

General aim: to comprehensively assess the impact of habitat, species, diet and fish maturation on the intestinal microbiota of three different classes of fish: herbivores (grass carp; Ctenopharyngodon idella), omnivores (common carp; Cyprinus carpio) and carnivores (rainbow trout; Oncorhynchus mykiss). In addition to quantitatively determine the levels of microbial-derived metabolites within the intestinal tracts of the same fish.

Obtaining fish from different locations (farms) we aim to identify i) whether host type or habitat has the greater influence on intestinal microbiota and metabolites, within the same species, ii) inter-individual variation within the same species raised in the same habitat, and iii) differences in microbiota and metabolites between disparate species.

Growing fry under controlled conditions and maintained on defined diets we aim to determine influences of diet and/or ontogenetic signals on resident gut microbiota and metabolites during different maturation stages (fry; adolescent; adult).

Bacterial metabolites will be measured in intestinal contents.
Feed consumption rate, fish weight and gut morphology will be assessed during feeding trials.

Methodology: Bacterial DNA will be extracted from fish intestinal contents, purified and quantified. Next-generation sequencing will be used to analyze 16S rRNA gene sequences. Raw sequencing reads will be subjected to bioinfomatic analyses in order to provide the phylogenetic framework for describing community structure, allowing characterization of microbial ecosystems, its richness and diversity. Bacterial metabolites will be measured by gas chromatography and quantified in relation to an internal standard. Feed consumption rate, weight and gut morphology will be determined by measuring various body parameters and by morphometric analyses.

Planned Impact

Academic Impact:
The outcome of this research will act as a platform for further studies in fish addressing issues such as the identification of safe and effective replacements for antibiotics, effect of environmental change and exposure to toxins and pollutants.

The outcome of this project will provide new knowledge that enhances scientific advancement nationally and internationally. It will offer valuable information for scientists and practitioners of diverse disciplines including physiologists, veterinarians, nutritionists, microbiologists, molecular biologists and geneticists. It will contribute to developing expertise and knowledge in multi-disciplinary areas.

Economic and Social Impact:
The audience benefiting from this research are:
The public: Increasing availability of fish means better health and more diverse diet.
Society and Economy: Contributing towards wealth creation and economic prosperity. Commercial aquaculture has the potential to stimulate economic growth and create jobs.
R&D investment from global business.
The commercialisation and exploitation of scientific knowledge, leading to interaction with industries.

Publications

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Daly K (2019) Host selectively contributes to shaping intestinal microbiota of carnivorous and omnivorous fish. in The Journal of general and applied microbiology

 
Description During the first year of research, we developed an innovative method for isolating bacterial DNA from fish intestinal content. Using this methodology and subsequently sequencing the DNA isolated from the intestine of a carnivorous fish (trout) and an omnivorous fish (carp), we have identified that in both fish types: 1) there are distinct differences in gut bacteria of two fish with different dietary habits, 2) gut bacterial populations are very different to those present in the mammalian intestine, with fish having a comparatively simple microbiota, and 3) the host has a profound effect on the composition and diversity of fish gut microbiota. These findings are significant and demonstrate the importance of host in shaping fish gut bacterial composition. This work has now been published.

Subsequently, we performed controlled trials to assess the impact of dietary change on fish gut microbiota. We maintained 2 omnivorous fish (carp and tilapia) and a carnivorous fish (trout) from fingerling stage to adulthood on their natural diet and an alternative diet (carnivorous vs. omnivorous or reverse). Using bacterial DNA extracted from intestinal contents we generated 16S rRNA gene amplicon libraries for characterisation of the gut microbiota by next-generation sequencing. Due to significant delays in sequencing and subsequent analysis caused by the coronavirus pandemic, we are only now in a position to analyse those datasets and prepare results for publication. Preliminary analysis shows that changes in diet and development can have significant effects on microbiota composition and that these changes differ between the fish species used. We aim to publish these results in two manuscripts in 2021.
Exploitation Route The findings from year 1 of the award provided the important basis for the phase 2 of the work in which the laboratory work has been completed, and the microbiome sequence data are under bioinformatics analysis. Unfortunately, this part of the work was severely impacted and delayed by the coronavirus pandemic. We aim to demonstrate in a well-controlled study (where we know precisely dietary compositions and environmental conditions) the contribution of fish development and dietary change on composition and diversity of fish gut microbiota.
Our findings so far have a significant impact for the academic community and fish feed industry. Results show that there are host influences on the gut microbiota of fish leading to species-specific variations. Our new data (Manuscripts in preparation) have indicated that dietary factors and developmental cues influence strongly bacterial community structure, demonstrating plasticity of fish gut microbiota, and the importance of diet in overriding host influences. This knowledge will assist in developing strategies to promote a beneficial gut microbiome for the improvement of health and productivity in farmed fish.
Sectors Agriculture, Food and Drink,Environment,Other

 
Title Fish Gut Microbiota Sequencing Data 
Description Illumina 16S rRNA gene sequencing dataset of intestinal microbiota of common carp and rainbow trout sourced from various farm locations in the United Kingdom. 
Type Of Material Database/Collection of data 
Year Produced 2019 
Provided To Others? Yes  
Impact Using bacterial 16S rRNA sequencing, we have identified that in the intestine of a carnivorous fish (rainbow trout) and an omnivorous fish (common carp): 1) there are distinct differences in gut bacteria of two fish with different dietary habits, 2) gut bacterial populations are very different to those present in the mammalian intestine, with fish having a comparatively simple microbiota, and 3) the host has a profound effect on the composition and diversity of fish gut microbiota. These findings have a significant impact for the academic community and fish feed industry and demonstrate that the gut microbiota of fish is influenced by host-derived factors leading to species-specific variations. 
URL https://www.ebi.ac.uk/ena/browser/view/PRJEB18770