The role of hindgut microbes in protein supply to marine herbivorous fishes

Abstract

This thesis examines the role that hindgut microbes play in the protein acquisition of herbivorous fishes in temperate and tropical reefs. The aim of this study is to quantify the contribution of microbial protein to host fish tissues using stable isotope techniques. Four temperate species were collected from Great Barrier Island in the Hauraki Gulf of New Zealand, and seven tropical species were collected from Lizard Island, Great Barrier Reef, Australia. These species were used to examine trophic interactions among the liver and muscle tissues and their dietary sources (algal diet and symbiotic hindgut microbes). Using bulk carbon and nitrogen stable isotope analysis (SIA), patterns were observed in the 815N and 813C values which showed evidence of N2-fixation within the microbial community inhabiting the guts of several species. These results support previous findings by Bojarski (2014). Trophic relationships and the potential contributions of microbial protein to the fish hosts were investigated further using compound-specific analysis of carbon and nitrogen stable isotopic composition of the amino acids (AAs) within the fish tissues and potential dietary sources. An in-depth examination of isotopic routing and fractionation of dietary sources using nitrogen compound-specific stable isotope analyses of AAs (CSIA-AA), elucidated the contributions of microbial protein to the host fish tissues and the incorporation of “new” nitrogen derived from microbial N2-fixation. Carbon CSIAAA analyses further revealed the contribution of microbial AAs to the host fish, specifically the routing of essential amino acids (EAAs) biosynthesized de novo by the hindgut microbiota to the fish tissues. Carbon and nitrogen CSIA-AA analyses were also extended to identify potential metabolic pathways of AAs within the algal diet, the hindgut microbes, and the liver and muscle tissues. Finally, the data from these analyses were incorporated into a Bayesian mixing model to quantify relative contributions of the algal diet and isolated hindgut microbes to the protein in the tissues of these fish for several amino acids. The estimated contribution of microbial amino acids to host fish tissue was similar to, or in some cases, greater than the estimated contribution of the algal diet. Therefore, the results described here are the first to provide clear evidence of the contribution of microbial protein to these herbivorous fish species. This study also provides the only evidence of assimilation of “new” nitrogen, created through N2-fixation by hindgut microorganisms, to the tissues of vertebrate hosts. These findings will undoubtedly help further our understanding of microbial symbionts and the isotopic relationships between gut microbes, herbivorous diets, and host tissues.