An integrated gene catalog of the zebrafish gut microbiome reveals significant homology with mammalian microbiomes

Abstract

24Gut microbiome research increasingly utilizes zebrafish (Danio rerio) given their amenability to 25 high-throughput experimental designs. However, the utility of zebrafish for discerning 26 translationally relevant host-microbiome interactions is constrained by a paucity of knowledge 27 about the biological functions that zebrafish gut microbiota can execute, how these functions 28 associate with zebrafish physiology, and the degree of homology between the genes encoded by 29 the zebrafish and human gut microbiomes. To address this knowledge gap, we generated a 30 foundational catalog of zebrafish gut microbiome genomic diversity consisting of 1,569,102 non-31 redundant genes from twenty-nine individual fish. We identified hundreds of novel microbial 32 genes as well as dozens of biosynthetic gene clusters of potential clinical interest. The genomic 33 diversity of the zebrafish gut microbiome varied significantly across diets and this variance 34 associated with altered expression of intestinal genes involved in inflammation and immune 35 activation. Zebrafish, mouse, and human fecal microbiomes shared > 50% of their total genomic 36 diversity and the vast majority of gene family abundance for each individual metagenome 37 (~99%) was accounted for by genes that comprised this shared fraction. These results indicate 38 that the zebrafish gut houses a functionally diverse microbial community that manifests 39 extensive homology to that of humans and mice despite substantial disparities in taxonomic 40 composition. We anticipate that the gene catalog developed here will enable future mechanistic 41 study of host-microbiome interactions using the zebrafish model. 42 43 Keywords: microbiome, metagenome, intestine, vertebrate, zebrafish, diet 44 45 46 also identify substantial homology between zebrafish, human, and mouse metagenomic diversity, 55 indicating that these microbiomes may operate similarly. 56 57 58 59 60 61 62 63 64 65 66 67 68 69 Nextera XT kit (Illumina, San Diego, CA USA) and sequenced on and Illumina HiSeq 3000 130 using a 150bp PE sequencing kit (Illumina). 131 132 Metagenome Assembly and Functional Annotation 133Each metagenomic library was demultiplexed and quality filtered with shotcleaner 134 (https://github.com/sharpton/shotcleaner) using the zebrafish genome (GRCz10). The cleaned 135 sequence files were then assembled simultaneously using MEGAHIT 15 with a minimum kmer 136 size of 27 and a kmer increment size of 10. The resulting contigs were used to predict genes 137 using prodigal in metagenome mode. CD-HIT 16 was used to collapse redundant genes (95% 138

Opportunistic gill infection is associated with TiO2 nanoparticle-induced mortality in zebrafish

“…Through metagenomic analysis, we discovered that bacteria found in the gill samples of zebrafish with NP-induced injury, such as the phyla Proteobacteria, Bacteroidetes, and Actinobacteria, accounted for more than 95% of the total bacteria count; these types of bacteria are all found in normal zebrafish gut microflora [52][53][54][55]. This result is likely attributable to the fact that the zebrafish in the experiment were kept under pathogen-free conditions; no pathogens other than normal flora were present.…”

“…The relative abundance (% relative to the total) of the bacteria populations was analyzed in specific hypervariable regions of 16S ribosomal RNA through new-generation sequencing analyses (Fig 4 and S2 Fig in S1 Data). We found that the phyla Proteobacteria, Bacteroidetes, and Actinobacteria, all of which are bacteria found in normal zebrafish gut microbiomes [52][53][54][55], accounted for more than 95% of the total bacteria counts (Fig 4). Because the fish culture conditions did not include pathogens, these results indicate that TiO 2 NP-induced zebrafish gill injury is associated with opportunistic infection of the gut normal flora.…”

Genetic Approaches Using Zebrafish to Study the Microbiota–Gut–Brain Axis in Neurological Disorders

“…Indeed, host gut microbiota interactions [ 124 ] and neurological disorders [ 125 , 126 ] that mirror human conditions have been independently investigated using zebrafish. However, the combined efforts of the characterization of recently developed zebrafish disease models for ASD and AD, the establishment of microbial approaches including GF culture [ 127 , 128 ] and metagenomic analyses of the microbiota [ 129 , 130 ], and the expansion of their neurobehavioral repertoires [ 131 , 132 , 133 , 134 ] may allow dissection of the signaling pathways for the host–bacteria interactions in neurological disorders, discussed in the following sections. The key advantages of the zebrafish model for studying the MGBA are summarized in Figure 2 .…”

“…Although the exact taxonomies of the microbial communities in humans, mice, and zebrafish differ, there is a trend towards interspecies conservation at the phylum level [ 187 , 188 ]. In addition, host responses to gut colonization by microbiota have been reported to be significantly shared between zebrafish and mouse models [ 189 ], and zebrafish, mouse, and human microbiomes are known to have similar abundances of functional pathways (i.e., DEG pathways) [ 130 ] which suggests that functional and mechanistic findings using the zebrafish microbiota may have direct translational implications for understanding human microbiota functions.…”

Conserved anti-inflammatory effects and sensing of butyrate in zebrafish

“…9 Previous genomic investigation has identified the presence of butyrate metabolic genes in the zebrafish gut metagenome; however, the key bacterial enzyme for butyrate synthesis butyryl coenzyme A (CoA):acetate-CoA transferase has not been specifically detected. 9,10 Acetate, butyrate, and propionate have been measured in several species of teleosts. [11][12][13][14][15] We have used conservation between mammalian intestinal function and immunity to create zebrafish models of human intestinal inflammation.…”