Host-microbiome associations in livebearing fishes adapted to toxic streams rich in hydrogen sulfide: Code for analyzing 16S rRNA dataset

Organisms inhabiting extreme environments must tolerate a variety of physiochemical stressors. In some cases, host-associated microbial communities facilitate survival of their hosts in extreme environments, but extremophile symbioses have not been identified in vertebrates. We used 16S rRNA amplicon sequencing to investigate commonalities and differences in the gut microbial communities of livebearing fish (Poecilia mexicana species complex, Poeciliidae) that have repeatedly colonized toxic sulfide streams in southern Mexico. We found shared core gut microbiomes across habitat types and drainages, but also differences in the microbiomes between sulfidic and nonsulfidic populations, both in terms of patterns of diversity and community composition. Most importantly, we documented convergent changes in microbiome composition across evolutionarily independent sulfide spring lineages. These patterns were consistent when we analyzed whole gut microbiomes and primarily host-associated microbi..., We sampled 8 populations of P. mexicana throughout the Río Grijalva basin in southern Mexico, dissected the intestines, preserved the whole intestines in ethanol, and used 16S rRNA amplicon sequencing to characterize the gut microbiomes. Fish were sampled from four river drainages (Tacotalpa, Ixtapangajoya, Pichucalco, Puyacatengo), and each drainage contained a sulfide-adapted and a sulfide-intolerant population of P. mexicana. Additionally, we collected water and sediment samples from all of the sites to characterize the environmental microbiomes, and we collected intestine samples from lab-raised fish following the same methods as in the field. The amplicon library was prepared following the Illumina MiSeq System 16S Metagenomic Sequencing Library Preparation protocol (Illumina), and it was sequenced on an Illumina MiSeq at the Kansas State University Integrated Genomics Facility. The sequences were processed using QIIME2, and the taxonomy count matrices are included here as inp..., , # Host-microbiome associations in livebearing fishes adapted to toxic streams rich in hydrogen sulfide: Code for analyzing 16S rRNA dataset

[https://doi.org/10.5061/dryad.dz08kps7v](https://doi.org/10.5061/dryad.dz08kps7v)

## Description of the data and file structure

We sampled four population pairs of sulfide-adapted and sulfide-intolerant populations of livebearing fishes in the *P. mexicana* species complex throughout the Río Grijalva basin in southern Mexico to investigate patterns of gut microbial diversity between ecotypes and across river drainages. We additionally sampled environmental microbiomes from these sites, as well as gut microbiomes of *P. mexicana* raised in a laboratory setting. We aimed to understand whether sulfide-adapted lineages have commonalities in their gut microbiomes and identify microbes of interest for future research investigating potentially adaptive host-microbiome associations.

### Files and variables

#### File: Labonly-merged-reads-level-6-taxon...,