Genome-resolved metagenomics, based on shotgun sequencing, has become a powerful strategy for investigating animal-associated microbiomes, due its heightened capability for delivering detailed taxonomic, phylogenetic, and functional insights compared to amplicon sequencing-based approaches. While genome-resolved metagenomics holds promise across various non-lethal sample types, their effectiveness in yielding high-quality metagenome-assembled genomes (MAGs) remains largely unexplored. Our investigation of fecal and cloacal microbiota of the mesquite lizards (Sceloporus grammicus) using genome-resolved metagenomics revealed that fecal samples contributed 97% of the 127 reconstructed bacterial genomes, whereas only 3% were recovered from cloacal swabs, which were largely enriched with host DNA. Taxonomic, phylogenetic and functional alpha microbial diversity was greater in fecal samples than in cloacal swabs. We also observed significant differences in microbial community composition between sampling methods, and higher inter-individual variation in cloacal swabs. Bacteroides, Phocaeicola and Parabacteroides (all Bacteroidota) were more abundant in the feces, whereas Hafnia and Salmonella (both Pseudomonadota) increased in the cloaca. Functional analyses showed that metabolic capacities of the microbiota to degrade polysaccharides, sugars and nitrogen compounds were enriched in fecal samples, likely reflecting the role of the microbiota in nutrient metabolism. Overall, our results indicate that fecal samples outperform cloacal swabs in characterizing microbial assemblages within lizards using genome-resolved metagenomics.

Freshwater habitats are under increasing pressure from numerous anthropogenic forces, including the introduction of alien species capable of altering ecosystems and threatening native species. Although alien species themself are likely to experience loss of genetic diversity when colonising novel environments, some manage to become invasive, suggesting that other factors might facilitate their adaptive capacity. Using a hologenomic approach, we elucidate population genomic trends, the gut microbiota composition and genome-environment-microbiota interaction in the endemic and endangered Spanish toothcarp (Aphanius iberus) and the highly invasive Eastern mosquitofish (Gambusia holbrooki). We found clear genetic signatures of captive breeding in the populations of A. iberus, while G. holbrooki are characterised by an overall low level of heterozygosity and likely signs of multiple introductions. Gut microbial communities of the two species differed significantly across locations, but no sign of increased microbial plasticity was detected in G. holbrooki. However, we report that the genetic profile of each fish was able to explain a considerable part of the microbiota variation measured across individuals. Using shotgun metagenomics, we observed an overall high functional capacity of the microbiota in both species, but we identified no significant differences in the functional capacity between them. The role of the gut microbiota in invasive species and conservation warrant further research using direct comparisons or controlled mesocosm setups, but based on the results of the current study, the gut microbiota of invasive species

The gut microbiomes that associate with animals can represent labile units of cooperating and competing microbes. This lability, sometimes referred to as metagenomic plasticity, has been posited to have an important role as an additional axis of hosts’ phenotypic plasticity. However, whether and how metagenomic plasticity varies across hosts with different ecological and evolutionary features remains unclear. To address this, we utilised faecal-derived genome-resolved metagenomics and compared how the taxonomic, phylogenetic and functional microbial dynamics varied across a series of disturbances in two mammal species; namely, the insectivorous-specialist, Crocidura russula (N = 29) and the omnivorous-generalist Apodemus sylvaticus (N = 22). Although faecal microbial diversity of both species remained stable, compositional dynamics differed significantly. C. russula exhibited substantially higher variability and directionality of microbial responses, with higher predictability associated with each disturbance, compared to A. sylvaticus. Predictions of functional traits using joint-species distribution modelling supported these observations. C. russula showed strong functional response to perturbations, with marked directional variation of various metabolic functions. In contrast, the significantly higher functional diversity and redundancy of the A. sylvaticus microbiome likely buffered its functional response to perturbations, which remained more constant across time. Our results indicate that the intrinsic properties (e.g., diversity, redundancy) of gut microbiomes associated with animals with different biological attributes shape the taxonomic, phylogenetic, and functional response to environmental stressors. This level of plasticity might affect the capacity of animal hosts to acclimate and adapt to changing environments.