Bioinformática aplicada à conservação de anfíbios: probióticos em microbiomas de ninho de espuma

Abstract

Amphibians are important environmental bioindicators; however, many species face population declines due to chytridiomycosis caused by the fungus Batrachochytrium dendrobatidis (Bd). Foam nests are reproductive structures of some amphibians that, in addition to protecting eggs, harbor beneficial bacteria, such as those belonging to the genus Pseudomonas, which may have anti-Bd activity. Given this, this project aimed to assess changes in the bacteriome of Leptodactylus vastus foam nests after bioaugmentation — a widely used inoculation technique to increase the proportion of a bacterium in a community — using a strain of Pseudomonas putida. The bacterial community structure and its impacts over time were analyzed using bioinformatics tools. For this purpose, samples of Leptodactylus vastus foam nests were subjected to bioaugmentation with Pseudomonas and analyzed by sequencing the V4 region of 16S rRNA. The sequences were processed in RStudio using DADA2 and classified with the SILVA 138 SSU database. Diversity analyses and differential abundance assessments were conducted using the Phyloseq, Vegan, and igraph packages, allowing for the evaluation of microbiome structure and bacterial interactions. A total of 3,675 ASVs were obtained after the removal of eukaryotic and archaeal sequences. Rarefaction curves indicated good sample coverage. Alpha diversity analysis revealed significant differences between treatments, with bioaugmentation impacting microbial structure. Abundance analysis showed that Proteobacteria was dominant, but Pseudomonas inoculation altered microbial composition, especially at Time 1. Beta diversity analysis demonstrated that time had a greater influence on microbial structure than bioaugmentation. Interaction networks indicated that the addition of Pseudomonas affected bacterial community connectivity and modularity over time. Thus, the study showed that the addition of the strain significantly influenced microbial diversity in foam nests, with bioaugmentation duration being the most significant factor. Additionally, the bacterium exhibited low adhesion to the community but indirectly influenced the microbiome by altering ecological interactions. However, further research on the relationship between the bacterium and the bacteriome, as well as Bd testing, is necessary to understand the role of the probiotic and its viability for future applications.