探索斑腿樹蛙腸道菌以及其網絡關係

Abstract

無中文摘要The concerted activity of intestinal microbes is crucial to the health and development of their host organisms. Studies have suggested that microbial assemblages in the intestine of animals are engines of globally important host physiological processes between hibernating and non-hibernating states. The advances in Next Generation Sequencing (NGS) technologies facilitate our understanding of gut microbiota with high resolutions in diversity and metabolic functioning between hibernating and non-hibernating seasons. Polypedates megacephalus is an invasive species in Taiwan since 2006. The approach of habitat usage and population dispersal of this invasive treefrogs across seasons have suggested a rapid expansion across counties in Taiwan. However, it still lacks an effective solution to control the expansion of invasive P. megacephalus. Due to the reciprocal interactions between gut microbiota and host physiology, I attempt to explore gut microbiota of P. megacephalus, decipher microbial interactions to understand the potential mechanisms of microbial ecosystem, and further manipulate host response by modulating gut microbiota according to the guidance by computational network analysis. This study not only delineated seasonal changes of gut microbiota in composition and metabolic functioning but demonstrated the potentials of computational network inference toward practical applications on animal systems. The compositional and predicted functional changes of gut microbiota across non-hibernating and artificial hibernating seasons were identified based on 16S rRNA amplicon analysis. The abundance profile and predicted functions of microbial community significantly change between artificial hibernating (AH) and non-hibernating (NH) treefrogs. Artificial hibernation significantly reduces microbial diversity and the level of Firmicutes and increases the level of Proteobacteria in the treefrog gut microbiota. In addition, AH treefrogs harbor core taxonomic units that are rarely abundant in NH treefrogs. Moreover, artificial hibernation significantly increased relative abundance of red-leg syndrome-related genera such as Citrobacter and Aeromonas. Functional predictions via PICRUSt and Tax4Fun suggested that artificial hibernation has effects on most pathways including metabolism and signal transduction. These results suggest that artificial hibernation restructure gut microbiota in treefrogs and significantly reduce microbial complexity of gut microbiome. The use of computational methods to decipher microbial interactions have been applied on microbiome data in a time-series fashion. A time-series microbiome data could monitor the population changes of each bacterium in the community over time. Due to the adjustable gut microbiome complexity of hibernating animals, the growth microbiome time-series (GMT) dataset is proposed to apply on the computational network inference methods. Among varieties of network inference tools, regression-based network model is selected and utilized due to its better performance tested by using in silico dataset. Lotka-Volterra models, also known as predator–prey equations, are the most currently used regression-based method, and predict both dynamics of microbial communities and how communities are structured and sustained. The interaction network of gut microbiotaat the genus level in the treefrog was constructed using Metagenomic Microbial Interaction Simulator (MetaMIS) package. The interaction network contained 1,568 commensal, 1,737 amensal, 3,777 mutual, and 3,232 competitive relationships, e.g., Lactococcus garvieae has a commensal relationship with Corynebacterium variabile. To validate the interacting relationships, I took advantage of probiotic system to evaluate the responses of gut microbiota to the probiotic trials. The trials involved different groups including single strain (L. garvieae, C. variabile, and Bacillus coagulans, respectively) and a combination of L. garvieae, C. variabile, and B. coagulans, because of the cooperative relationship among their respective genera identified in the interaction network. After a two-week trial, the combination of cooperative microbes yielded significantly higher probiotic concentrations than single strains, and the immune response (interleukin-10 expression) also significantly changed in a manner consistent with improved probiotic effects.