Microorganisms play crucial roles in nutrient cycling, water quality maintenance, and farmed animal health. Increasing evidences have revealed a close association between unstable microbial environments and disease occurrences in aquaculture. Thereupon, we used high-throughput sequencing technology to comprehensively compare the bacterial communities of water, sediment, and intestine in mariculture ponds at the middle and late stages of Litopenaeus vannamei farming and analyzed whether changes of their microbiota assemblages were associated with environmental factors and shrimp physiological health. Results showed that bacterial community structures were significantly distinct among water, sediment, and intestine; meanwhile, the relative abundances of intestinal dominant taxa were significantly changed between different rearing stages. Compared with intestine and water, shrimp intestine and sediment had a similar profile of the dominant bacterial genera by cluster analysis, and the observed species, diversity indexes, and shared OTUs of bacterial communities in intestine and sediment were simultaneously increased after shrimp were farmed for 90 days. These results reflected a closer relationship between microbiotas in sediment and intestine, which was further proved by nonmetric multidimensional scaling analysis. However, bacterial communities in water, sediment, and intestine responded differently to environmental variables by redundancy and correlation analysis. More importantly, shrimp physiological parameters were closely associated with bacterial variations in the gut and/or ambient, especially the gut microbiota owning significantly high levels of predicted functional pathways involved in disease emergence. These findings may greatly add to our understanding of the microbiota characteristics of the shrimp pond ecosystem and the complex interactions among shrimp, ambient microflora, and environmental variables.