Low-dose (LD, 100 mg kg(-1) day(-1)), moderate-dose (MD, 200 mg kg(-1) day(-1)), and high-dose (HD, 600 mg kg(-1) day(-1)) krill oil treatments have a stepwise, enhanced effect on alleviating hyperlipidemia, and 16S rRNA sequencing of the fecal samples demonstrates that krill oil treatment alters microbial communities. Feces may not represent all microbial communities in the gastrointestinal (GI) tract. Therefore, in this study, the stored ileal and colon samples collected from LD and HD groups were sequenced, and the location-specific modulations of microbial communities were observed after krill oil treatments. The 16S rRNA sequencing of the ileal samples showed that the LD and HD groups have similar patterns between control and high-fat diet (HFD) treatments, and six most abundant genera and 40 operational taxonomic units that respond to krill oil treatment were identified. However, the 16S rRNA sequencing of the colon samples showed that LD krill oil shifts the structure from the HFD to that of the control, whereas the HD group was distributed between the control and HFD groups. The corresponding most abundant genera and responsive OTUs totaled 4 and 45, respectively. In conclusion, different gastrointestinal tract locations contain different microbial communities. These results will help to provide a comprehensive understanding of the role of dietary krill oil in modulating the gut microbiota and alleviating hyperlipidemia.