Avian coccidiosis is an intestinal parasitic disease introduced by Eimeria spp., causing a major economic loss in the world poultry industry. Eimeria necatrix is the most pathogenic species that causes acute coccidiosis in chickens, leading to high mortality. Studies have shown that disruption of the gut environment due to Eimeria infection causes an imbalance in intestinal homeostasis. However, changes in the intestinal microbiota of chickens infected with E. necatrix remain unclear. In the present study, we performed full-length 16S ribosomal RNA amplicon sequencing to assess the effects of E. necatrix infection on jejunal and cecal microbiota at 4 and 10 days post-infection (dpi). The results showed that in both the infected and not infected groups at both time points, the most abundant phyla were Firmicutes, Proteobacteria and Bacteroidetes in the jejunum, and Firmicutes, Bacteroidetes and Proteobacteria in the cecum. The most common genera in the jejunum were Lactobacillus, Limosilactobacillus and Ligilactobacillus at 4 dpi, and Lactobacillus, Limosilactobacillus and Enterococcus in the infected group, and Lactobacillus, Limosilactobacillus and Streptococcus in the control group at 10 dpi. In the cecum, the most common genera were Phocaeicola, Lactobacillus and Alistipes at 4 dpi, and Lactobacillus, Phocaeicola and Alistipes in the infected group, and Lactobacillus, Phocaeicola and Bacteroides in the control group at 10 dpi. A total of 1528 species was annotated, and differences in relative abundance at the species level were analyzed using Lefse method. The results showed that the relative abundance of 23 species, including Acetilactobacillus jinshanensis, Anaerotruncus colihominis, Bacteroides heparinolyticus, Bacteroides ndongoniae, Bariatricus comes, Bifidobacterium gallinarum, Blautia coccoides, Butyricimonas paravirosa, Caproiciproducens galactitolivorans, Clostridioides difficile, Enterococcus cecorum, Escherichia coli, Intestinimonas timonensis, Lachnoanaerobaculum umeaense, Lactobacillus acetotolerans, Ligilactobacillus aviarius, Ligilactobacillus aviarius _B, Limosilactobacillus oris, Limosilactobacillus vaginalis, Megamonas funiformis, Plesiomonas shigelloides, Streptococcus pneumoniae, and Veillonella denticariosi, were significantly different between the infected and not infected groups. Our data reveal that E. necatrix infenction disrupts the integrity of gut microbiota, potentially promoting the establishment and growth of pathogenic bacteria; some species such as Bariatricus comes and Ligilactobacillus aviarius_B may be associated with the pathogenicity of the coccidian parasite and recovery of coccidiosis.