Egg production, one of the most important economic traits of chickens, is largely regulated by interactions of
genetic, endocrine, and environmental factors. Gut microbiota is the main environmental factor that is closely
related to egg production. However, the key gut microbiota affecting egg production is still unknown. Therefore,
Golden Montazah (GM) chickens (Rhode Island Red × Dokki-4 chickens) of 44 weeks of age were housed
separately. After keeping track of egg laying for 90 consecutive days, 10 laying hens were selected for the H
group (with a higher egg laying rate) based on the laying level, and 10 laying hens were assigned to the L group
(with a lower egg laying rate). Grossly, the number of eggs (P < 0.0001) and the number of hierarchical follicles
(P < 0.001) were significantly higher in the ovaries of high egg production chickens. HE staining results indicated
that the granulosa cell thickness of large white follicles (LWFs) was significantly higher (P < 0.001) in high
egg production chickens. 16S rRNA gene sequencing results showed that the relative abundance of Firmicutes
was higher both in the ileum and cecum of high egg production chickens, yet the relative abundance of Proteobacteria was higher in the ileum of low egg production chickens. Further, Spearman correlation analysis
indicated that the relative abundance of cecal Lachnoclostridium was positively correlated with the egg number
and hierarchical follicle number, while the relative abundance of ileal Olsenella was significantly (P < 0.05) and
positively correlated with the egg number. Yet the relative abundance of cecal Collinsella was significantly (P <
0.05) and negatively correlated with the egg number. Our findings indicated that gut microbiota is associated
with the egg-laying performance of chickens.