Dynamic distribution of gut microbiota during embryonic development in chicken

Abstract

The gut microbiota is a complex ecological community and widely recognized in many aspects of research, but little is known on the relation between gut microbiota and embryonic development in chickens. The aim of this study was to explore the dynamic distribution of gut microbiota in chickens' embryos during stages of developments (chicken embryos that had incubated until day 3 [E3], day 12 [E12], and day 19 [E19]). Here, 16S rRNA gene sequencing was performed on the gut microbiota in chicken embryos across different developmental stages. Twenty-one phyla and 601 genera were present in chicken embryos, and 96 genera such as Ochrobactrum, Phyllobacterium, and Amycolatopsis were the core microbiota in the 3 stages of development. Second, 94 genera of microbes were found to change significantly between E3 and E12, and 143 genera significantly differed between E12 and E19 in chicken embryos (P < 0.05). Ochrobactrum and Amycolatopsis decreased with growth changes: E3 (30.4%), E12 (25.1%), and E19 (13.6%) and E3 (11.5%), E12 (7.4%), and E19 (5.6%), respectively. Contrarily, Phyllobacterium increased to 47.9% at E19, indicating the growing trend of microbial diversity among the embryos' development. Moreover, the principal component analysis showed a high level of similarities between E3 and E12 compared with E19, whereas the alpha analysis showed more diversity of gut microbiota at E19. Furthermore, the functional predictions showed that metabolic pathways such as energy metabolism and genetic information processing were significantly enriched on day 3 and day 12 in our study, suggesting their strong influence on growth, development, and immunity of chicken embryos. Our findings provide insights into the understanding of dynamic shifts of gut microbiota during chicken embryonic growth.

Keywords: 16S rRNA gene sequencing; chicken; embryo development; gut microbiota.

Figure 1

Composition of gut microbiota in the chicken embryo. (A) Phylogenetic tree constructed from the taxa. Each classification unit is distinguished with different colors. Colored blocks in the outermost circle indicate phyla and in the inner circle indicate genera. (B) The abundant microbes at each level (genus, family, order, class, and phylum).

Figure 2

Embryonic gut microbial difference at different developmental stages. (A) PCA shows a high level of similarities between chicken embryos that had incubated until day 3 and 12 compared with those incubated until day 19. (B) Alpha analysis using the Shannon index showing the diversity of gut microbiota across different growth stages. (C) The relative abundance of gut microbiota in the chicken embryo at different days. (D) The Venn diagram shows the core microbes shared at different stages of development. (E) Taxonomic cladogram generated from LEfSe showing significant difference in the microbiota profile of 3 stages of development. Green, red, and blue represent the enriched taxa in chicken embryos that had been incubated until day 3, 12, and 19, respectively. Abbreviations: LEfSe, linear discriminant analysis effect size; PCA, principal component analysis.

Figure 3

Functional profiles of the microbial community at different developmental stages (A) Functional pathways that changed significantly (P < 0.05) between E3 and E12. (B) Functional pathways that changed significantly (P < 0.05) between E12 and E19.

Figure 4

Embryonic gut microbiota composition across different breeds: Beijing You (Y), Shiqiza (S), and SPF (Babcock) (B). (A) Beta analysis using NMDS across different breeds. Blue, red, and green represent SPF (Babcock) (B), Shiqiza (S), and Beijing You (Y) breeds, respectively. (B) The Shannon diversity index was not significant among the 3 groups (P > 0.05). The box shows the quartiles above and below the median, with a dark line at the center of the box denoting the median and black dots outside the box showing the outlier. (C) Relative abundance of bacteria among the 3 breeds at the genus level. (d) Relative abundance of bacteria among the 3 breeds at the phylum level. Abbreviations: NMDS, nonmetric multidimensional scaling; SPF, specific pathogen-free.