Table_3_Incubation determines favorable microbial communities in Chinese alligator nests.DOCX

Nest materials are a major heat source due to rotting promoted by microbial activity. Additionally, they are a potential microbial source given their direct contact with eggshells. Microbial dynamics during incubation have been studied in wild birds; however, similar studies in reptiles remain elusive. Here, the study characterized microbial communities in the nest materials of Chinese alligator (Alligator sinensis) using high-throughput sequencing of bacterial 16S rRNA genes and fungal internal transcribed spacer (ITS) region sequences. The results showed that significant changes in the diversity and structure of microbial communities according to different incubation periods. The diversity and richness of bacterial species increased significantly over time, but the relative abundance of the most dominant bacteria in pre-incubation period, including some pathogenic bacteria, declined after incubation. In contrast, fungal species diversity and richness decreased significantly with time. Additionally, nest material composition significantly influenced microbial community structure rather than species diversity and richness. Notably, the fungal community structure showed a stronger response than bacteria to nest material composition, which varied due to differences in plant litter composition. Our results demonstrate the significant response of microbial community diversity and structure to differences in incubation periods and nest material composition in reptiles. It is further emphasized that the importance of incubation period in the conservation of the Chinese alligator and could inform similar studies in other reptiles and birds.

巢材因微生物活动促进腐烂而成为主要热源，同时因其直接接触蛋壳，也成为潜在的微生物来源。目前针对野生鸟类孵化期的微生物动态已开展较多研究，但爬行动物中的同类研究仍较为匮乏。本研究通过对细菌16S rRNA基因与真菌内转录间隔区（ITS）序列进行高通量测序，解析了扬子鳄（Alligator sinensis）巢材中的微生物群落特征。结果显示，微生物群落的多样性与结构随孵化期的不同呈现显著变化：细菌物种的多样性与丰富度随孵化时间显著升高，但孵化前期占优势的细菌类群（含部分致病菌）的相对丰度在孵化后有所下降；与之相反，真菌物种的多样性与丰富度则随孵化时间显著降低。此外，巢材组成显著影响微生物群落结构，但对物种多样性与丰富度无显著作用；值得注意的是，相较于细菌群落，真菌群落结构对因植物凋落物组成差异而形成的巢材组成变化响应更为强烈。本研究结果证实，爬行动物巢材中的微生物群落多样性与结构对孵化期差异及巢材组成变化具有显著响应，进一步凸显了孵化期在扬子鳄保护中的重要意义，同时可为其他爬行动物与鸟类的同类研究提供参考。