During early ontogeny, microbiome affects development of the gastrointestinal tract, immunity and survival in vertebrates. Bird eggs are thought to be (1) initially sterile (sterile egg hypothesis) and (2) colonised after oviposition through horizontal trans-shell migration, or (3) initially seeded with bacteria by vertical transfer from mother oviduct. To date, however, little empirical data illuminate the contribution of these mechanisms to gut microbiota formation in avian embryos. We investigated microbiome of the egg content (day 0; E0-egg), embryonic gut at day 13 (E13) and female faeces in a free-living passerine, the great tit (Parus major), using a methodologically advanced procedure combining 16S rRNA gene sequencing and microbe-specific qPCR assays. Our metabarcoding revealed that the avian egg is (nearly) sterile, yet obtaining a slightly richer microbiome during the embryonic development. Of the three potentially pathogenic bacteria targeted by qPCR, only Dietzia was found in E0-egg (yet also in controls), E13 gut and female samples, which might indicate possible vertical transfer. Unlike in poultry, we have shown in passerines that major bacterial colonisation of the gut does not occur before hatching. We emphasise that protocols that carefully check for environmental contamination are critical in studies with low-bacterial biomass samples.

中文翻译：

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雀形目鸟类中几乎（？）不育的禽蛋

在个体发育早期，微生物组影响脊椎动物胃肠道的发育、免疫力和生存。鸟蛋被认为是（1）最初是无菌的（无菌蛋假说）和（2）产卵后通过水平跨壳迁移定植，或（3）最初通过从母输卵管垂直转移接种细菌。然而，迄今为止，很少有经验数据阐明这些机制对禽类胚胎肠道微生物群形成的贡献。我们使用结合 16S rRNA 的方法先进程序，研究了自由生活的雀形目大山雀 (Parus Major) 中的卵含量（第 0 天；E0-卵）、第 13 天的胚胎肠道 (E13) 和雌性粪便的微生物组基因测序和微生物特异性 qPCR 检测。我们的元条形码显示，禽蛋（几乎）是无菌的，但在胚胎发育过程中获得了稍微丰富的微生物组。在 qPCR 靶向的三种潜在致病菌中，只有迪茨氏菌在 E0 卵（但也在对照中）、E13 肠道和雌性样本中发现，这可能表明可能存在垂直转移。与家禽不同，我们在雀形目动物中发现，孵化前不会发生肠道内主要细菌的定植。我们强调，仔细检查环境污染的方案对于低细菌生物量样品的研究至关重要。