Table 1.

Sterility of the gastrointestinal tract in neonatal animals

Model	Method	Key findings	Controversies	Study
Cattle	16S rRNA gene sequencing	Microbiota, dominated by Proteobacteria and Firmicutes, were detected in bovine fetal tissues and fluids at 12 weeks gestation	The findings challenge the sterile womb hypothesis but face scrutiny regarding contamination risks and the biological implications of detected low-biomass microbiomes	Amat et al. [17]
Lamb	16S rRNA gene sequencing	The study found no evidence of bacterial DNA in the fetal environment or intestine during the third trimester of sheep pregnancy, reinforcing the concept of a sterile fetal environment	The findings challenge recent claims of in utero microbial colonization, sparking debates over the reliability of low-biomass DNA sequencing and the influence of contamination on reported microbiomes	Malmuthuge and Griebel [18]
Lamb	Multi-omics analysis	Detected Escherichia coli as predominant species in fetal lamb intestines, suggesting maternal microbial transfer	Data might be questioned due to contamination or errors in low-biomass sample analysis	Bi et al. [19]
Calves	16S rRNA gene sequencing	Fetal calf GIT and amniotic fluid harbor distinct, viable microbial communities, with colonization starting as early as 5 months of gestation	The study's findings contradict the sterile womb hypothesis and face skepticism regarding contamination control	Guzman et al. [20]
Sheep	16S rRNA gene sequencing	With strict contamination controls, this study identified trace bacterial DNA in fetal tissues, supporting the potential for maternal–fetal transfer of bacteria or their genetic material	The detection of low-biomass bacterial DNA raises concerns about contamination, and the functional significance of this DNA in fetal development remains uncertain	Rodriguez et al. [21]