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Background: Breast milk is a rich nutrient with a temporally dynamic nature. In particular, numerous alterations in the nutritional, immunological and microbiological content occur during the transition from colostrum to mature milk. The objective of our study was to evaluate the potential impact of delivery mode on the microbiota of colostrum, at both the quantitative and qualitative levels (bacterial abundance and microbiota network). Methods: Twenty-nine Italian mothers (15 vaginal deliveries vs 14 Cesarean sections) were enrolled in the study. The microbiota of colostrum samples was analyzed by next generation sequencing (Ion Torrent Personal Genome Machine). The colostrum microbiota network associated with Cesarean section and vaginal delivery was evaluated by means of the Auto Contractive Map (AutoCM), a mathematical methodology based on Artificial Neural Network (ANN) architecture. Results: Numerous differences between Cesarean section and vaginal delivery colostrum were observed. Vaginal delivery colostrum had a significant lower abundance of Pseudomonas spp., Staphylococcus spp. and Prevotella spp. when compared to Cesarean section colostrum samples. Furthermore, the mode of delivery had a strong influence on the microbiota network, as Cesarean section colostrum showed a higher number of bacterial hubs if compared to vaginal delivery, sharing only 5 hubs. Interestingly, the colostrum of mothers who had a Cesarean section was richer in environmental bacteria than mothers who underwent vaginal delivery. Finally, both Cesarean section and vaginal delivery colostrum contained a greater number of anaerobic bacteria genera. Conclusions: The mode of delivery had a large impact on the microbiota composition of colostrum. Further studies are needed to better define the meaning of the differences we observed between Cesarean section and vaginal delivery colostrum microbiota. [ABSTRACT FROM AUTHOR]