3.1 Bacterial microbiome: community analysis
16S rRNA amplification was successfully carried out for 12/22 VKC samples (54.5%), and in 4/20 HC samples (20%). High-throughput amplicon sequencing produced a total of 734.157 high-quality reads (average of 45.885 reads per sample), which were clustered into 1.241 OTUs (97% sequence identity) and classified according to the Greengenes database. Rarefaction curves tended to saturation indicating that that the sequencing depth was adequate to capture the diversity in the microbiome in each conjunctival sample (Figure 1S).
Overall, α diversity was significantly higher in VKC patients compared to HC (p=0.05) (Figure 2S). Alpha diversity was significantly higher in IgE-negative patients than HC (p=0.03) as well as in tarsal VKC patients compared to the mixed type (p=0.02) and HC (p=0.03). Alpha diversity was also significantly higher in formula-fed than breastfed children (p=0.03) and in children with a history of atopic dermatitis (AD) during the first year of life (p=0.01) (Figure 1), but not significant considering the type of birth, socioeconomical condition, diet, or contact with pets.
Beta diversity metrics highlighted the differences of conjunctival microbiota composition in VKC patients and HC (Figure 3S). PCoA unveiled also the different clustering of bacterial microbiomes depending on different variables such as the presence of atopic dermatitis during the first year of life, the type of feeding and the severity of ocular surface inflammation according to the QUICK score (Figure 2).