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).