Analysis of 16S rRNA Gene Sequence of Nasopharyngeal Exudate Reveals Changes in Key Microbial Communities Associated with Aging

Abstract

Functional or compositional perturbations of the microbiome can occur at different sites, of the body and this dysbiosis has been linked to various diseases. Changes in the nasopharyngeal microbiome are associated to patient’s susceptibility to multiple viral infections, supporting the idea that the nasopharynx may be playing an important role in health and disease. Most studies on the nasopharyngeal microbiome have focused on a specific period in the lifespan, such as infancy or the old age, or have other limitations such as low sample size. Therefore, detailed studies analyzing the age- and sex-associated changes in the nasopharyngeal microbiome of healthy people across their whole life are essential to understand the relevance of the nasopharynx in the pathogenesis of multiple diseases, particularly viral infections. One hundred twenty nasopharyngeal samples from healthy subjects of all ages and both sexes were analyzed by 16S rRNA sequencing. Nasopharyngeal bacterial alpha diversity did not vary in any case between age or sex groups. Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes were the predominant phyla in all the age groups, with several sex-associated. Acinetobacter, Brevundimonas, Dolosigranulum, Finegoldia, Haemophilus, Leptotrichia, Moraxella, Peptoniphilus, Pseudomonas, Rothia, and Staphylococcus were the only 11 bacterial genera that presented significant age-associated differences. Other bacterial genera such as Anaerococcus, Burkholderia, Campylobacter, Delftia, Prevotella, Neisseria, Propionibacterium, Streptococcus, Ralstonia, Sphingomonas, and Corynebacterium appeared in the population with a very high frequency, suggesting that their presence might be biologically relevant. Therefore, in contrast to other anatomical areas such as the gut, bacterial diversity in the nasopharynx of healthy subjects remains stable and resistant to perturbations throughout the whole life and in both sexes. Age-associated abundance changes were observed at phylum, family, and genus levels, as well as several sex-associated changes probably due to the different levels of sex hormones present in both sexes at certain ages. Our results provide a complete and valuable dataset that will be useful for future research aiming for studying the relationship between changes in the nasopharyngeal microbiome and susceptibility to or severity of multiple diseases.

Age- and disease severity-associated changes in the nasopharyngeal microbiota of COVID-19 patients

“…was not significantly different in any case when comparing between age groups in the uninfected cohort used as a control in this study (18). Similarly, analysing bacterial species richness in the nasopharynx of subjects of the different age and COVID-19 disease severity groups established for this study, revealed no age-or severity-associated relevant differences in alpha diversity, with the only striking exception of COVID-19 patients with severe symptoms, whose alpha diversity decreased progressively as their age increased.…”

“…3c). In addition, we have previously demonstrated the absence of any significant changes in alpha diversity in the uninfected control cohort (18).…”

“…After characterizing the healthy human nasopharyngeal microbiota (18), we sought to study any possible links between alterations of this microbiota and SARS-CoV-2 infection, with special attention to the COVID-19 clinical outcome, as this is a crucial question from a biomedical and clinical point of view that remains unresolved. But the problem is not the lack of studies on this, since dozens of works have already addressed this topic, but rather that their results are extremely variable and contradictory (14).…”

“…The uninfected nasopharyngeal control samples were selected, collected, and classified as previously described (18). Besides those uninfected samples, and according to our experimental design and economic resources, we decided to select and collect a maximum of 360 nasopharyngeal samples from SARS-CoV-2 infected individuals with different ages and COVID-4 19 disease severity, as will be detailed here after (Table S1).…”

“…Available at: https://www.random.org]. The reason that some groups contained fewer patients than planned in our experimental design, or even none such as patients under 20 years of age with moderate or severe COVID-19 disease severity, is that patients with these characteristics are rare and we simply enrolled all patients with such characteristics who were in our parent cohort (Table S1 Finally, while the exclusion criteria for the uninfected nasopharyngeal samples were already described (18), the only SARS-CoV-2 infected individuals excluded from this study were those younger than 1 year as the microbiome of infants is known to be highly fluctuating with age and, therefore, it could significantly increase the variability of our analyses.…”