**1. Background**

Among the remaining challenges in biomedical sciences, one of the most important is to fully understand the effect of aging on human biological processes, health, and wellness. In recent years, solid evidence has been collected to support the idea that the microbial communities that inhabit the different anatomical areas of the human body could play a key role in these processes, and there has been much speculation about possible medical interventions [1–6]. Although most research has focused on the well-studied gut microbiome, there is growing evidence that variations in microbial communities in other sites of the body are also responsible for wide-ranging health effects [7–11]. The case of the respiratory tract is curious, since the lungs were long believed to be sterile, despite the fact that they are constantly exposed to microorganisms in inhaled air and the upper respiratory tract [12], which has been the main cause that the respiratory microbiome has barely been studied until very recently [13]. However, new culture-independent microbial identification techniques, such as metagenomics, have revealed that the respiratory tract is a dynamic ecosystem, and this has raised the interest of the scientific community in the role of the respiratory microbiota in health and disease [12,13].

The human upper respiratory tract that comprises the anterior nares, nasal cavity, sinuses, nasopharynx, Eustachian tube, middle ear cavity, oral cavity, oropharynx, and larynx is the major portal of entry for infectious droplet- or aerosol-transmitted microorganisms [14]. Among these different areas, the nasopharynx is anatomically unique because it presents a common meeting place for the ear, nose, and mouth cavities [15], but has not gained special prominence until the outbreak of the current COVID-19 pandemic [16]. Importantly, dozens of studies have already detected unquestionable correlations between the composition of the nasopharyngeal microbiota and susceptibility to different viral infections in humans [13], and some evidence is emerging, although still controversial, that it may be playing a role in the susceptibility to SARS-CoV-2 infection, too [17]. Elucidating this might shed light on the still unexplained fact that some COVID-19 patients, such as the elderly, are more susceptible and present more severe forms of COVID-19 than others [18].

Large cohort studies of human microbiome data with appropriate controls are particularly valuable, especially of all ages and both sexes, as these datasets are difficult to obtain due to multiple factors, including our long lifespans, heterogeneity in consent and other sample access issues, and because of socioeconomic confounds. Therefore, human studies have tended to focus on a specific component of the lifespan, such as infanthood, or studies of the elderly, rather than examining variation across an entire population. Knowledge about the relationships between changes in the nasopharyngeal microbiota and susceptibility to viral infections is a good example of this, since most studies have focused only on children [13].

A crucial factor to consider in studies of variation across the lifespan is sex as a biological variable. For aging research, this includes the understanding that females and males may have different aging trajectories [19–21], including in key systems such as the digestive tract. For example, the gut microbiome and sex hormones may interact to predispose women to autoimmune diseases [22] and dietary interventions are known to have sex-specific effects on gut microbiota [23]. There are no studies analyzing the possible sex-associated differences in the nasopharyngeal microbiome at different life stages.

Here, we analyze, for the first time, the diversity and relative abundance of the nasopharyngeal microbiota across the whole lifespan in 120 healthy individuals of all ages and both sexes, the taxonomic changes in the nasopharynx associated to age or sex, and the possible biological relevance of several taxa whose frequency of appearance in the population is high. We therefore provide a very comprehensive and valuable dataset that will be the base for future research aimed at identifying relationships between age- and sex-associated changes in nasopharyngeal microbiome and susceptibility to or severity of the diseases of interest.
