Background Molecular assays are important tools for pathogen detection but need to be periodically re-evaluated with the discovery of additional genetic diversity that may cause assays to exclude target taxa or include non-target taxa. A single well-developed assay can find broad application across research, clinical, and industrial settings. Pathogen prevalence within a population is estimated using such assays and accurate results are critical for formulating effective public health policies and guiding future research. A variety of assays for the detection of Staphylococcus aureus are currently available. The utility of commercial assays for research is limited, given proprietary signatures and lack of transparent validation. Results In silico testing of existing peer-reviewed assays show that most suffer from a lack of sensitivity and specificity. We found no assays that were specifically designed and validated for quantitative use. Here we present a qPCR assay, SaQuant, for the detection and quantification of S. aureus as might be collected on sampling swabs. Sensitivity and specificity of the assay was 95.6 and 99.9 %, respectively, with a limit of detection of between 3 and 5 genome equivalents and a limit of quantification of 8.27 genome equivalents. The presence of DNA from non-target species likely to be found in a swab sample, did not impact qualitative or quantitative abilities of the assay. Conclusions This assay has the potential to serve as a valuable tool for the accurate detection and quantification of S. aureus collected from human body sites in order to better understand the dynamics of prevalence and transmission in community settings.

Disease control relies on pathogen identification and understanding reservoirs. Staphylococcus aureus infection prevention is based upon decades of research on colonization and infection, but diminishing returns from mitigation efforts suggest significant knowledge gaps. Existing knowledge and mitigation protocols are founded upon culture-based detection, with almost no information about pathogen quantities. We employed a qPCR assay on samples from three body sites to characterize colonization more comprehensively than previous studies by describing both prevalence and pathogen quantity. We show a much higher overall prevalence (65.9%) than previously documented, with higher quantities and prevalence associated with the nares, non-Hispanic males (86.9%), and correlating with colonization in other body sites. These results suggest that research and clinical practices likely misclassify over half of colonized persons, limiting mitigation measures and their impact. This work begins the process of rebuilding foundational knowledge of S. aureus carriage with more accurate and wholistic approaches.

Staphylococcus aureus exists as a pathogen and commensal. Individuals with asymptomatic carriage serve as a reservoir for transmission and are at increased risk of infecting themselves. In order to characterize the genomic diversity of S. aureus circulating in the community, we sequenced 166 genomes collected from individuals in Yuma, AZ.

Asymptomatic carriage of Staphylococcus aureus is a major risk factor for subsequent clinical infection. Diminishing returns from mitigation efforts emphasize the need to better understand colonization, spread, and transmission of this opportunistic pathogen. While contact with other people presents opportunities for pathogen exposure and transmission, diversity of social connections may be protective against pathogens such as the common cold. This study examined whether social relationship resources, including the amount and diversity of social contacts, are associated with S. aureus colonization. Participants were community members (N = 443; 68% Hispanic) in naturally occurring social groups in southwestern Arizona. Four types of social relationships and loneliness were assessed, and samples from the skin, nose and throat were obtained to ascertain S. aureus colonization. Overall S. aureus prevalence was 64.8%. Neither the amount nor the diversity of social contacts were associated with S. aureus colonization. The concurrent validity of the social relationship assessments was supported by their moderate intercorrelations and by their positive association with self-rated health. The results suggest that the association of social network diversity and susceptibility to the common cold does not extend to S. aureus colonization. Conversely, colonization prevalence was not higher among those with more social contacts. The latter pattern suggests that social transmission may be relatively infrequent or that more intimate forms of social interaction may drive transmission and colonization resulting in high community prevalence of S. aureus colonization. These data inform communicable disease control efforts.

Staphylococcus aureus is a frequent cause of mild and severe infections that occur when these commensal bacteria penetrate the outer layers of skin or mucosa. As most S. aureus infections are the result of autoinfection, and community-acquired infections are increasingly common, it is important to better understand S. aureus colonization characteristics in the community setting. Using standard culture technique and a quantitative PCR assay (SaQuant), we detected and quantified S. aureus across the nares, throat, and palm of 548 community-dwelling individuals in southwestern Arizona. Using culture-based methods, we detected S. aureus colonization in the nares of 26.3% of individuals (n = 144); however, the combination of two detection methods across multiple body sites resulted in much higher prevalence than has been reported previously. Overall, 65.9% of participants were colonized, with significantly higher prevalence in males (compared to females) and non-Hispanics (compared to Hispanics), with this pattern especially evident in nares and throat samples. Colonizing quantities in the nares were slightly higher in males and significantly greater among non-Hispanics. The clear sex and ethnicity patterns warrant further investigation in order to identify and leverage protective factors that may drive these disparities. In the nares, S. aureus density was the highest, most variable, and correlates with colonization in other body sites such as throat and palm. Our results demonstrate that screening by culture-based methods only can miss individuals colonized by S. aureus and that previous carriage statistics are likely underestimates. By including a highly sensitive quantitative assay, this work provides a roadmap towards more comprehensive and accurate characterization of S. aureus carriage and the potential for more effective mitigation.AUTHOR SUMMARYEffective disease control and prevention is tied to pathogen identification and understanding reservoirs. Staphylococcus aureus infection prevention efforts and protocols are based upon decades of research on colonization patterns and associated links to subsequent infection. Unfortunately, efforts to prevent S. aureus infections have been met with diminishing returns, suggesting significant gaps in fundamental knowledge of colonization. However, this knowledge and resulting protocols, are founded upon culture-based detection. By employing a new quantitative PCR assay on samples from three body sites in 548 individuals, we can characterize colonization more comprehensively than previous studies by describing both prevalence and pathogen quantity. Our highly sensitive detection resulted in an overall prevalence of 65.9%. Higher quantities were associated with the nares and were highest among non-Hispanic males (86.9%). Overall prevalence was much higher than has been previously documented. Common research practices, such as culture-based detection from a single body site, may misclassify over half of colonized persons. Future studies incorporating quantitative data, especially with longitudinal sampling at more body sites will provide a more wholistic understanding of community carriage, colonization dynamics, and likelihood of autoinfection and transmission.