Chromosome-Scale, De Novo, Phased Genome Assemblies of Three Australian Limes: Citrus australasica, C. inodora, and C. glauca.

Huanglongbing (HLB) is a severe citrus disease worldwide. Wild Australian limes like Citrus australasica, C. inodora, and C. glauca possess beneficial HLB resistance traits. Individual trees of the three taxa were extensively used in a breeding program for over a decade to introgress resistance traits into commercial-quality citrus germplasm. We generated high-quality, phased, de novo genome assemblies of the three Australian limes using PacBio long-read sequencing. The genome assembly sizes of the primary and alternate haplotypes were determined for C. australasica (337 Mb/335 Mb), C. inodora (304 Mb/299 Mb), and C. glauca (376 Mb/379 Mb). The nine chromosome-scale scaffolds included 86-91% of the genome sequences generated. The integrity and completeness of the assembled genomes were estimated to be at 97.2-98.8%. Gene annotation studies identified 25,461 genes in C. australasica, 27,665 in C. inodora, and 30,067 in C. glauca. Genes belonging to 118 orthogroups were specific to Australian lime genomes compared to other citrus genomes analyzed. Significantly fewer canonical resistance (R) genes were found in C. inodora and C. glauca (319 and 449, respectively) compared to C. australasica (576), C. clementina (579), and C. sinensis (651). Similar patterns were observed for other gene families associated with potential HLB resistance, including Phloem protein 2 (PP2) and Callose synthase (CalS) genes predicted in the Australian lime genomes. The genomic information on Australian limes developed in the present study will help understand the genetic basis of HLB resistance.

Long-read, chromosome-scale assembly of Vitis rotundifolia cv. Carlos and its unique resistance to Xylella fastidiosa subsp. fastidiosa.

BACKGROUND: Muscadine grape (Vitis rotundifolia) is resistant to many of the pathogens that negatively impact the production of common grape (V. vinifera), including the bacterial pathogen Xylella fastidiosa subsp. fastidiosa (Xfsf), which causes Pierce's Disease (PD). Previous studies in common grape have indicated Xfsf delays host immune response with a complex O-chain antigen produced by the wzy gene. Muscadine cultivars range from tolerant to completely resistant to Xfsf, but the mechanism is unknown. RESULTS: We assembled and annotated a new, long-read genome assembly for 'Carlos', a cultivar of muscadine that exhibits tolerance, to build upon the existing genetic resources available for muscadine. We used these resources to construct an initial pan-genome for three cultivars of muscadine and one cultivar of common grape. This pan-genome contains a total of 34,970 synteny-constrained entries containing genes of similar structure. Comparison of resistance gene content between the 'Carlos' and common grape genomes indicates an expansion of resistance (R) genes in 'Carlos.' We further identified genes involved in Xfsf response by transcriptome sequencing 'Carlos' plants inoculated with Xfsf. We observed 234 differentially expressed genes with functions related to lipid catabolism, oxidation-reduction signaling, and abscisic acid (ABA) signaling as well as seven R genes. Leveraging public data from previous experiments of common grape inoculated with Xfsf, we determined that most differentially expressed genes in the muscadine response were not found in common grape, and three of the R genes identified as differentially expressed in muscadine do not have an ortholog in the common grape genome. CONCLUSIONS: Our results support the utility of a pan-genome approach to identify candidate genes for traits of interest, particularly disease resistance to Xfsf, within and between muscadine and common grape.

Paleoecology provides context for conserving culturally and ecologically important pine forest and barrens communities.

In fire-prone ecosystems, knowledge of vegetation-fire-climate relationships and the history of fire suppression and Indigenous cultural burning can inform discussions of how to use fire as a management tool, particularly as climate continues to change rapidly. On Wiisaakodewan-minis/Stockton Island in the Apostle Islands National Lakeshore of Wisconsin, USA, structural changes in a pine-dominated natural area containing a globally rare barrens community occurred after the cessation of cultural burning by the Indigenous Ojibwe people and the imposition of fire-suppression policies, leading to questions about the historical role of fire in this culturally and ecologically important area. To help understand better the ecological context needed to steward these pine forest and barrens communities, we developed palaeoecological records of vegetation, fire, and hydrological change using pollen, charcoal, and testate amoebae preserved in peat and sediment cores collected from bog and lagoon sediments within the pine-dominated landscape. Results indicated that fire has been an integral part of Stockton Island ecology for at least 6000 years. Logging in the early 1900s led to persistent changes in island vegetation, and post-logging fires of the 1920s and 1930s were anomalous in the context of the past millennium, likely reflecting more severe and/or extensive burning than in the past. Before that, the composition and structure of pine forest and barrens had changed little, perhaps due to regular low-severity surface fires, which may have occurred with a frequency consistent with Indigenous oral histories (~4-8 years). Higher severity fire episodes, indicated by large charcoal peaks above background levels in the records, occurred predominantly during droughts, suggesting that more frequent or more intense droughts in the future may increase fire frequency and severity. The persistence of pine forest and barrens vegetation through past periods of climatic change indicates considerable ecological resistance and resilience. Future persistence in the face of climate changes outside this historical range of variability may depend in part on returning fire to these systems.

Chemical profile and analysis of biosynthetic pathways and genes of volatile terpenes in Pityopsis ruthii, a rare and endangered flowering plant.

It is critical to gather biological information about rare and endangered plants to incorporate into conservation efforts. The secondary metabolism of Pityopsis ruthii, an endangered flowering plant that only occurs along limited sections of two rivers (Ocoee and Hiwassee) in Tennessee, USA was studied. Our long-term goal is to understand the mechanisms behind P. ruthii's adaptation to restricted areas in Tennessee. Here, we profiled the secondary metabolites, specifically in flowers, with a focus on terpenes, aiming to uncover the genomic and molecular basis of terpene biosynthesis in P. ruthii flowers using transcriptomic and biochemical approaches. By comparative profiling of the nonpolar portion of metabolites from various tissues, P. ruthii flowers were rich in terpenes, which included 4 monoterpenes and 10 sesquiterpenes. These terpenes were emitted from flowers as volatiles with monoterpenes and sesquiterpenes accounting for almost 68% and 32% of total emission of terpenes, respectively. These findings suggested that floral terpenes play important roles for the biology and adaptation of P. ruthii to its limited range. To investigate the biosynthesis of floral terpenes, transcriptome data for flowers were produced and analyzed. Genes involved in the terpene biosynthetic pathway were identified and their relative expressions determined. Using this approach, 67 putative terpene synthase (TPS) contigs were detected. TPSs in general are critical for terpene biosynthesis. Seven full-length TPS genes encoding putative monoterpene and sesquiterpene synthases were cloned and functionally characterized. Three catalyzed the biosynthesis of sesquiterpenes and four catalyzed the biosynthesis of monoterpenes. In conclusion, P. ruthii plants employ multiple TPS genes for the biosynthesis of a mixture of floral monoterpenes and sesquiterpenes, which probably play roles in chemical defense and attracting insect pollinators alike.

PanoMRT: Panoramic infrared thermography to model human thermal exposure and comfort.

As summer heat waves become the new normal worldwide, modeling human thermal exposure and comfort to assess and mitigate urban overheating is crucial to uphold livability in cities. We introduce PanoMRT, an open source human-biometeorological model to calculate Mean Radiant Temperature (TMRT), Physiologically Equivalent Temperature (PET), and the Universal Thermal Climate Index (UTCI) from thermal equirectangular 360° panoramas and standard weather information (air temperature, relative humidity, wind speed). We validated the model for hot, dry, clear summer days in Tempe, Arizona, USA with in-situ observations using a FLIR Duo Pro R thermal camera on a rotating arm and the mobile human-biometeorological instrument platform MaRTy. We observed and modeled TMRT and thermal comfort for 19 sites with varying ground cover (grass, concrete, asphalt), sky view factor, exposure (sun, shade), and shade type (engineered, natural) six times per day. PanoMRT performed well with a Root Mean Square Error (RMSE) of 4.1 °C for TMRT, 2.6 °C for PET, and 1.2 °C for UTCI, meeting the accuracy requirement of ±5 °C set in the ISO 7726 standard for heat and cold stress studies. RayMan reference model runs without measured surface temperature forcing reveal that accurate longwave radiative flux estimations are crucial to meet the ±5 °C threshold, particularly for shaded locations and during midday when surface temperatures peak and longwave modeling errors are largest. This study demonstrates the importance of spatially resolved 3D surface temperature data for thermal exposure and comfort modeling to capture complex longwave radiation exposure patterns resulting from heterogeneity in built configuration and material radiative and thermal properties in the built environment.

Development and Characterization of 20 Genomic SSR Markers for Ornamental Cultivars of Weigela.

Weigela (Caprifoliaceae) is a genus of ornamental plants popular for its phenotypic variation and hardiness, that includes species hybridized to produce the commercially available cultivars. Despite its popularity, limited genetic resources exist for the genus. Twenty genomic simple sequence repeat (gSSR) markers distributed across the genome were developed using low coverage whole-genome sequencing data of Weigela Spilled Wine®. A cross-amplification evaluation with these 20 gSSR markers on a collection of 18 Weigela cultivars revealed a total of 111 unique alleles, including 36 private alleles. A diagrammatic key was constructed to identify cultivars using only six of the gSSR markers, demonstrating the newly developed gSSR markers are immediately useful for cultivar identification. Future uses could include breeding with marker-assisted selection, determining the history of hybridization of the current cultivated lines, aiding in the construction of genetic maps, and assessing the patterns of population genetic structure of Weigela spp.

Characterization and microsatellite marker development for a common bark and ambrosia beetle associate, Geosmithia obscura.

Symbioses between Geosmithia fungi and wood-boring and bark beetles seldom result in disease induction within the plant host. Yet, exceptions exist such as Geosmithia morbida, the causal agent of Thousand Cankers Disease (TCD) of walnuts and wingnuts, and Geosmithia sp. 41, the causal agent of Foamy Bark Canker disease of oaks. Isolates of G. obscura were recovered from black walnut trees in eastern Tennessee and at least one isolate induced cankers following artificial inoculation. Due to the putative pathogenicity and lack of recovery of G. obscura from natural lesions, a molecular diagnostic screening tool was developed using microsatellite markers mined from the G. obscura genome. A total of 3256 candidate microsatellite markers were identified (2236, 789, 137 di-, tri-, and tetranucleotide motifs, respectively), with 2011, 703, 101 di-, tri-, and tetranucleotide motifs, respectively, containing markers with primers. From these, 75 microsatellite markers were randomly selected, screened, and optimized, resulting in 28 polymorphic markers that yielded single, consistently recovered bands, which were used in downstream analyses. Five of these microsatellite markers were found to be specific to G. obscura and did not cross-amplify into other, closely related species. Although the remaining tested markers could be useful, they cross-amplified within different Geosmithia species, making them not reliable for G. obscura detection. Five novel microsatellite markers (GOBS9, GOBS10, GOBS41, GOBS43, and GOBS50) were developed based on the G. obscura genome. These species-specific microsatellite markers are available as a tool for use in molecular diagnostics and can assist future surveillance studies.

Chloroplast genome of the invasive Pyrus calleryana underscores the high molecular diversity of the species.

The complete chloroplast genome of Pyrus calleryana (GenBank OM541581.1) was developed by de novo assembly from whole-genome sequencing data. Reference-guided (P. phaeocarpa) read mapping and assembly were followed by annotation and phylogenetic comparisons. The 159,965 bp P. calleryana chloroplast genome represented 36.56% GC content with a classical quadripartite architecture and two inverted repeats regions (IRs; each 26,392 bp) separating the large single-copy region (LSC; 87,942 bp) and the small single-copy region (SSC; 19.239 bp). In total, 125 unique features were annotated in that genome, including 83 protein coding genes, 38 tRNA coding genes, and 4 rRNA coding genes. Phylogenetic analyses based on the whole chloroplast genome sequences placed the P. calleryana among other Rosaceae plants, specifically among the Asian species of Pyrus.

Integrated Genomic and Bioinformatics Approaches to Identify Molecular Links between Endocrine Disruptors and Adverse Outcomes.

Exposure to Endocrine Disrupting Chemicals (EDC) has been linked with several adverse outcomes. In this review, we examine EDCs that are pervasive in the environment and are of concern in the context of human, animal, and environmental health. We explore the consequences of EDC exposure on aquatic life, terrestrial animals, and humans. We focus on the exploitation of genomics technologies and in particular whole transcriptome sequencing. Genome-wide analyses using RNAseq provides snap shots of cellular, tissue and whole organism transcriptomes under normal physiological and EDC perturbed conditions. A global view of gene expression provides highly valuable information as it uncovers gene families or more specifically, pathways that are affected by EDC exposures, but also reveals those that are unaffected. Hypotheses about genes with unknown functions can also be formed by comparison of their expression levels with genes of known function. Risk assessment strategies leveraging genomic technologies and the development of toxicology databases are explored. Finally, we review how the Adverse Outcome Pathway (AOP) has exploited this high throughput data to provide a framework for toxicology studies.

Development and Characterization of 15 Novel Genomic SSRs for Viburnum farreri.

The Viburnum genus is of particular interest to horticulturalists, phylogeneticists, and biogeographers. Despite its popularity, there are few existing molecular markers to investigate genetic diversity in this large genus, which includes over 160 species. There are also few polymorphic molecular tools that can delineate closely related species within the genus. Viburnum farreri, a member of the Solenotinus subclade and one of the centers of diversity for Viburnum, was selected for DNA sequencing and development of genomic simple sequence repeats (gSSRs). In this study, 15 polymorphic gSSRs were developed and characterized for a collection of 19 V. farreri samples. Number of alleles per locus ranged from two- to- eight and nine loci had four or more alleles. Observed heterozygosity ranged from 0 to 0.84 and expected heterozygosity ranged from 0.10 to 0.80 for the 15 loci. Shannon diversity index values across these loci ranged from 0.21 to 1.62. The markers developed in this study add to the existing molecular toolkit for the genus and will be used in future studies investigating cross-transferability, genetic variation, and species and cultivar delimitation in the Viburnum genus and closely allied genera in the Adoxaceae and Caprifoliaceae.

The "Neglected Viruses" of Taihu: Abundant Transcripts for Viruses Infecting Eukaryotes and Their Potential Role in Phytoplankton Succession.

Drivers of algal bloom dynamics remain poorly understood, but viruses have been implicated as important players. Research addressing bloom dynamics has generally been restricted to the virus-infection of the numerically dominant (i.e. bloom forming) taxa. Yet this approach neglects a broad diversity of viral groups, limiting our knowledge of viral interactions and constraints within these systems. We examined hallmark virus marker genes in metatranscriptomic libraries from a seasonal and spatial survey of a Microcystis aeruginosa bloom in Lake Tai (Taihu) China to identify active infections by nucleocytoplasmic large DNA viruses (NCLDVs), RNA viruses, ssDNA viruses, bacteriophage, and virophage. Phylogenetic analyses revealed a diverse virus population with seasonal and spatial variability. We observed disproportionately high expression of markers associated with NCLDVs and ssRNA viruses (consistent with viruses that infect photosynthetic protists) relative to bacteriophage infecting heterotrophic bacteria or cyanobacteria during the height of the Microcystis bloom event. Under a modified kill-the-winner scheme, we hypothesize viruses infecting protists help suppress the photosynthetic eukaryotic community and allow for the proliferation of cyanobacteria such as Microcystis. Our observations provide a foundation for a little considered factor promoting algal blooms.

Pipeline for Integrated Microarray Expression Normalization Tool Kit (PIMENTo) for Tumor Microarray Profiling Experiments.

We have developed a Pipeline for Integrated Microarray Expression & Normalization Tool kit (PIMENTo) with the aim of streamlining the processes necessary for gene expression analysis in tumor tissue using DNA microarrays. Built with the R programming language and leveraging several open-source packages available through CRAN and Bioconductor, PIMENTo enables researchers to perform complex tasks with a minimal number of operations. Here, we describe the pipeline, review necessary data inputs, examine data outputs and quality control assessments and explore the commands to perform such analysis.

Genome-Wide Analysis of Low Dose Bisphenol-A (BPA) Exposure in Human Prostate Cells.

Endocrine disrupting compounds (EDCs) have the potential to cause adverse effects on wild-life and human health. Two important EDCs are the synthetic estrogen 17α-ethynylestradiol (EE2) and bisphenol-A (BPA) both of which are xenoestrogens (XEs) as they bind the estrogen receptor and dis-rupt estrogen physiology in mammals and other vertebrates. In the recent years the influence of XEs on oncogenes, specifically in relation to breast and prostate cancer has been the subject of considerable study. METHODOLOGY: In this study, healthy primary human prostate epithelial cells (PrECs) were exposed to environmentally relevant concentrations of BPA (5nM and 25nM BPA) and interrogated using a whole genome microarray. RESULTS: Exposure to 5 and 25nM BPA resulted in 7,182 and 7,650 differentially expressed (DE) genes, respectively in treated PrECs. Exposure to EE2 had the greatest effect on the PrEC transcriptome (8,891 DE genes). CONCLUSION: We dissected and investigated the nature of the non-estrogenic gene signature associated with BPA with a focus on transcripts relevant to epigenetic modifications. The expression of transcripts encoding nuclear hormone receptors as well as histone and DNA methylation, modifying enzymes were significantly perturbed by exposure to BPA.

Systems analysis of the liver transcriptome in adult male zebrafish exposed to the non-ionic surfactant nonylphenol.

Nonylphenol (NP) arises from the environmental degradation of nonylphenol ethoxylates. It is a ubiquitous environmental contaminant and has been detected at levels up to 167 nM in rivers in the United States. NP is an endocrine disruptor (ED) that can act as an agonist for estrogen receptors. The Adverse Outcome Pathway (AOP) framework defines an adverse outcome as the causal result of a series of molecular initiating events (MIEs) and key events (KEs) that lead to altered phenotypes. This study examined the liver transcriptome after a 21 day exposure to NP and 17ß-estradiol (E2) by exploiting the zebrafish (Danio rerio) as a systems toxicology model. The goal of this study was to tease out non-estrogenic genomic signatures associated with NP exposure using DNA microarray and RNA sequencing. Our experimental design included E2 as a positive and potent estrogenic control in order to effectively compare and contrast the 2 compounds. This approach allowed us to identify hepatic transcriptomic perturbations that could serve as MIEs for adverse health outcomes in response to NP. Our results revealed that exposure to NP was associated with differential expression (DE) of genes associated with the development of steatosis, disruption of metabolism, altered immune response, and metabolism of reactive oxygen species, further highlighting NP as a chemical of emerging concern (CEC).

Haplotyping of Cornus florida and C. kousa chloroplasts: Insights into species-level differences and patterns of plastic DNA variation in cultivars.

Chloroplast DNA is a part of plant non-nuclear genome, and is of particular interest for lineage studies. Moreover, the non-coding regions of cpDNA display higher mutation rates than the conserved coding cpDNA, which has been employed for phylogenetic and population research. We analyzed the cpDNA of 332 gDNA samples from collections of Cornus florida and C. kousa (commercial cultivars, breeding selections, and wild kousa accessions from Asia), using the chlorotyping system developed on North America-native, wild accessions of C. florida. Our results indicated significant differences in chlorotype frequencies between the two species. Cornus florida samples were represented by all major chlorotypes previously described, whereas all C. kousa samples analyzed had only one of the chlorotype patterns shown by C. florida. The chlorotyping analytic panel was then expanded by sequencing the targeted three non-coding cpDNA regions. Results indicated a major difference in the maternally-inherited cpDNA between the two closely related Big-Bracted Cornus species. Chlorotype diversity and differences in the proportion of informative sites in the cpDNA regions of focus emphasized the importance of proper loci choice for cpDNA-based comparative studies between the closely related dogwood species. Phylogenetic analyses of the retrieved sequences for the other species of Cornus provided information on the relative utility of the cpDNA regions studied and helped delineate the groups (Big-Bracted, Cornelian Cherries, Blue/White-Fruited) within the genus. Genealogical relationships based on the cpDNA sequences and the inferred chlorotype networks indicated the need for continued analyses across further non-coding cpDNA regions to improve the phylogenetic resolution of dogwoods.

Systems Analysis of the Liver Transcriptome in Adult Male Zebrafish Exposed to the Plasticizer (2-Ethylhexyl) Phthalate (DEHP).

The organic compound diethylhexyl phthalate (DEHP) represents a high production volume chemical found in cosmetics, personal care products, laundry detergents, and household items. DEHP, along with other phthalates causes endocrine disruption in males. Exposure to endocrine disrupting chemicals has been linked to the development of several adverse health outcomes with apical end points including Non-Alcoholic Fatty Liver Disease (NAFLD). This study examined the adult male zebrafish (Danio rerio) transcriptome after exposure to environmental levels of DEHP and 17α-ethinylestradiol (EE2) using both DNA microarray and RNA-sequencing technologies. Our results show that exposure to DEHP is associated with differentially expressed (DE) transcripts associated with the disruption of metabolic processes in the liver, including perturbation of five biological pathways: 'FOXA2 and FOXA3 transcription factor networks', 'Metabolic pathways', 'metabolism of amino acids and derivatives', 'metabolism of lipids and lipoproteins', and 'fatty acid, triacylglycerol, and ketone body metabolism'. DE transcripts unique to DEHP exposure, not observed with EE2 (i.e. non-estrogenic effects) exhibited a signature related to the regulation of transcription and translation, and ruffle assembly and organization. Collectively our results indicate that exposure to low DEHP levels modulates the expression of liver genes related to fatty acid metabolism and the development of NAFLD.

Comparison of multiplex PCR hybridization-based and singleplex real-time PCR-based assays for detection of low prevalence pathogens in spiked samples.

Molecular diagnostic devices are increasingly finding utility in clinical laboratories. Demonstration of the effectiveness of these devices is dependent upon comparing results from clinical samples tested with the new device to an alternative testing method. The preparation of mock clinical specimens will be necessary for the validation of molecular diagnostic devices when a sufficient number of clinical specimens is unobtainable. Examples include rare pathogens, some of which are pathogens posing a biological weapon threat. Here we describe standardized steps for developers to follow for the culture and quantification of three organisms used to spike human whole blood to create mock specimens. The three organisms chosen for this study were the Live Vaccine Strain (LVS) of Francisella tularensis, surrogate for a potential biothreat pathogen, Escherichia coli, a representative Gram-negative bacterium and Babesia microti (Franca) Reichenow Peabody strain, representing a protozoan parasite. Mock specimens were prepared with blood from both healthy donors and donors with nonspecific symptoms including fever, malaise, and flu-like symptoms. There was no significant difference in detection results between the two groups for any pathogen. Testing of the mock samples was compared on two platforms, Target Enriched Multiplex-PCR (TEM-PCR™) and singleplex real-time PCR (RT-PCR). Results were reproducible on both platforms. The reproducibility demonstrated by obtaining the same results between two testing methods and between healthy and symptomatic mock specimens, indicates the standardized methods described for creating the mock specimens are valid and effective for evaluating diagnostic devices.

Biochar-surface oxygenation with hydrogen peroxide.

Biochar was produced from pinewood biomass by pyrolysis at a highest treatment temperature (HTT) of 400 °C. This biochar was then treated with varying concentrations of H2O2 solution (1, 3, 10, 20, 30% w/w) for a partial oxygenation study. The biochar samples, both treated and untreated, were then tested with a cation exchange capacity (CEC) assay, Fourier Transformed Infrared Resonance (FT-IR), elemental analysis, field water-retention capacity assay, pH assay, and analyzed for their capacity to remove methylene blue from solution. The results demonstrated that higher H2O2 concentration treatments led to higher CEC due to the addition of acidic oxygen functional groups on the surface of the biochar, which also corresponds to the resultant lowering of the pH of the biochar with respect to the H2O2 treatment. Furthermore, it was shown that the biochar methylene blue adsorption decreased with higher H2O2 concentration treatments. This is believed to be due to the addition of oxygen groups onto the aromatic ring structure of the biochar which in turn weakens the overall dispersive forces of π-π interactions that are mainly responsible for the adsorption of the dye onto the surface of the biochar. Elemental analysis revealed that there was no general augmentation of the elemental composition of the biochar samples through the treatment with H2O2, which suggests that the bulk property of biochar remains unchanged through the treatment.

Seasonal variation of respiratory pathogen colonization in asymptomatic health care professionals: A single-center, cross-sectional, 2-season observational study.

BACKGROUND: The purpose of this study was to determine the seasonal variance of potentially pathogenic bacterial and viral organisms in nasopharyngeal specimens obtained from asymptomatic health care professionals (HCPs) during the 2014 winter and summer months. METHODS: Nasopharyngeal specimens from 100 HCPs were collected from Huntsville Hospital (Huntsville, AL) during the winter and from 100 HCPs during the summer. All subjects were tested for 22 viruses and 19 bacteria using Target Enriched Multiplex Polymerase Chain Reaction. Both seasonal cohorts were composed of students, nurses, physicians, and residents. RESULTS: Of the 100 HCPs tested during the winter, 34 subjects were colonized with at least 1 bacterium, and 11 tested positive for at least 1 virus. Methicillin-resistant Staphylococcus aureus (MRSA), Moraxella catarrhalis, and coronavirus were the most frequently detected potentially infectious agents. Of the 100 HCPs tested during the summer, 37 tested positive for at least 1 bacterium, and 4 tested positive for a viral agent. The most prevalent bacteria were MRSA and Klebsiella pneumonia. CONCLUSION: Nasopharyngeal carriage among asymptomatic HCPs was common, but the frequency and presence of potential pathogens varied with each season. Understanding the colonization and infection potential of upper respiratory organisms is important, particularly for viruses. Although asymptomatic HCPs certainly harbor a number of different potentially infectious agents, future studies are needed to determine whether colonized pathogens are transmitted or initiate infection in at-risk patient populations.