Microbial burden of chronic wounds is believed to play an important role in impaired healing and development of infection-related complications. However, clinical cultures have little predictive value of wound outcomes, and culture-independent studies have been limited by cross-sectional design and small cohort size. We systematically evaluated the temporal dynamics of the microbiota colonizing diabetic foot ulcers (DFU), a common and costly complication of diabetes, and its association with healing and clinical complications. Dirichlet multinomial mixture modeling, Markov chain analysis, and mixed-effect models were used to investigate shifts in the microbiota over time and its associations with healing. Here we show to our knowledge previously unreported temporal dynamics of the chronic wound microbiome. Microbiota community instability was associated with faster healing and improved outcomes. DFU microbiota were found to exist in one of four community types that experienced frequent and non-random transitions. Transition patterns and frequencies associated with healing time. Exposure to systemic antibiotics destabilized the wound microbiota, rather than altering overall diversity or relative abundance of specific taxa. This study provides to our knowledge previously unreported evidence that the dynamic wound microbiome is indicative of clinical outcomes and may be a valuable guide for personalized management and treatment of chronic wounds.

Chronic nonhealing wounds have been heralded as a silent epidemic, causing significant morbidity and mortality especially in elderly, diabetic, and obese populations. Polymicrobial biofilms in the wound bed are hypothesized to disrupt the highly coordinated and sequential events of cutaneous healing. Both culture-dependent and -independent studies of the chronic-wound microbiome have almost exclusively focused on bacteria, omitting what we hypothesize are important fungal contributions to impaired healing and the development of complications. Here we show for the first time that fungal communities (the mycobiome) in chronic wounds are predictive of healing time, associated with poor outcomes, and form mixed fungal-bacterial biofilms. We longitudinally profiled 100, nonhealing diabetic-foot ulcers with high-throughput sequencing of the pan-fungal internal transcribed spacer 1 (ITS1) locus, estimating that up to 80% of wounds contain fungi, whereas cultures performed in parallel captured only 5% of colonized wounds. The “mycobiome” was highly heterogeneous over time and between subjects. Fungal diversity increased with antibiotic administration and onset of a clinical complication. The proportions of the phylum Ascomycota were significantly greater (P = 0.015) at the beginning of the study in wounds that took >8 weeks to heal. Wound necrosis was distinctly associated with pathogenic fungal species, while taxa identified as allergenic filamentous fungi were associated with low levels of systemic inflammation. Directed culturing of wounds stably colonized by pathogens revealed that interkingdom biofilms formed between yeasts and coisolated bacteria. Combined, our analyses provide enhanced resolution of the mycobiome during impaired wound healing, its role in chronic disease, and impact on clinical outcomes.

Highlights d Wound microbiota was profiled longitudinally in patients with diabetic foot ulcers d Staphylococcus aureus strains were associated with poor outcomes d S. aureus and other wound isolates promoted differential wound healing responses d Debridement depleted anaerobic bacteria in wounds with favorable outcomes

Summary Skin microbiota can impact allergic and autoimmune responses, wound healing and anti-microbial defense. We investigated the role of skin microbiota in cutaneous leishmaniasis and found that human patients infected with Leishmania braziliensis develop dysbiotic skin microbiota, characterized by increases in the abundance of Staphylococcus and/or Streptococcus. Mice infected with L. major exhibit similar changes depending upon disease severity. Importantly, this dysbiosis is not limited to the lesion site, but is transmissible to normal skin distant from the infection site, and to skin from co-housed naïve mice. This observation allowed us to test whether a preexisting dysbiotic skin microbiota influences disease, and we found that challenging dysbiotic naïve mice with L. major or testing for contact hypersensitivity results in exacerbated skin inflammatory responses. These findings demonstrate that a dysbiotic skin microbiota is not only a consequence of tissue stress, but also enhances inflammation, which has implications for many inflammatory cutaneous diseases.

Plaque psoriasis, a chronic inflammatory disease primarily affecting the skin, is thought to have a multifactorial etiology, including innate immune system dysregulation, environmental triggers, and genetic susceptibility. We sought to further understand the role of skin microbiota in psoriasis pathogenesis, as well as their response to therapy. We systematically analyzed dynamic microbiota colonizing psoriasis lesions and adjacent nonlesional skin in 114 patients prior to and during ustekinumab treatment in a phase 3b clinical trial. By sequencing the bacterial 16S ribosomal RNA gene from skin swab samples obtained at six anatomical sites, we identified minor, site-specific differences in microbial diversity and composition between pretreatment lesional and nonlesional skin. During therapy, microbial communities within lesional and nonlesional skin diverged, and body-site dispersion increased, reflecting microbial skin site-specificity. Microbiota demonstrated greater pretreatment heterogeneity in psoriatic lesions than in nonlesional skin, and variance increased as treatment progressed. Microbiota colonizing recurrent lesions did not overlap with pretreatment lesional microbiota, suggesting colonization patterns varied between initial and recurrent psoriatic lesions. While plaque psoriasis does not appear to be associated with specific microbes and/or microbial diversity, this large dataset provides insight into microbial variation associated with (i) disease in different body locations, (ii) initial versus recurrent lesions, and (iii) anti-IL12/23 therapy.

Cutaneous leishmaniasis is a disease characterized by ulcerating skin lesions, the resolution of which requires an effective, but regulated, immune response that limits parasite growth without causing permanent tissue damage. While mechanisms that control the parasites have been well studied, the factors regulating immunopathologic responses are less well understood. IL-22, a member of the IL-10 family of cytokines, can contribute to wound healing, but in other instances promotes pathology. Here we investigated the role of IL-22 during leishmania infection, and found that IL-22 limits leishmania-induced pathology when a certain threshold of damage is induced by a high dose of parasites. Il22 -/- mice developed more severe disease than wild-type mice, with significantly more pathology at the site of infection, and in some cases permanent loss of tissue. The increased inflammation was not due to an increased parasite burden, but rather was associated with the loss of a wound healing phenotype in keratinocytes. Taken together, these studies demonstrate that during cutaneous leishmaniasis, IL-22 can play a previously unappreciated role in controlling leishmania-induced immunopathology.

Objectives: Difficult intravenous access (DIVA) is common in the emergency department (ED). We investigated the extent to which DIVA is associated with care delay outcomes including time to first laboratory draw, therapies, imaging, and ED disposition. Methods: An observational retrospective cohort analysis of patients with DIVA treated between 2018 and 2020 at 2 urban academic EDs was performed. DIVA was defined as patients requiring ultrasound-guided intravenous access placed by physicians or advanced practice providers (APPs) as opposed to landmark-based intravenous placement by nurses. ED throughput variables and disposition time were compared. We correlated DIVA with time to administration of intravenous pain medications, fluids, imaging contrast, laboratory results, and ED disposition. Results: A total of 108,256 subjects with 161,122 total encounters were included. DIVA occurred in 4961 (3.1%) of ED visits. Patients with DIVA were more likely to be female (3.5% vs 2.6% for males, odds ratio [OR] 1.34, 95% confidence interval [CI]: 1.27-1.42), self-identify as black (OR 1.78, 95% CI: 1.66-1.91), and have higher acuity of illness (P < 0.001). Among pediatric patients, DIVA occurred most often in the first year of life at a rate of 3.25%. In adults, DIVA occurred in 2 age peaks; at 35 years (4.02%), and at 63 years (3.44%). In all workflow metrics, the presence of DIVA was associated with significant delays in median time to completion: 50 minutes for pain

Given widespread use of spike antibody in generating coronavirus disease vaccines, SARS-CoV-2 nucleocapsid antibodies are increasingly used to indicate previous infection in serologic surveys. However, longitudinal kinetics and seroreversion are poorly defined. We found substantial seroreversion of nucleocapsid total immunoglobulin, underscoring the need to account for seroreversion in seroepidemiologic studies.