Postbiotic lactobacilli induce cutaneous antimicrobial response and restore the barrier to inhibit the intracellular invasion of Staphylococcus aureus in vitro and ex vivo.

Intracellular pathogens including Staphylococcus aureus contribute to the non-healing phenotype of chronic wounds. Lactobacilli, well known as beneficial bacteria, are also reported to modulate the immune system, yet their role in cutaneous immunity remains largely unknown. We explored the therapeutic potential of bacteria-free postbiotics, bioactive lysates of lactobacilli, to reduce intracellular S. aureus colonization and promote healing. Fourteen postbiotics derived from various lactobacilli species were screened, and Latilactobacillus curvatus BGMK2-41 was selected for further analysis based on the most efficient ability to reduce intracellular infection by S. aureus diabetic foot ulcer clinical isolate and S. aureus USA300. Treatment of both infected keratinocytes in vitro and infected human skin ex vivo with BGMK2-41 postbiotic cleared S. aureus. Keratinocytes treated in vitro with BGMK2-41 upregulated expression of antimicrobial response genes, of which DEFB4, ANG, and RNASE7 were also found upregulated in treated ex vivo human skin together with CAMP exclusively upregulated ex vivo. Furthermore, BGMK2-41 postbiotic treatment has a multifaceted impact on the wound healing process. Treatment of keratinocytes stimulated cell migration and the expression of tight junction proteins, while in ex vivo human skin BGMK2-41 increased expression of anti-inflammatory cytokine IL-10, promoted re-epithelialization, and restored the epidermal barrier via upregulation of tight junction proteins. Together, this provides a potential therapeutic approach for persistent intracellular S. aureus infections.

Race and ethnicity are inadequate predictors of ambulatory visit length and utilization of preventive services.

Health disparities can be experienced by any disadvantaged group who has limited access to healthcare or decreased quality of care. Quality of care can be measured by physician-patient communication measures such as length of visit, health outcomes, patient satisfaction, or by the services one receives such as screening or health education. This study aims to determine the relationship between length of physician-patient encounter, number of preventive services, ethnicity, and race. This study utilizes data from the National Ambulatory Medical Care Survey (NAMCS) from 2007 to 2016. Visits with a single diagnosis were selected. Visits with the five most frequent diagnoses were selected by International Classification of Diseases, Ninth or Tenth Revision (ICD-9/ICD-10) classification. The primary outcome is time spent with a physician in minutes and the number of preventive services provided represented by the Preventive Service Index (PSI). Of 255,916 visits, non-white individuals made up 16.2% (95% Confidence Interval 15.9-16.4) while Latinos represented 13.4% (95%CI 13.2-13.6) of individuals. Multivariate analysis revealed minimal differences in visit length in race and ethnic groups regardless of diagnosis. Greater PSI was associated with individuals less than 43 years old (Odds Ratio (OR) 2.0, 95% CI 1.8-2.3, p =< 0.0001), those who reside in metropolitan statistical areas (MSAs) (OR 1.2, 95% CI 1.1-1.4, p = 0.006), non-white individuals (OR 1.2, 95% CI 1.1-1.3, p = 0.004), and those with private insurance (OR 1.3, 95% CI 1.1-1.4, p =< 0.0001). Race and ethnicity do not predict length of time with a physician regardless of diagnosis. Age, race, location within a metropolitan area, and insurance are significant but minimal predictors of receiving preventive services in the rank-order leading five most frequent diagnoses. This large, population-based study highlights improvements in the distribution of healthcare services from previous studies.

Increased biologic utilization in Latino patients with psoriasis.

INTRODUCTION: Hispanic/Latino populations are more likely to have extensive psoriasis than the non-Hispanic/Latino population. Biologics are indicated for moderate/severe psoriasis or psoriasis with comorbidities. No studies have assessed ethnicity as a predictor of biologic utilization. We aimed to determine if biologic utilization differs between Latinos and non-Latinos with psoriasis. METHODS: This study utilizes data from the National Ambulatory Medical Care Survey (NAMCS) from 2003 to 2016. Psoriasis visits were selected by International Classification of Diseases, Ninth or Tenth Revision (ICD-9/ICD-10) classification. The primary outcome is biologic use at the time of visit. RESULTS: Of 1202 psoriasis visits, Latinos consisted of 9.7% (95% Confidence Interval 7.3-12) of the study population and 65% (95%CI 61-69) used private insurance. Multivariate analysis reveals the increased likelihood of biologic utilization in Latinos compared to non-Latinos (Odds Ratio (OR) 2.4, 95% CI 1.1-5.4, p = .03). Independently, private insurance status is associated with increased biologic use compared to public insurance (OR 2.4, 95% CI 1.2-5.0, p = .02). Private insurance status did not differ between Latinos and non-Latinos with psoriasis (OR 1.0, 95% CI 0.5-1.9, p = .98). CONCLUSIONS: Hispanic/Latino ethnicity and private insurance status are independent predictors of biologic utilization, suggesting that biologic mechanisms may influence the increased use of biologics in Latinos with psoriasis.

Biofilm propensity of Staphylococcus aureus skin isolates is associated with increased atopic dermatitis severity and barrier dysfunction in the MPAACH pediatric cohort.

BACKGROUND: Atopic dermatitis (AD) patients are often colonized with Staphylococcus aureus, and staphylococcal biofilms have been reported on adult AD skin lesions. The commensal S epidermidis can antagonize S aureus, although its role in AD is unclear. We sought to characterize S aureus and S epidermidis colonization and biofilm propensity and determine their associations with AD severity, barrier function, and epidermal gene expression in the first US early-life cohort of children with AD, the Mechanisms of Progression of Atopic Dermatitis to Asthma in Children (MPAACH). METHODS: The biofilm propensity of staphylococcal isolates was assessed by crystal violet assays. Gene expression of filaggrin and antimicrobial alarmins S100A8 and S100A9 was measured in keratinocyte RNA extracted from skin tape strips. Staphylococcal biofilms sampled from MPAACH skin were visualized using scanning electron microscopy. RESULTS: Sixty-two percent of staphylococcal isolates (sampled from 400 subjects) formed moderate/strong biofilms. Sixty-eight percent of subjects co-colonized with both staphylococcal species exhibited strains that formed cooperative mixed-species biofilms. Scanning electron microscopy verified the presence of staphylococcal biofilms on the skin of MPAACH children. Staphylococcus aureus strains showing higher relative biofilm propensity compared with S epidermidis were associated with increased AD severity (P = .03) and increased lesional and nonlesional transepidermal water loss (P = .01, P = .03). CONCLUSIONS: Our data suggest a pathogenic role for S aureus biofilms in AD. We found that strain-level variation in staphylococcal isolates governs the interactions between S epidermidis and S aureus and that the balance between these two species, and their biofilm propensity, has important implications for AD.

On the surface: Skin microbial exposure contributes to allergic disease.

OBJECTIVE: To discuss the skin microbiome modulates immunity by interactions between skin immunology with keratinocytes to combat pathogens. Allergic disorders are classified by immunoglobulin E sensitivity and aberrant TH2 cell responses, and an increasing number of studies have described the associations with skin microbiome fluctuations. In this review, we discuss commensal-epidermal homeostasis and its influence on allergic disease. DATA SOURCES: All included references were obtained from the PubMed database. STUDY SELECTIONS: Studies addressing relevant aspects of commensal-epidermal homeostasis, skin microbiome dysbiosis, microbiome-targeted therapeutics, and prevention in allergy were included. RESULTS: Homeostasis between the commensal microbiome and the epidermis is important in protecting against allergic disease. Commensals promote antiallergic TH1 and TH17 immunophenotypes within the skin and induce keratinocytes to secrete antimicrobial peptides and alarmins that enhance barrier function and antagonize proallergic organisms. Perturbations in this homeostasis, however, is associated with allergic disease development. Atopic dermatitis is associated with decreases in skin commensals and increases in the pathogen, Staphylococcus aureus. Fluctuations in the skin microbiome contributes to decreased barrier dysfunction, allergic sensitization, and TH2 cytokine secretion. Little is known about how the skin microbiome affects food allergy, allergic rhinitis, and asthma, and it is poorly understood how cutaneous inflammation influences systemic allergic responses. Therapies are targeted toward maintenance of the skin barrier, replacement of healthy commensals, and anti-TH2 biologic therapy. CONCLUSION: Although the effects of commensal-epidermal homeostasis on allergy within the skin are becoming increasingly clear, future studies are necessary to assess its effects on extracutaneous allergic disorders and explore potential therapeutics targeting the skin microbiome.

Events in Normal Skin Promote Early-Life Atopic Dermatitis-The MPAACH Cohort.

BACKGROUND: Nonlesional skin in atopic dermatitis (AD) is abnormal, but the pathobiology of lesional and nonlesional skin and the definition of endotypes are poorly understood. OBJECTIVE: To define lesional and nonlesional endotypes of AD by building the first US-based early-life prospective cohort of children with AD, the Mechanisms of Progression from AD to Asthma in Children cohort. METHODS: We assessed lesional and nonlesional skin transepidermal water loss, filaggrin (FLG) and alarmin (S100A8, S100A9) expression, staphylococcal colonization, and patterns of aeroallergen and food sensitization to define nonlesional and lesional phenotypes and endotypes. RESULTS: Pathophysiologic changes were present in lesional and nonlesional skin and were associated with SCORing for Atopic Dermatitis. Nonlesional skin had features characteristic of diseased skin including low FLG and high alarmin expression, and increased colonization with Staphylococcus aureus. In a multivariate model, nonlesional, but not lesional, FLG expression was associated with the development of cosensitization and moderate to severe AD. Lesional skin was characterized by further deficits in FLG expression (P < .001), but alarmin expression was the same as observed in nonlesional skin. CONCLUSIONS: This study reveals that events in the nonlesional, not the lesional, skin promote the subsequent development of AD severity and cosensitization, which is a key risk factor for allergic comorbidities. Collectively, these data suggest the presence of a subclinical eczema endotype that may predispose to the development of allergic disease in the absence of overt eczema. This may represent a new definition of the atopic march that starts with skin barrier dysfunction rather than eczema.

Commensal Candida albicans Positively Calibrates Systemic Th17 Immunological Responses.

Mucosal barriers are densely colonized by pathobiont microbes such as Candida albicans, capable of invasive disseminated infection. However, systemic infections occur infrequently in healthy individuals, suggesting that pathobiont commensalism may elicit host benefits. We show that intestinal colonization with C. albicans drives systemic expansion of fungal-specific Th17 CD4+ T cells and IL-17 responsiveness by circulating neutrophils, which synergistically protect against C. albicans invasive infection. Protection conferred by commensal C. albicans requires persistent fungal colonization and extends to other extracellular invasive pathogens such as Staphylococcus aureus. However, commensal C. albicans does not protect against intracellular influenza virus infection and exacerbates allergic airway inflammation susceptibility, indicating that positively calibrating systemic Th17 responses is not uniformly beneficial. Thus, systemic Th17 inflammation driven by CD4+ T cells responsive to tonic stimulation by commensal C. albicans improves host defense against extracellular pathogens, but with potentially harmful immunological consequences.

An in vitro proof-of-principle study of sonobactericide.

Infective endocarditis (IE) is associated with high morbidity and mortality rates. The predominant bacteria causing IE is Staphylococcus aureus (S. aureus), which can bind to existing thrombi on heart valves and generate vegetations (biofilms). In this in vitro flow study, we evaluated sonobactericide as a novel strategy to treat IE, using ultrasound and an ultrasound contrast agent with or without other therapeutics. We developed a model of IE biofilm using human whole-blood clots infected with patient-derived S. aureus (infected clots). Histology and live-cell imaging revealed a biofilm layer of fibrin-embedded living Staphylococci around a dense erythrocyte core. Infected clots were treated under flow for 30 minutes and degradation was assessed by time-lapse microscopy imaging. Treatments consisted of either continuous plasma flow alone or with different combinations of therapeutics: oxacillin (antibiotic), recombinant tissue plasminogen activator (rt-PA; thrombolytic), intermittent continuous-wave low-frequency ultrasound (120-kHz, 0.44 MPa peak-to-peak pressure), and an ultrasound contrast agent (Definity). Infected clots exposed to the combination of oxacillin, rt-PA, ultrasound, and Definity achieved 99.3 ± 1.7% loss, which was greater than the other treatment arms. Effluent size measurements suggested low likelihood of emboli formation. These results support the continued investigation of sonobactericide as a therapeutic strategy for IE.

Staphylococcal Biofilms in Atopic Dermatitis.

PURPOSE OF REVIEW: Atopic dermatitis (AD) is a chronic, relapsing inflammatory skin disorder that is a major public health burden worldwide. AD lesions are often colonized by Staphylococcus aureus and Staphylococcus epidermidis. An important aspect of Staphylococcus spp. is their propensity to form biofilms, adhesive surface-attached colonies that become highly resistant to antibiotics and immune responses, and recent studies have found that clinical isolates colonizing AD skin are often biofilm-positive. Biofilm formation results in complex bacterial communities that have unique effects on keratinocytes and host immunity. This review will summarize recent studies exploring the role of staphyloccocal biofilms in atopic dermatitis and the implications for treatment. RECENT FINDINGS: Recent studies suggest an important role for biofilms in the pathogenesis of numerous dermatologic diseases including AD. S. aureus biofilms have been found to colonize the eccrine ducts of AD skin, and these biofilms influence secretion of keratinocyte cytokines and trigger differentiation and apoptosis of keratinocytes. These activities may act to disrupt barrier function and promote disease pathogenesis as well as allergen sensitization. Formation of biofilm is a successful strategy that protects the bacteria from environmental danger, antibiotics, and phagocytosis, enabling chronic persistence in the host. An increasing number of S. aureus skin isolates are resistant to conventional antibiotics, and staphylococcal biofilm communities are prevalent on the skin of individuals with AD. Staphylococcal colonization of the skin impacts skin barrier function and plays multiple important roles in AD pathogenesis.

Escherichia coli lipoprotein binds human plasminogen via an intramolecular domain.

Escherichia coli lipoprotein (Lpp) is a major cellular component that exists in two distinct states, bound-form and free-form. Bound-form Lpp is known to interact with the periplasmic bacterial cell wall, while free-form Lpp is localized to the bacterial cell surface. A function for surface-exposed Lpp has yet to be determined. We hypothesized that the presence of C-terminal lysinses in the surface-exposed region of Lpp would facilitate binding to the host zymogen plasminogen (Plg), a protease commandeered by a number of clinically important bacteria. Recombinant Lpp was synthesized and the binding of Lpp to Plg, the effect of various inhibitors on this binding, and the effects of various mutations of Lpp on Lpp-Plg interactions were examined. Additionally, the ability of Lpp-bound Plg to be converted to active plasmin was analyzed. We determined that Lpp binds Plg via an atypical domain located near the center of mature Lpp that may not be exposed on the surface of intact E. coli according to the current localization model. Finally, we found that Plg bound by Lpp can be converted to active plasmin. While the consequences of Lpp binding Plg are unclear, these results prompt further investigation of the ability of surface exposed Lpp to interact with host molecules such as extracellular matrix components and complement regulators, and the role of these interactions in infections caused by E. coli and other bacteria.

BB0347, from the lyme disease spirochete Borrelia burgdorferi, is surface exposed and interacts with the CS1 heparin-binding domain of human fibronectin.

The causative agent of Lyme disease, Borrelia burgdorferi, codes for several known fibronectin-binding proteins. Fibronectin a common the target of diverse bacterial pathogens, and has been shown to be essential in allowing for the development of certain disease states. Another borrelial protein, BB0347, has sequence similarity with these other known fibronectin-binding proteins, and may be important in Lyme disease pathogenesis. Herein, we perform an initial characterization of BB0347 via the use of molecular and biochemical techniques. We found that BB0347 is expressed, produced, and presented on the outer surface of intact B. burgdorferi. We also demonstrate that BB0347 has the potential to be important in Lyme disease progression, and have begun to characterize the nature of the interaction between human fibronectin and this bacterial protein. Further work is needed to define the role of this protein in the borrelial infection process.

Evaluation of RevA, a fibronectin-binding protein of Borrelia burgdorferi, as a potential vaccine candidate for lyme disease.

Previous studies indicated that the Lyme disease spirochete Borrelia burgdorferi expresses the RevA outer surface protein during mammalian infection. As an adhesin that promotes bacterial interaction with fibronectin, RevA appears to be a good target for preventive therapies. RevA proteins are highly conserved across all Lyme borreliae, and antibodies against RevA protein are cross-reactive among RevA proteins from diverse strains. Mice infected with B. burgdorferi mounted a rapid IgM response to RevA, followed by a strong IgG response that generally remained elevated for more than 12 months, suggesting continued exposure of RevA protein to the immune system. RevA antibodies were bactericidal in vitro. To evaluate the RevA antigen as a potential vaccine, mice were vaccinated with recombinant RevA and challenged with B. burgdorferi by inoculation with a needle or by a tick bite. Cultured tissues from all treatment groups were positive for B. burgdorferi. Vaccinated animals also appeared to have similar levels of B. burgdorferi DNA compared to nonvaccinated controls. Despite its antigenicity, surface expression, and the production of bactericidal antibodies against it, RevA does not protect against Borrelia burgdorferi infection in a mouse model. However, passive immunization with anti-RevA antibodies did prevent infection, suggesting the possible utility of RevA-based immunotherapeutics or vaccine.