Role of the extracellular matrix in Candida biofilm antifungal resistance.

Clinical infection due to Candida species frequently involve growth in biofilm communities. Recalcitrance despite antifungal therapy leads to disease persistence associated with high morbidity and mortality. Candida possesses several tools allowing evasion of antifungal effects. Among these, protection of biofilm cells via encasement by the extracellular matrix is responsible for a majority drug resistance phenotype. The Candida matrix composition is complex and includes a mannan-glucan complex linked to antifungal drug sequestration. This mechanism of resistance is conserved across the Candida genus and impacts each of the available antifungal drug classes. The exosome pathway is responsible for delivery and assembly of much of the Candida extracellular matrix as functional vesicle protein and polysaccharide cargo. Investigations demonstrate the vesicle matrix delivery pathway is a useful fungal biofilm drug target. Further elucidation of the vesicle pathway, as well as understanding the roles of biofilm driven cargo may provide additional targets to aid the diagnosis, prevention, and treatment of Candida biofilms.

Extracellular Vesicles Contribute to Mixed-Fungal Species Competition during Biofilm Initiation.

Extracellular vesicles commonly modulate interactions among cellular communities. Recent studies demonstrate that biofilm maturation features, including matrix production, drug resistance, and dispersion, require the delivery of a core protein and carbohydrate vesicle cargo in Candida species. The function of the vesicle cargo for these advanced-phase biofilm characteristics appears to be conserved across Candida species. Mixed-species interactions in mature biofilms indicate that vesicle cargo serves a cooperative role in preserving the community. Here, we define the function of biofilm-associated vesicles for biofilm initiation both within and among five species across the Candida genus. We found similar vesicle cargo functions for several conserved proteins across species, based on the behavior of mutants. Repletion of the adhesion environment with wild-type vesicles returned the community phenotype toward reference levels in intraspecies experiments. However, cross-species vesicle complementation did not restore the wild-type biology and in fact drove the phenotype in the opposite direction for most cross-species interactions. Further study of mixed-species biofilm adhesion and exogenous wild-type vesicle administration similarly demonstrated competitive interactions. Our studies indicate that similar vesicle cargoes contribute to biofilm initiation. However, vesicles from disparate species serve an interference competitive role in mixed-Candida species scenarios. IMPORTANCE Candida species commonly form mixed-species biofilms with other Candida species and bacteria. In the established biofilm state, vesicle cargo delivers public goods to support the mature community. At biofilm initiation, however, vesicles play a negative role in cross-species interactions, presumably to allow species to gain a survival advantage. These observations and recent reports reveal that vesicle cargo has both cooperative and competitive roles among Candida species, depending on the needs of the community biofilm formation.

Rapid breath analysis for acute respiratory distress syndrome diagnostics using a portable two-dimensional gas chromatography device.

Acute respiratory distress syndrome (ARDS) is the most severe form of acute lung injury, responsible for high mortality and long-term morbidity. As a dynamic syndrome with multiple etiologies, its timely diagnosis is difficult as is tracking the course of the syndrome. Therefore, there is a significant need for early, rapid detection and diagnosis as well as clinical trajectory monitoring of ARDS. Here, we report our work on using human breath to differentiate ARDS and non-ARDS causes of respiratory failure. A fully automated portable 2-dimensional gas chromatography device with high peak capacity (> 200 at the resolution of 1), high sensitivity (sub-ppb), and rapid analysis capability (~ 30 min) was designed and made in-house for on-site analysis of patients' breath. A total of 85 breath samples from 48 ARDS patients and controls were collected. Ninety-seven elution peaks were separated and detected in 13 min. An algorithm based on machine learning, principal component analysis (PCA), and linear discriminant analysis (LDA) was developed. As compared to the adjudications done by physicians based on the Berlin criteria, our device and algorithm achieved an overall accuracy of 87.1% with 94.1% positive predictive value and 82.4% negative predictive value. The high overall accuracy and high positive predicative value suggest that the breath analysis method can accurately diagnose ARDS. The ability to continuously and non-invasively monitor exhaled breath for early diagnosis, disease trajectory tracking, and outcome prediction monitoring of ARDS may have a significant impact on changing practice and improving patient outcomes. Graphical abstract.

Direct observation of Medicaid beneficiary attempts to fill prescriptions for nicotine replacement medications.

OBJECTIVES: Although many states have expanded Medicaid coverage of cessation medications, utilization remains low. Anecdotal reports suggest that beneficiaries are at times denied coverage of cessation medications at the pharmacy counter. We conducted an observational community-wide case study of Medicaid beneficiary attempts to fill over-the-counter nicotine replacement therapy at pharmacies. METHODS: We recruited tobacco-using beneficiaries from a Federally Qualified Health Center, whose providers wrote paper prescriptions for nicotine patches. Study staff escorted beneficiaries to all eligible pharmacies (n = 18) in a Midwestern community to observe fill attempts. Study staff recorded encounters via smartphone into a secure database on a university server. RESULTS: Seven of 18 pharmacies (39%) did not fill the prescription on the day of the attempt. Of these, 6 offered to order the patch for pick-up at a later date. All (4/4) chain pharmacies filled the prescription; 2/3 mass merchant pharmacies failed to fill. Combining successful same-day fills with offers to order for pick-up, 17/18 (94%) would ultimately have been able to obtain patches. CONCLUSION: This pilot study found that many beneficiaries left pharmacies without a prescription in hand. Successful same-day fills varied markedly by store type. For people with low incomes, transportation presents a major barrier for delayed pick-up. In addition, delays can fuel ambivalence toward quitting. Future research based on this pilot study might address whether patients who fail to secure a same-day prescription ever fill the prescription and, if not, the degree to which this barrier contributes to success or failure in quitting.

Partial immune reconstitution of X-linked hyper IgM syndrome with recombinant CD40 ligand.

X-linked hyper IgM syndrome (XHM) is a combined immune deficiency disorder caused by genetic alterations in CD40 ligand. The purpose of this study was to investigate the safety and efficacy of recombinant CD40 ligand (rCD40L) in the treatment of the disease. Three children were administered rCD40L subcutaneously 3 times per week at 0.03 mg/kg for 22 weeks, and after a 12-week drug-free interval, the dose was increased to 0.05 mg/kg for an additional 22 weeks of treatment. Although specific antibody responses to T cell-dependent antigens was lacking, administration of rCD40 resulted in acquisition of the capacity to mount cutaneous delayed type hypersensitivity reactions that disappeared during the drug-free interval as well as the postbiologic follow-up period. With rCD40L treatment, patient T cells developed a new capacity to respond to T-cell mitogens with synthesis of IFN-γ and TNF-α. Intracellular cytokine staining studies showed that both CD4(+) and CD8(+) T cells participated in this response. Finally, CD40L therapy was associated with changes in lymph node size and architecture based on comparison of biopsies taken before and after therapy. This clinical study showed that rCD40L is capable of improving T cell-immune function in patients with XHM.