How Three Self-Secreted Biofilm Exopolysaccharides of Pseudomonas aeruginosa, Psl, Pel, and Alginate, Can Each Be Exploited for Antibiotic Adjuvant Effects in Cystic Fibrosis Lung Infection.

In cystic fibrosis (CF), pulmonary infection with Pseudomonas aeruginosa is a cause of increased morbidity and mortality, especially in patients for whom infection becomes chronic and there is reliance on long-term suppressive therapies. Current antimicrobials, though varied mechanistically and by mode of delivery, are inadequate not only due to their failure to eradicate infection but also because they do not halt the progression of lung function decline over time. One of the reasons for this failure is thought to be the biofilm mode of growth of P. aeruginosa, wherein self-secreted exopolysaccharides (EPSs) provide physical protection against antibiotics and an array of niches with resulting metabolic and phenotypic heterogeneity. The three biofilm-associated EPSs secreted by P. aeruginosa (alginate, Psl, and Pel) are each under investigation and are being exploited in ways that potentiate antibiotics. In this review, we describe the development and structure of P. aeruginosa biofilms before examining each EPS as a potential therapeutic target for combating pulmonary infection with P. aeruginosa in CF, with a particular focus on the current evidence for these emerging therapies and barriers to bringing these therapies into clinic.

Non-nutritive bioactive components in maternal milk and offspring development: a scoping review.

Lactation is a critical time in mammalian development, where maternal factors shape offspring outcomes. In this scoping review, we discuss current literature concerning maternal factors that influence lactation biology and highlight important associations between changes in milk composition and offspring outcomes. Specifically, we explore maternal nutritional, psychosocial, and environmental exposures that influence non-nutritive bioactive components in milk and their links to offspring growth, development, metabolic, and behavioral outcomes. A comprehensive literature search was conducted using the Preferred Reporting Items for Systematic Reviews and Meta-Analysis Extension for Scoping Reviews (PRISMA-ScR) guidelines. Predetermined eligibility criteria were used to analyze 3,275 papers, and the final review included 40 primary research articles. Outcomes of this review identify maternal obesity to be a leading maternal factor influencing the non-nutritive bioactive composition of milk with notable links to offspring outcomes. Offspring growth and development are the most common modes of programming associated with changes in non-nutritive milk composition due to maternal factors in early life. In addition to discussing studies investigating these key associations, we also identify knowledge gaps in the current literature and suggest opportunities and considerations for future studies.

Pseudomonas aeruginosa aggregation and Psl expression in sputum is associated with antibiotic eradication failure in children with cystic fibrosis.

We previously demonstrated that P. aeruginosa isolates that persisted in children with cystic fibrosis (CF) despite inhaled tobramycin treatment had increased anti-Psl antibody binding in vitro compared to those successfully eradicated. We aimed to validate these findings by directly visualizing P. aeruginosa in CF sputum. This was a prospective observational study of children with CF with new-onset P. aeruginosa infection who underwent inhaled tobramycin eradication treatment. Using microbial identification passive clarity technique (MiPACT), P. aeruginosa was visualized in sputum samples obtained before treatment and classified as persistent or eradicated based on outcomes. Pre-treatment isolates were also grown as biofilms in vitro. Of 11 patients enrolled, 4 developed persistent infection and 7 eradicated infection. P. aeruginosa biovolume and the number as well as size of P. aeruginosa aggregates were greater in the sputum of those with persistent compared with eradicated infections (p < 0.01). The amount of Psl antibody binding in sputum was also greater overall (p < 0.05) in samples with increased P. aeruginosa biovolume. When visualized in sputum, P. aeruginosa had a greater biovolume, with more expressed Psl, and formed more numerous, larger aggregates in CF children who failed eradication therapy compared to those who successfully cleared their infection.

Comparison of methods for pre-processing, exosome isolation, and RNA extraction in unpasteurized bovine and human milk.

Milk is a highly complex, heterogeneous biological fluid that contains non-nutritive, bioactive extracellular vesicles called exosomes. Characterization of milk-derived exosomes (MDEs) is challenging due to the lack of standardized methods that are currently being used for milk pre-processing, storage, and exosome isolation. In this study, we tested: 1) three pre-processing methods to remove cream, fat, cellular debris, and casein proteins from bovine milk to determine whether pre-processing of whole milk prior to long-term storage improves MDE isolations, 2) the suitability of two standard exosome isolation methods for MDE fractionation, and 3) four extraction protocols for obtaining high quality RNA from bovine and human MDEs. MDEs were characterized via Transmission Electron Microscopy (TEM), Nanoparticle Tracking Analysis (NTA), and western immunoblotting for CD9, CD63, and Calnexin protein markers. We also present an optimized method of TEM sample preparation for MDEs. Our results indicate that: 1) Removal of cream and fat globules from unpasteurized bovine milk, prior to long-term storage, improves the MDE yield but not purity, 2) Differential ultracentrifugation (DUC) combined with serial filtration is better suited for bovine MDE isolation compared to ExoQuick (EQ) combined with serial filtration, however both methods were comparable for human milk, and 3) TRIzol LS is better suited for RNA extraction from bovine MDEs isolated by EQ and DUC methods. 4) TRIzol LS, TRIzol+RNA Clean and Concentrator, and TRIzol LS+RNA Clean and Concentrator methods can be used for RNA extractions from human MDEs isolated by EQ, yet the TRIzol LS method is better suited for human MDEs isolated by DUC. The QIAzol + miRNeasy Mini Kit produced the lowest RNA yield for bovine and human MDEs.