Rapidly growing interest in nanoparticle-mediated delivery of DNA and RNA to plants requires a better understanding of how nanoparticles and their cargoes translocate in plant tissues and into plant cells. However, little is known about how the size and shape of nanoparticles influences transport in plants and use of their cargoes, limiting development and deployment of nanotechnology in plant systems. Here, we employ nonbiolistically delivered DNA-modified gold nanoparticles (AuNP) spanning various sizes (5 -20 nm) and shapes (spheres and rods) to systematically investigate their transport following infiltration into Nicotiana benthamiana (Nb) leaves. Generally, smaller AuNPs demonstrate more rapid, higher, and longer-lasting levels of association with plant cell walls compared to larger AuNPs. We observe internalization of rod-shaped but not spherical AuNPs into plant cells, yet surprisingly, 10 nm spherical AuNP functionalized with small-interfering RNA (siRNA) are most efficient at siRNA delivery and inducing gene silencing in mature plant leaves. These results indicate the importance of nanoparticle size in efficient biomolecule delivery, and, counterintuitively, demonstrate that efficient cargo delivery is possible and potentially optimal in the absence of nanoparticle cellular internalization. Our results highlight nanoparticle features of importance for transport within plant tissues, providing a mechanistic overview of how nanoparticles can be designed to achieve efficacious bio-cargo delivery for future developments in plant nanobiotechnology.

No abstract

Bullous pemphigoid (BP) is an autoimmune blistering disease characterized by antibodies (IgG and IgE) targeting cell-substrate adhesion proteins. A variety of BP models suggest that autoantibody-dependent neutrophil degranulation is essential for blister formation. However, lesional biopsies reveal a predominance of eosinophils and few neutrophils. Our goal was to evaluate the role of antibodies and complement in eosinophil localization, degranulation, and split formation at the dermo-epidermal junction (DEJ) utilizing a human skin cryosection model of BP paired with a human eosinophilic cell line, 15HL-60. Expression of receptors for IgG (FcγRII), IgE (FcεRI), and complement (CR1 and CR3) was confirmed on 15HL-60 cells using flow cytometry. 15HL-60 expression of granule protein (eosinophil derived neurotoxin (EDN) and eosinophil peroxidase (EPO)) mRNA and their degranulation in vitro was confirmed using RT-PCR and ELISA, respectively. For cryosection experiments, BP or control sera or IgG and IgE antibodies purified from BP sera were utilized in combination with 15HL-60 cells ± fresh complement. Both BP serum and fresh complement were required for localization of 15-HL60 cells to the DEJ. Interestingly, eosinophil localization to the DEJ was dependent on IgG, but not IgE, and complement. However, no subepidermal split was observed. Additionally, the 15HL-60 cells did not degranulate under any experimental conditions and direct application of cell lysate to cryosections did not result in a split. Our observation that eosinophil localization to the DEJ is dependent on IgG mediated complement fixation provides additional insight into the sequence of events during the development of BP lesions.

Summary Objectives As individuals may be colonized with multiple strains of Staphylococcus aureus at different body sites, the objectives of this study were to determine whether S. aureus polyclonal colonization exists within one body niche and the optimal sampling sites and culture methodology to capture the diversity of S. aureus strains in community-dwelling individuals. Methods Swabs were collected from the nares, axillae, and inguinal folds of 3 children with community-associated S. aureus infections and 11 household contacts, all with known S. aureus colonization. S. aureus isolates were recovered from each body niche using 4 culture methods and evaluated for polyclonality using phenotypic and genotypic strain characterization methodologies. Results Within individuals, the mean (range) number of phenotypes and genotypes was 2.4 (1–4) and 3.1 (1–6), respectively. Six (43%) and 10 (71%) participants exhibited phenotypic and genotypic polyclonality within one body niche, respectively. Broth enrichment yielded the highest analytical sensitivity for S. aureus recovery, while direct plating to blood agar yielded the highest genotypic strain diversity. Conclusions This study revealed S. aureus polyclonality within a single body niche. Culture methodology and sampling sites influenced the analytical sensitivity of S. aureus colonization detection and the robustness of phenotypic and genotypic strain recovery.

Vitamin D promotes epithelial immunity by upregulating antimicrobial peptides (AMPs), including LL-37, that have bactericidal activity against Staphylococcus aureus. We found that children with vitamin D deficiency or insufficiency [25(OH)D <30 ng/mL] were more likely to present with recurrent, rather than primary, S. aureus skin or soft tissue infection (SSTI). Vitamin D sufficiency may be one of a myriad of host and environmental factors that can be directly impacted to reduce the frequency of S. aureus SSTI.