Comparing microbiotas in the upper aerodigestive and lower respiratory tracts of lambs

Abstract Background Recently, the importance of the lung microbiota during health and disease has been examined in humans and in small animal models. Whilst sheep have been proposed as an appropriate large animal model for studying the pathophysiology of a number of important human respiratory diseases, it is clearly important to continually define the limits of agreement between these systems as new concepts emerge. In humans, it has recently been established that the lung microbiota is seeded by microbes from the oral cavity. We sought to determine whether the same was true in sheep. Results We took lung fluid and upper aerodigestive tract (oropharyngeal) swab samples from 40 lambs (7 weeks old). DNA extraction was performed, and the V2-V3 region of the 16S rRNA gene was amplified by PCR then sequenced via Illumina Miseq. Oropharyngeal swabs were either dominated by bacteria commonly associated with the rumen or by bacteria commonly associated with the upper aerodigestive tract. Lung microbiota samples did not resemble either the upper aerodigestive tract samples or reagent-only controls. Some rumen-associated bacteria were found in lung fluids, indicating that inhalation of ruminal bacteria does occur. We also identified several bacteria which were significantly more abundant in lung fluids than in the upper aerodigestive tract swabs, the most predominant of which was classified as Staphylococcus equorum. Conclusions In contrast to humans, we found that the lung microbiota of lambs is dissimilar to that of the upper aerodigestive tract, and we suggest that this may be related to physiological and anatomical differences between sheep and humans. Understanding the comparative physiology and anatomy underlying differences in lung microbiota between species will provide a foundation upon which to interpret changes associated with disease and/or environment

A sheep cannulation model for evaluation of nasal vaccine delivery

C1 - Journal Articles RefereedWe have developed and validated a novel model to investigate the efficacy of nasal vaccine delivery. Based on lymphatic cannulation of the tracheal lymph trunk of sheep, the model allows collection of lymph draining from the Nasal Associated Lymphoid Tissue. The model is suitable for determining both the amount of material that is absorbed into the lymphatic system, following intra-nasal delivery and the immune response that occurs following vaccination into the nasal area. The cell populations that track in this duct were phenotyped and found to be similar to those previously reported to be present in efferent lymph draining from peripheral lymph nodes. Following intra-nasal spray, we demonstrated that the amount of material recovered in draining lymph is only a very small fraction of the total delivered. Nevertheless, intra-nasal spraying of a vaccine could activate local immune cells. The method described will be invaluable for optimising intra-nasal delivery systems by allowing a separate optimisation of vaccine uptake and immune responses induction

Comparison on In Vitro Characterization of Fucospheres and Chitosan Microspheres Encapsulated Plasmid DNA (pGM-CSF): Formulation Design and Release Characteristics

Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a cytokine used in the treatment of serious conditions resulting from chemotherapy and bone marrow transplantation such as neutropenia and aplastic anemia. Despite these effects, GM-CSF has a very short biological half-life, and it requires frequent injection during the treatment. Therefore, the cytokine production is possible in the body with plasmid-encoded GM-CSF (pGM-CSF) coding for cytokine administered to the body. However, the selection of the proper delivery system for the plasmid is important. In this study, two different delivery systems, encapsulated plasmid such as fucoidan–chitosan (fucosphere) and chitosan microspheres, were prepared and the particle physicochemical properties evaluated. Fucospheres and chitosan microspheres size ranges are 151–401 and 376–681 nm. The zeta potential values of the microspheres were changed between 8.3–17.1 mV (fucosphere) and +21.9–28.9 mV (chitosan microspheres). The encapsulation capacity of fucospheres changed between 84.2% and 94.7% depending on the chitosan molecular weight used in the formulation. In vitro plasmid DNA release from both delivery systems exhibited slower profiles of approximately 90–140 days. Integrity of released samples was checked by agarose gel electrophoresis, and any additional band was not seen. All formulations were analyzed kinetically. The calculated regression coefficients showed a higher r2 value with zero-order kinetics. In conclusion, the characterizations of the microspheres can be modulated by changing the formulation variables, and it can be concluded that fucospheres might be a potential carrier system for the controlled delivery of GM-CSF encoding plasmid DNA