Spatial Distribution of Mycobacterium tuberculosis mRNA and Secreted Antigens in Acid-Fast Negative Human Antemortem and Resected Tissue

BACKGROUND: The ability to detect evidence of Mycobacterium tuberculosis (Mtb) infection within human tissues is critical to the study of Mtb physiology, tropism, and spatial distribution within TB lesions. The capacity of the widely-used Ziehl-Neelsen (ZN) staining method for identifying Mtb acid-fast bacilli (AFB) in tissue is highly variable, which can limit detection of Mtb bacilli for research and diagnostic purposes. Here, we sought to circumvent these limitations via detection of Mtb mRNA and secreted antigens in human tuberculous tissue. METHODS: We adapted RNAscope, an RNA in situ hybridisation (RISH) technique, to detect Mtb mRNA in ante- and postmortem human TB tissues and developed a dual ZN/immunohistochemistry staining approach to identify AFB and bacilli producing antigen 85B (Ag85B). FINDINGS: We identified Mtb mRNA within intact and disintegrating bacilli as well as extrabacillary mRNA. Mtb mRNA was distributed zonally within necrotic and non-necrotic granulomas. We also found Mtb mRNA within, and adjacent to, necrotic granulomas in ZN-negative lung tissue and in Ag85B-positive bronchiolar epithelium. Intriguingly, we observed accumulation of Mtb mRNA and Ag85B in the cytoplasm of host cells. Notably, many AFB were negative for Ag85B staining. Mtb mRNA was observed in ZN-negative antemortem lymph node biopsies. INTERPRETATION: RNAscope and dual ZN/immunohistochemistry staining are well-suited for identifying subsets of intact Mtb and/or bacillary remnants in human tissue. RNAscope can identify Mtb mRNA in ZN-negative tissues from patients with TB and may have diagnostic potential in complex TB cases. FUNDING: Wellcome Leap Delta Tissue Program, Wellcome Strategic Core Award, the National Institutes of Health (NIH, USA), the Mary Heersink Institute for Global Health at UAB, the UAB Heersink School of Medicine

The catalytic evaluation of structured zeolite catalysts

ZSM-5 films, with thicknesses between 150 and 2300 nm, supported on 3 mm diameter alumina beads are characterised in terms of their catalytic activity and selectivity for tri-isopropylbenzene cracking and para-xylene isomerisation at 450°C. A reaction-diffusion model adequately represents the experimental data and is used to estimate intrinsic reactivity and diffusion properties of the zeolite films. Results show that the external activity, overall activity and the diffusivity increase with increasing film thickness. The variation in reactivity and diffusivity, when compared to physio-chemical data, are a result of structural defects and non-homogeneities of the zeolite film. The measured diffusivities are within an order of magnitude of those extrapolated from literature. Thick zeolite films have the largest Thiele modulus as the increase in zeolite film thickness dominates over the structural defects. The largest selectivity improvements are observed for thick zeolite films</p

ZSM-5 structured catalysts coated with silicalite-1

Alumina beads were coated with ZSM-5 films ranging from 150 nm to 2300 nm ill thickness. The ZSM-5 boated alumina beads were Subsequently hydrothermally treated in a silicalite-1 synthesis solution ill two steps Whereupon a dense silicalite-1 film was formed oil top of the ZSM-5 film. The materials were tested with two probe reactions and the reactivity was compared before and after coating with silicalite-1. As expected, the para-xylene (pX) isomerization reactivity showed no change for samples with and without the top layer of silicalite-1 for equal amounts Of zeolite. Surprisingly, the triisopropylbenzene (TIPB) conversion did not decrease after the silicalite-1 film was introduced. As measured by XPS, the aluminum concentration at the Surface of the uncalcined silicalite-1 film surface was lower compared to that at the Surface of the calcined ZSM-5 film. However, after calcination the concentration of aluminum was higher at the silicalite-1 film surface than at the ZSM-5 film Surface. These results Suggest that aluminum migrates from the ZSM-5 film into the silicalite-1 film during calcination and testing which results in all active top layer.</p