Viral polyhedroses are very common diseases of insects. They were first identified as leading causes of losses in the silk industry. This heterogeneous group of diseases is characterized by the formation of crystals in infected cells that are called viral polyhedra or occlusion bodies and represent the infectious form of the viruses. Polyhedra have similar role in the infectious cycle of the two groups of viruses responsible for polyhedroses, the cypoviruses - members of the Reoviridae family - and the Baculoviridae. Polyhedra embed virus particles within infected cells in a robust crystalline matrix that protects viral infectivity after release in the environment. Upon ingestion by a new host, crystals dissolve readily thereby releasing the infectious particles to initiate a new viral cycle. Owing to their unique molecular organization, these atypical infectious forms have long intrigued virologists and biochemists alike. They attracted particular interest because of the in vivo crystallization process and the contrast between rapid release upon ingestion and extreme stability. It is only recently that novel approaches and technologies allowed the structure determination of such tiny crystals by X-ray crystallography. Cypovirus and baculovirus polyhedra share the same role in the virus cycle, the same crystalline lattice with a cubic centered symmetry, and matrix proteins called polyhedrins of similar sizes. However, their building blocks differ by their folds and packing in polyhedra. The two classes of polyhedra therefore harbour distinct molecular architectures and appear to have emerged independently in the virosphere. The role of tyrosine clusters in polyhedra dissolution and the use of molecular arms to achieve in vivo crystallization may thus represent striking cases of convergent evolution. This review summarizes our understanding of viral polyhedra with an emphasis on the recent structural studies. We also provide examples of biotechnological applications entailing structure-based engineering of polyhedra as novel types of crystalline microparticules.

Crystal-storing histiocytosis (CSH) is an uncommon histiocytic proliferation reported to involve diverse organs and tissues, but involvement of the central nervous system (CNS) is rare. In most cases CSH is identified in association with underlying lymphoproliferative, plasma cell diseases or rarely with various inflammatory or infectious conditions. CSH is characterized by the cytoplasmic accumulation of crystalline material in histiocytes, most commonly of kappa immunoglobulin light chain. We report a unique case of localized CSH involving the left cerebellum and caudal brain stem in a young man with a history of gout but without known lymphoproliferative or plasma cell disorders. Awareness of this entity is important diagnostically, but also to ensure appropriate management and follow-up, particularly in the absence of apparent underlying malignancy.

Clofazimine (CFZ) is an optically active, red-colored chemotherapeutic agent that is FDA approved for the treatment of leprosy and is on the World Health Organization\'s list of essential medications. Interestingly, CFZ massively accumulates in macrophages where it forms crystal-like drug inclusions (CLDIs) after oral administration of the drug in animals and humans. The analysis of the fluorescence spectra of CLDIs formed by resident tissue macrophages revealed that CFZ, when accumulated as CLDIs, undergoes a red shift in fluorescence excitation (from Ex: 540-570 to 560-600 nm) and emission (Em: 560-580 to 640-700 nm) signal relative to the soluble and free-base crystal forms of CFZ. Using epifluorescence microscopy, CLDI(+) cells could be identified, relative to CLDI(-) cells, based on a >3-fold increment in mean fluorescence signal at excitation 640 nm and emission at 670 nm. Similarly, CLDI(+) cells could be identified by flow cytometry, based on a >100-fold increment in mean fluorescence signal using excitation lasers at 640 nm and emission detectors >600 nm. CLDI\'s fluorescence excitation and emission was orthogonal to that of cell viability dyes such as propidium iodide and 4,6-diamidino-2-phenylindole dihydrochloride (DAPI), cellular staining dyes such as Hoechst 33342 (nucleus) and FM 1-43 (plasma membrane), as well as many other fluorescently tagged antibodies used for immunophenotyping analyses. In vivo, >85% of CLDI(+) cells in the peritoneal exudate were F4/80(+) macrophages and >97% of CLDI(+) cells in the alveolar exudate were CD11c(+). Most importantly, the viability of cells was minimally affected by the presence of CLDIs. Accordingly, these results establish that CFZ fluorescence in CLDIs is suitable for quantitative flow cytometric phenotyping analysis and functional studies of xenobiotic sequestering macrophages.

In mammals, highly lipophilic small molecule chemical agents can accumulate as inclusions within resident tissue macrophages. In this context, we characterized the biodistribution, chemical composition, and structure of crystal-like drug inclusions (CLDIs) formed by clofazimine (CFZ), a weakly basic lipophilic drug. With prolonged oral dosing, CFZ exhibited a significant partitioning with respect to serum and fat due to massive bioaccumulation and crystallization in the liver and spleen. The NMR, Raman, and powder X-ray diffraction (p-XRD) spectra of CLDIs isolated from the spleens of CFZ-treated mice matched the spectra of pure, CFZ hydrochloride crystals (CFZ-HCl). Elemental analysis revealed a 237-fold increase in chlorine content in CLDIs compared to untreated tissue samples and a 5-fold increase in chlorine content compared to CFZ-HCl, suggesting that the formation of CLDIs occurs through a chloride mediated crystallization mechanism. Single crystal analysis revealed that CFZ-HCl crystals had a densely packed orthorhombic lattice configuration. In vitro, CFZ-HCl formed at a pH of 4-5 only if chloride ions were present at sufficiently high concentrations (>50:1 Cl(-)/CFZ), indicating that intracellular chloride transport mechanisms play a key role in the formation of CLDIs. While microscopy and pharmacokinetic analyses clearly revealed crystallization and intracellular accumulation of the drug in vivo, the chemical and structural characterization of CLDIs implicates a concentrative, chloride transport mechanism, paralleling and thermodynamically stabilizing the massive bioaccumulation of a weakly basic drug.