Negative staining of mycobacteria - A clue to the diagnosis in cytological aspirates: Two case reports

Large amounts of lipids present in the cell wall of mycobacteria render them impermeable to dyes used in routine stains. Special staining techniques like Ziehl-Neelsen (ZN), Auromine Rhodamine are needed to demonstrate them. However, they may appear as negatively stained ghosts in Leishman, Giemsa, Hematoxylin and Eosin stain (H and E) and Gram stained smears. Awareness of this feature prompts one to look for mycobacteria by special techniques, even in the absence of cytological features of tuberculous infection like epithelioid granulomas and caseous necrosis. In this paper, we aim to present two cases showing negatively stained images in the lymphnode aspirates of Human Immunodeficiency Virus (HIV) positive patients. A study has been done of two cases of HIV positive patients with cervical lymphadenopathy. Fine needle aspiration cytology (FNAC) was carried out for them from the enlarged lymph nodes which revealed purulent material. Smears of FNAC material were prepared for histopathological examination. Fixed smears were stained with Papanicolaou stain and air dried smears were stained with Giemsa and ZN-stain and studied. Smear study showed scant cellularity, cells composed of neutrophils, lymphocytes, plasma cells and macrophages. Background was necrotic. Giemsa stained smears showed, in addition to cells mentioned, negatively stained ghostly rod shaped structures in the cytoplasm of macrophages and also in the background. ZN-stain showed numerous acid fast bacilli. Both the aspirates were signed out as tuberculous lymphadenitis. In both the cases, cultures grew Mycobacterium avium Intracellulare. Classical cytological features of tuberculosis may not be present in immunocompromised patients and on the contrary there may be suppuration rich in neutrophils or sheets of histiocytes in tuberculosis. One may miss these cases if mycobacteria are not looked for specifically by special stains. Negatively stained ghost images of Tuberculous bacilli on different types of strains are a very helpful finding in such cases.