Tuberculosis is a potentially deadly infectious disease caused by the Mycobacterium tuberculosis (M. tuberculosis). Tuberculosis is diagnosed by proving the M. tuberculosis in sputum samples based on the results of acid-resistant staining, culture, and nucleic acid amplification tests. However, there is a report that the detection rate of M. tuberculosis is low in acid-resistant staining using tissue specimens. It has been suspected that the cause is a potential loss of acid resistance by the organic solvents used for tissue specimen preparation. Therefore, this study was pursued to find out if Gram staining and fluorescent staining in addition to acid-resistant staining would be helpful in diagnosing tuberculosis. We used four tissue (lung, small intestine, large intestine, and lymph node) samples with chronic granulomatous inflammation observed in HE staining and positive results in real-time PCR. These detection rates and staining properties were investigated through microscopic examination using the Ziehl-Neelsen, Gram, and Auramin rhodamine staining. In this studies, M. tuberculosis were observed by Ziehl-Neelsen, Gram, and Auramin rhodamine staining in all four samples. In the evaluation of clinical microbiology proficiency testing (CMPT), the Ziehl-Neelsen and Gram staining were the same result, but the Auramin rhodamine staining was relatively low. These data indicated that Gram staining is useful for detecting M. tuberculosis in formalin-fixed tissue specimens. Therefore, if the Ziehl-Neelsen and Gram staining are combined as the M. tuberculosis staining method in tissue specimens, a better direction may be provided for tuberculosis diagnosis.

Tuberculosis (TB) is a significant disease for both humans and animals worldwide. The genus Mycobacterium includes several species that cause TB disease in humans and other animals. Amongst the members of the Mycobacterium tuberculosis complex (MTC), M. tuberculosis is mainly a human pathogen, whereas M. bovis has a broad host range and is the principal agent responsible for TB in domestic and wild mammals. M. bovis also infects humans, causing zoonotic TB through ingestion, inhalation. M. bovis accounts for only a small percentage of the reported cases of TB in humans. In recent years, TB in farmed deer has become a disease as public health importance in several countries. Nowadays, there has been rapid outbreak of bovine TB in cattle and deer in Korea. Investigations are needed to elucidate the relative importance of M. bovis on TB incidence in humans, especially in Korea where bovine TB remains a problem. Also, the human sources as the cause of animal infection, M. tuberculosis from the farm workers also important for TB control of animals in Korea. Differentiation between the causative organisms may only be achieved by sophisticated laboratory methods involving bacteriological characteristics, biochemical properties, and routine resistance to pyrazinamide (PZA). M. bovis shows inherently resistant to PZA whereas M. tuberculosis is susceptible to PZA. In this study, we developed a multiplex-PCR assay based on a 12.7-kb fragment for the differential detection of M. bovis and M. tuberculosis. A total of 131 M. tuberculosis complex isolates were randomly obtained from cattle and deers that were PPD positive. they all yielded M. bovis. M. tuberculosis was not isolated from animals. and, a total of 25 M. tuberculosis complex isolates which is resistant to PZA were obtained from patient. PZA resistant MTC in humans was caused entirely by M. tuberculosis. The multiplex-PCR protocol was highly species-specific and time saving for identification of M. bovis and M. tuberculosis. This multiplex-PCR assay will be easily used as a routine monitoring tool in veterinary and medical laboratories.

Pulmonary tuberculosis is an important public health problem. Early diagnosis and treatment is necessary for successful patient outcomes. However, diagnosis of pulmonary tuberculosis is difficult in cases with an unusual presentation and often requires a lung biopsy. The tuberculosis polymerase chain reaction results in tissue sample have potential for early diagnosis and treatment initiation for pulmonary tuberculosis. The relatively low sensitivity limit the use of this test in the detection of the pulmonary tuberculosis.