Potential Value of Qiagen and PrepIT•MAX Kits in Extraction of Mycobacterial DNA From Presumptive Tuberculosis Archived Formalin-Fixed Paraffin-Embedded Tissues

Abstract

Background: DNA analysis has potential for screening for and diagnosing a variety of conditions as well as the characterization of various pathogens for many purposes including to identify genetic disorders and mutations, study genetic diversity, and establish evolutional trends. Methods:Our study compared the performance of 2 DNA extraction kits: Qiagen and prepITMAX. The study tested 160 formalin-fixed paraffin-embedded (FFPE) human tissue samples that had been collected at Muhimbili National Hospital (MNH) between 2010 and 2016. For each sample, DNA extraction was performed using both the Qiagen and prepITMAX kits followed by polymerase chain reaction (PCR) tests to target the RNA polymerase gene and gel electrophoresis.Results: The findings showed that the Qiagen was 3 times superior to the prepITMAX kit in successfully extracting mycobacterial DNA from presumptive tuberculosis (TB) FFPE tissues. Of the 160 previously Ziehl-Neelsen stain-negative Mycobacterium tuberculosis suspicious tissue samples, 12 (7.5%) tested positive with the PCR. Of the 12 PCR-detected positive samples, 8 (66.7%) yielded positive results with the Qiagen kit only and 4 (33.3%) yielded positive results with both Qiagen and prepITMAX kits. Additionally, 10 (83.3%) came from well-formed granuloma, 1 (8%) from caseous necrosis, and 1 (8.3%) Langhans-type giant cells endorsing their potential for housing infection such as TB adenitis.Conclusions: A combination of molecular techniques, microscopy, and pathological features increases detection of M. tuberculosis from FFPE tissues. Both the Qiagen and the prepITMAX DNA extraction kits have shown a remarkable capability for extracting DNA from M. tuberculosis, although examination of FFPE tissues is not an intended use for the prepITMAX, according to the manufacturer. In resource-limited countries, however, these kits may complement each other. We recommend further studies for validation and optimization, which includes the cost effectiveness of prepITMAX extraction kit to advocate for its use in extraction of mycobacterial DNA from FFPE tissues.

Drug Resistance and Molecular Characteristics of Mycobacterium tuberculosis: A Single Center Experience

“…It is often necessary to obtain tissue sample for diagnosis and management of complications through surgery or by the use of computerized tomography (CT), magnetic resonance imaging (MRI), and endoscopy to greatly assist in the anatomic localization of EPTB [25]. FFPE sample can be stored for years, extracting MTB DNA from samples in a short period of time, but require additional deparaffinization steps resulting in partial degradation of the extracted DNA [26]. FFPE specimen is an important pathological specimen for the diagnosis of TB [27].…”