Rapid Detection of Mycobacteria in Inflammatory Necrotizing Granulomas From Formalin-Fixed, Paraffin-Embedded Tissue by PCR in Clinically High-Risk Patients With Acid-Fast Stain and Culture-Negative Tissue Biopsies

Abstract

A collection of inflammatory necrotizing granulomas (INGs) negative by acid-fast stain and culture (AFSC) were analyzed by polymerase chain reaction (PCR) for the presence of mycobacteria. Forty-two paraffin-embedded specimens with INGs were collected from patients at high risk for contracting tuberculosis. Twenty biopsies were positive and 22 were negative for mycobacteria by AFSC. Two universal primers specific for all mycobacteria were used to detected a 414 base pair (bp) fragment of 16S rRNA gene. Twenty of 20 biopsies were positive for mycobacteria by both AFSC and PCR (100%), whereas 19 of 22 biop-A resurgence in tuberculosis has become a very serious public health concern. 1 " 8 In 1992, there were 26,532 new cases in the United States. Georgia ranked 5th among the states with 950 new cases. Grady Memorial Hospital (GMH), a 1,200 bed inner-city hospital in Atlanta, Georgia, where this study took place, had 257 new cases of tuberculosis in 1992, a total higher than that for all but 25 states in the United States. These developments in tuberculosis infection have generated a sense of urgency to develop more efficient methods for detecting mycobacteria.

Application of RLEP Real-Time PCR for Detection of M. leprae DNA in Paraffin-Embedded Skin Biopsy Specimens for Diagnosis of Paucibacillary Leprosy

“…12 The PCR detecting the presence of M. leprae DNA in fixed tissues coupled with histopathology may help clinicians arrive at a more rapid and definitive diagnosis of the disease. 13 Recently, we have found that a nested PCR method that detects M. leprae DNA in patients' whole blood can be used for rapid and early diagnosis of leprosy. 14 In this study, we evaluated the use of a real-time PCR test targeting RLEP DNA sequence for adjunct diagnosis of PB leprosy in comparison to the nested PCR test 14 and also conventional histopathology for diagnosis of leprosy.…”

Detection of Mycobacterium tuberculosis Complex in Formalin-Fixed, Paraffin-Embedded Tissue Specimens with Necrotizing Granulomatous Inflammation by Strand Displacement Amplification

“…Previous studies have described the use of PCR on human FFPE specimens for detection of MTC DNA with promising results, but almost all of them used in-house PCR methods. [15][16][17][18][19] The BDProbeTec ET Direct Detection assay (Becton Dickinson, Sparks, MD, USA) is commercially available for the detection of MTC in clinical specimens. This assay is based on simultaneous strand displacement amplification of MTC-specific IS6110 target DNA of 95 bp and real-time detection by fluorescentlabeled probes.…”

Detection of <i>Mycobacterium leprae</i> in Ocular Tissues by Histopathology and Real-Time Polymerase Chain Reaction

“…Several investigators have used DNA amplification through PCR to amplify various genomic sequences of M. leprae to improve detection when low numbers of bacteria are present. Thus, PCR for identification of DNA that encodes M. leprae proteins of 65 kDa [16] , 18 kDa [17] , and repetitive sequences of M. leprae [18] have been suggested to be a more sensitive and specific method than the AFS examination [19] . Real-time polymerase chain reaction (RT-PCR) is more sensitive and precise for detecting DNA sequences from analyte compared to conventional PCR protocols.…”