Utility of Conventional ZN Staining and Modified Bleach Techniques in Detection of Acid Fast Bacilli from Aspirates of Suspected Cases of Tuberculous Lymphadenitis

Abstract

Introduction: Tuberculosis (TB) is a significant public health problem in many developing countries, including India. Diagnosing tuberculous lymphadenitis, in particular, poses challenges as it requires the demonstration of Acid-Fast Bacilli (AFB) using the conventional Ziehl-Neelsen (ZN) technique. The Modified Bleach Technique, which involves liquefying the aspirated specimen with sodium hypochlorite followed by centrifugation, has been described to enhance the yield of AFB, improving sensitivity and diagnostic accuracy. Aim: To highlight the role of the bleach concentration method compared to conventional ZN staining in detecting AFB in Fine-Needle Aspiration (FNA) material from clinically suspected cases of tuberculous lymphadenitis. Materials and Methods: A prospective cross-sectional study was conducted at the Department of Pathology, MVJ Medical College and Research Hospital, Bengaluru, Karnataka, India from November 2020 to December 2022. Fine needle aspiration material from 100 cases of clinically suspected tuberculous lymphadenitis was included. In each case, part of the aspirate was processed for ZN staining, while the remaining material was used for modified bleach staining and Cartridge Based Nucleic Acid Amplification Test (CBNAAT). Patient data, including age, sex, duration and site of swelling, nature of aspirate, and cytomorphological diagnosis, were documented. The data was entered into a Microsoft excel spreadsheet and analysed using the Chi-square test, with a p-value<0.05 considered statistically significant. Results: The age range of the study participants was 1-80 years. Out of 100 cases, 13 were excluded due to inadequate material. The most common lymph node site was cervical 69 (79.3%), followed by axillary 8 (9.1%) and supraclavicular 6 (6.89%). Among the 87 cases, reactive lymphadenitis was diagnosed in 36 (41.3%), suppurative lymphadenitis in 12 (13.8%), necrotising granulomatous lymphadenitis in 12 (13.8%), granulomatous lymphadenitis in 15 (17.3%), and necrotising lymphadenitis in 12 (13.8%). Out of the 100 cases included in the study, 32 cases were confirmed as tubercular lymphadenitis by CBNAAT. Among these 32 cases, conventional ZN stain was positive for AFB in only 3 cases (9%), whereas the Modified bleach stain was positive in 16 cases (50%). The results were compared with CBNAAT findings, and the modified bleach technique showed sensitivity (59.4%), specificity (100%), Positive Predictive Value (PPV) (100%), Negative Predictive Value (NPV) (80%), and Accuracy (85%). Conclusion: The Modified bleach method of ZN staining is a simple, safe, inexpensive, and easy-to-perform technique. With the bleach method, AFB was easily visible and detectable in the majority of AFB-positive cases. Additionally, the bleach method preserved the morphology of AFB better. It improves the microscopic detection of AFB, leading to a definitive diagnosis of TB on cytology.