Novel Approach for Improving Sensitivity of Microscopic Detection of Acid-Fast Bacilli (AFB) by Use of the ReaSLR Method

Abstract

The ReaSLR methodology developed for sputum processing is a novel, low-cost, and simple technique that has improved the sensitivity of smear microscopy for the diagnosis of tuberculosis (TB). Sample processing consists of rapid liquefaction of the sputum specimen with the ReaSLR reagent, followed by syringe filtration, concentration by centrifugation, and use of the sediment for smear microscopy. The performance of the ReaSLR kit was evaluated on 150 sputum samples and was compared with that of the modified Petroff method for sputum decontamination and concentration. Ziehl-Neelsen staining was performed for smear microscopy after processing by these two techniques; simultaneously, culture on Lowenstein-Jensen (LJ) medium was done to evaluate the two methods. The efficiency of smear microscopy was 18/150 (12%) with the modified Petroff method compared to 47/150 (31.33%) with the ReaSLR method, and this difference was statistically significant (P < 0.001). The ReaSLR method for smear microscopy demonstrated a sensitivity and specificity of 90.47% and 91.6%, respectively, whereas the modified Petroff method showed a sensitivity and specificity of 40.47% and 99.07%, respectively, compared to those of culture, which was used as the gold standard. With the newer ReaSLR method, the kappa coefficient () was 0.8, which implies an excellent positive agreement. The ReaSLR method was found to be more sensitive than the conventional method for sputum smear microscopy. The newer ReaSLR method holds promise for adoption in TB control programs across the globe, as it was found suitable for the laboratory diagnosis of pulmonary TB. Further large-scale studies are needed to evaluate other aspects of this method.T uberculosis (TB), an airborne infectious disease, continues to be a major global health problem. About 9 million new cases of TB are diagnosed and close to 2 million people die of the disease each year. The majority of cases occur in Africa (30%) and Asia (55%), with India and China accounting for about 35% of all such cases (1). With the emergence of multidrug-resistant tuberculosis (MDR-TB) (around 0.4 to 0.5 million cases each year), the scenario is becoming more challenging (2).The threat of TB is alarming, and the disease can be controlled only by an effective TB control program. Early case detection is a crucial step in the control of the disease, and this relies solely on the detection of acid-fast bacilli (AFB) in clinical samples. Sputum microscopy with Ziehl-Neelsen (ZN) staining has been a reliable tool for decades, especially in the setting of developing countries, but it has only a 40 to 60% sensitivity for the diagnosis of pulmonary TB (3). It has the advantages of low cost, a need for minimal expertise, rapid processing, and simplicity but has the disadvantage of a lack of sensitivity (4, 5, 6). The methods of decontamination, liquefaction, and concentration before the ZN staining improve the yield and allow identification of the AFB in the sputum sample (7, 8). There is a need to upgrade the existing simp...

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