Determination of the biocidal effect of sodium hypochlorite in smear microscopy for diagnosis of tuberculosis in absence of safety cabinet

Abstract

Pulmonary tuberculosis (PTB) is the most common presentation of tuberculosis (TB) worldwide. To diagnose PTB, direct sputum smear microscopy is usually used because it is technically simple, non-invasive and cheap. The reliability of this technique for the diagnosis of TB has frequently been questioned due to its low sensitivity. Liquefaction and concentration of sputum using sodium hypochlorite (NaOCl-Jik bleach) before preparing a smear has been used to increase sensitivity in many settings. Results from these studies have, however, been conflicting. Nevertheless, the protective value of NaOCl in processing of sputum smears for diagnosis of TB particularly in places where adequate safety measures are lacking has not been ascertained. This study was therefore designed to determine the biocidal effect of NaOCl in terms of appropriate concentration and time of exposure required to completely sterilize the speciments and making recommendations for use in absence of a safety cabinet. Sputum specimens of over 10ml from newly diagnosed patients were processed for microscopy and culture. Two direct smears were prepared from each specimen, one stained with flourescence microscopy (FM) and the other using Ziehl Neelsen (ZN) method. The remaining sputum was divided into five equal portions after homogenization. One portion was processed for culture. The remaining four portions were treated with NaOCl at 3.5% (household bleach jikTM) and 5% (commercial bleach) concentrations. Two specimens, of each concentration were centrifuged and the other two sedimented. Smears were prepared from all sputum specimens that tested smear negative by direct FM and ZN, and examined by FM and ZN methods. Sedimented and centrifuged sputum specimens were cultured at various intervals ranging from ˝ hrs to 24 hours, to determine their viability upon which the biocidal effect of NaOCl was evaluated. The sensitivity of direct ZN was 66% and that of direct FM 77.5%. ZN sensitivity was 42.9% and 27.1%, respectively, after centrifugation and sentimentation with 3.5% NaOCl, while it was 37.1% and 10%, respectively, after centrifugation and sendimentation with 5%. The number of specimens that were positive with ZN after treatment with 3.5 was significantly higher than those treated with 5% after centrifugation and sedimentation ( = 0.0014;  = 0.001), respectively. There was also significant reduction in debris in sputum giving clear field defination under the microscope. The 3.5% NaOCl showed total immobilization of MTB, between 1 hr and 3 hr of exposure whereas 5% NaOCl showed total sterilization after 15 hrs. Use of appropriate concentration of NaOCl and appropriate time of exposure, may be recommended as a safety measure during smear microscopy for AFB in setups where a safety cabinet is not available.