Comparison of C ₁₈ -Carboxypropylbetaine and Standard N -Acetyl- l -Cysteine-NaOH Processing of Respiratory Specimens for Increasing Tuberculosis Smear Sensitivity in Brazil

Abstract

Techniques to improve the sensitivity of smear microscopy would facilitate early tuberculosis (TB) diagnosis and disease control, especially in low-income countries where the positive predictive value is high. C 18 -carboxypropylbetaine (CB-18) is a zwitterionic detergent that helps to compensate for the innate buoyancy of mycobacteria, potentially enhancing recovery by centrifugation. Previous data suggest that CB-18 may increase the sensitivity of smear, culture, and molecular amplification diagnostic testing. The goal of the present study was to evaluate if the sensitivity of the smear technique using light microscopy could be improved by treating respiratory samples with CB-18. In the first phase, respiratory specimens were collected consecutively from patients with suspected pulmonary tuberculosis in a tertiary-care hospital in Rio de Janeiro, Brazil (236 specimens were analyzed). After protocol modifications, another 120 respiratory specimens were evaluated. The standard technique was N-acetyl-L-cysteine with sodium hydroxide (NALC-NaOH) treatment, smear concentration with centrifugation, and Ziehl-Neelsen staining. Culture on Löwenstein-Jensen slants was performed on all specimens for use as the "gold standard." No specimens from patients undergoing active TB treatment were included. The initial protocol for CB-18 processing resulted in a sensitivity of 59.6% and specificity of 96.8% compared to standard processing with a sensitivity of 66.0% and specificity of 96.8%. Using the modified protocol, the sensitivity of CB-18 increased to 71.4% with a specificity of 97.0% versus standard processing with a sensitivity of 61.9% and a specificity of 99.0%. The diagnostic yield of acid-fast bacillus smear with CB-18 in the absence of fluorescence microscopy and PCR compared to standard processing with NALC-NaOH was not significantly different, although the power to detect a difference by the modified assay was low.Tuberculosis (TB) remains one of the deadliest diseases worldwide, with an estimated 8.4 million new cases and over 2 million deaths each year (21). Smear microscopy is currently the most feasible microbiological method for diagnosis of pulmonary TB in developing countries due to its rapidity, low cost, and high positive predictive value for Mycobacterium tuberculosis (3). However, sputum smear microscopy with ZiehlNeelsen staining is only 60 to 70% sensitive for the diagnosis of pulmonary TB compared with sputum culture (2, 8). Among human immunodeficiency virus-infected persons the sensitivity of sputum Ziehl-Neelsen smears has been reported to be even lower (2, 6, 10). Delayed or missed diagnoses contribute to M. tuberculosis transmission and mortality due to TB (9, 11).Improvements in the sensitivity of smear microscopy would allow earlier diagnosis. TB programs could then optimize disease control from a public health standpoint as well as improve individual patient management (20). In previously published reports, homogenization and concentration of respiratory specimens by centrifugation have been ...

Comparison of NALC-NAOH processing method with C18-carboxy propyl betaine method for the detection of mycobacterium in sputum

“…11 Moreover it is a zwitter ionic detergent that can be taken up by viable mycobacteria, making them denser and thereby increasing their recovery by centrifugation. 7 Our results are in parallel with Charles G. thornton 1998, K. F. Laserson 2005. 12,6 It has been shown CB-18 improves the recovery of mycobacteria other than tuberculosis among human respiratory specimens and also among animal products such as milk.…”

Colorimetric microwell plate reverse-hybridization assay for Mycobacterium tuberculosis detection

“…Thus, the false positive results may also be due to the presence of non-cultivable bacteria in the specimen or to the high sensitivity of the PCR assay, which can even identify asymptomatic individuals infected with tuberculous bacilli (de Lassence et al 1992, Chakravorty et al 2005. Another important aspect of these false-positive results may come from the widely-used definition of positivity based on microbiological rather than clinical evidence (Scott et al 2002, Piersimoni & Scarparo 2003.…”

Utility of Universal Sample Processing Methodology, Combining Smear Microscopy, Culture, and PCR, for Diagnosis of Pulmonary Tuberculosis

“…The inclusion of a concentration step enhanced the sensitivity of the direct smear method, from 54 to 57% to 63 to 80% (5,9). The N-acetyl L-cysteine (NALC)-NaOH concentration method has shown an improved sensitivity of ϳ66 to 83% (8,16). However, a conflicting report suggests that the overall diagnostic sensitivity of smear microscopy did not increase with sputum liquefaction and concentration (21).…”