sodium-hypochlorite and Tuberculosis

Background: Sputum smear microscopy (SSM) is a screening test used to diagnose tuberculosis (TB); however, its performance and sensitivity are relatively low, which can lead to false negatives. We designed a cross-sectional study to estimate the performance of SSM that includes a pretreatment based on sputum digestion with bleach (sodium hypochlorite) for the diagnosis of TB. Methods: We evaluated 73 sputum samples from patients with a diagnosis of TB confirmed by the Xpert MTB/RIF test and 114 samples from patients without TB. We performed sputum digestion using a 5% sodium hypochlorite solution, centrifuged at 2000 rpm for 15 min. We prepared smears for direct and bleach-treated SSM and used Ziehl−Neelsen staining. Results: The bleach-treated SSM obtained absolute identification of the cases of TB confirmed by the Xpert test, compared to 95.9% identified by the direct smear method (without bleach treatment). We also found a significant increase (p < 0.001) in the recovery of acid-fast bacilli (AFB) obtained by the bleach-treated SSM (293.8 ± 215.1 AFB) compared to the direct SSM method (222.9 ± 195.5 AFB). The AUC of the bleach-treated SSM and direct SSM was 100% and 96.6%, respectively. Conclusion: The bleach-treated SSM performs better than the direct SSM in identifying AFB and increasing the bacillary count in the sputum samples.

Topics: Cross-Sectional Studies; Humans; Microscopy; Mycobacterium tuberculosis; Peru; Sensitivity and Specificity; Sodium Hypochlorite; Sputum; Tuberculosis; Tuberculosis, Pulmonary

In Ethiopia, the most widely used disinfectant is 5% Hypochlorites. However, Ethiopian national health safety and infection prevention guideline recommendation on the use of bleach is not consistent and varying from 0.1%-4%. The purpose of this study was therefore to assess the effective time-concentration relationship of sodium hypochlorite against Mycobacterium tuberculosis complex isolates in the absence of any organic load.. This experimental study was conducted in Bahir Dar Regional laboratory from February-June 2013. Test suspensions of 1.5 X 10(8) CFU/ml prepared using normal saline containing 0.5% tween 80. From 5% stock, 0.1%, 0.5%, 1% and 2% bleach was prepared. A 1ml of test strain suspension and 1ml of bleach mixed and allowed to stand until the specified time achieved, neutralized by 48 ml phosphate buffer. 100µl from the diluted sediment were spread on two L-J mediums and incubated at 37°C for 8 weeks.. When 0. 1% bleach was used for 10 min, majority 11/20 of isolates showed 3 x 10(3) CFU/ml growth (ME = 4.4) which was inefficient. However, when the time increased, the log10 reduction was acceptable, ME >5 and it was effective. The bleach solution containing 0.5% and above was effective in all respective times. In this study, there is no difference observed in the tuberculocidal activity of bleach against resistant and sensitive strains.. Our study showed that in the absence of any organic load, 0.1% bleaches over 15 min and 0.5% bleaches over 10 min was found to be tuberculocidal.

Topics: Disinfectants; Drug Resistance, Bacterial; Ethiopia; Health Facilities; Humans; Mycobacterium tuberculosis; Practice Guidelines as Topic; Sodium Hypochlorite; Time Factors; Tuberculosis

Mawenzi Regional Hospital, northern Tanzania.. To determine the value of light-emitting diode (LED) microscopy in diagnosing tuberculosis (TB) on bleach-treated and direct sputum smears.. Sputum samples were collected from patients suspected of pulmonary TB who presented consecutively at the laboratory for smear evaluation between December 2009 and February 2010. Four smears were prepared from each specimen: conventional Ziehl-Neelsen (ZN), direct auramine, bleach centrifugation and bleach short sedimentation auramine smears. A light microscope was used to examine ZN smears and an LED fluorescent microscope to examine auramine-stained smears.. Of the 267 sputum samples examined, respectively 78 (29%), 62 (23%), 74 (28%) and 48 (18%) were acid-fast bacilli (AFB) positive by the bleach centrifugation, bleach short sedimentation, direct auramine and ZN methods. Bleach centrifugation identified 30 (11%) more positives than ZN microscopy (P < 0.001), but was not superior to the direct auramine method (P = 0.46), which yielded 26 (10%) more positives than ZN microscopy (P < 0.001). Fluorescent LED required a shorter smear reading time (1.5 min on average), while the light microscope took 4 min (P < 0.001).. Fluorescent LED microscopy with direct smear preparation is rapid and effective. Further studies are needed to ascertain its performance under routine conditions.

Topics: Bacteriological Techniques; Benzophenoneidum; Centrifugation; Humans; Microscopy, Fluorescence; Mycobacterium tuberculosis; Sodium Hypochlorite; Sputum; Staining and Labeling; Tanzania; Time Factors; Tuberculosis

The diagnosis of tuberculosis is seriously hampered in the absence of standard biosafety laboratory facilities for specimen concentration and Mycobacterium tuberculosis culture. Within a laboratory twinning arrangement, heat-fixed direct smear and sediment from 74 bleach-processed and 20 non-processed specimens from Cumura Hospital, Guinea-Bissau, were sent to Lisbon for molecular evaluation of rifampicin resistance. Sequence analysis of a 369 base-pair rpoB locus detected 3.2% (3/94) resistant specimens. To our knowledge, this represents the first report on the molecular analysis of M. tuberculosis from bleach-processed sputum, an alternative to current diagnostic practice in low-resource settings.

Topics: Antitubercular Agents; Bacterial Proteins; Base Sequence; Capacity Building; Clinical Laboratory Techniques; DNA-Directed RNA Polymerases; Guinea-Bissau; Humans; Laboratories; Microbial Sensitivity Tests; Mycobacterium tuberculosis; Portugal; Rifampin; Sequence Analysis; Sodium Hypochlorite; Specimen Handling; Sputum; Tuberculosis

Sputum microscopy is the only diagnostic for tuberculosis (TB) available at peripheral levels of health service in resource-poor countries. Its sensitivity is reduced in high HIV-prevalence settings. Sodium hypochlorite (NaOCl) specimen sedimentation prior microscopy and light-emitting diode (LED)-fluorescence microscopy (FM) can individually improve performance of microscopy. This study aimed to evaluate the performance of combined LED-FM and NaOCl sputum sedimentation for TB detection at peripheral level of health services.. A prospective study was conducted in an urban health clinic in Nairobi, Kenya. Three sputum specimens were collected over 2 days from consecutive TB suspects. Smears were prepared and stained with auramine O and Ziehl-Neelsen (ZN) methods. Bleach (3.5%) was added to the remaining specimen before overnight sedimentation at room temperature. Auramine O staining was performed on smears of sediment. A 4(th) specimen was collected for TB culture. Auramine smears were read under the same microscope as used for ZN smears, but equipped with the LED FluoLED™ fluorescence illuminator.. 497 patients were included, and 1394 specimens collected. The yield of positive specimen was significantly increased after NaOCl sedimentation (24.9%) compared to direct LED-FM (20.6%) and direct ZN (20.3%). In detecting smear-positive patients, sensitivity was 78.5% for LED-FM after NaOCl sedimentation compared to 73.2% and 72.0% for direct LED-FM (P = 0.06) and direct ZN (P = 0.06), respectively. Specificity was 87.8% for LED-FM after NaOCl sedimentation compared to 96.7% and 95.9% for direct LED-FM (P<0.01) and direct ZN (P<0.01), respectively. Inter-reading agreement (kappa = 0.7) and technicians' acceptability were good.. NaOCl sedimentation did not improve the performance of LED-FM in the diagnosis of pulmonary TB at peripheral health service level.

Topics: Adult; Female; Humans; Male; Microscopy, Fluorescence; Middle Aged; Prospective Studies; Sodium Hypochlorite; Specimen Handling; Tuberculosis; Young Adult

Bleach-sedimentation may improve microscopy for diagnosing tuberculosis by sterilising sputum and concentrating Mycobacterium tuberculosis. We studied gravity bleach-sedimentation effects on safety, sensitivity, speed and reliability of smear-microscopy.. This blinded, controlled study used sputum specimens (n = 72) from tuberculosis patients. Bleach concentrations and exposure times required to sterilise sputum (n = 31) were determined. In the light of these results, the performance of 5 gravity bleach-sedimentation techniques that sterilise sputum specimens (n = 16) were compared. The best-performing of these bleach-sedimentation techniques involved adding 1 volume of 5% bleach to 1 volume of sputum, shaking for 10-minutes, diluting in 8 volumes distilled water and sedimenting overnight before microscopy. This technique was further evaluated by comparing numbers of visible acid-fast bacilli, slide-reading speed and reliability for triplicate smears before versus after bleach-sedimentation of sputum specimens (n = 25). Triplicate smears were made to increase precision and were stained using the Ziehl-Neelsen method.. M. tuberculosis in sputum was successfully sterilised by adding equal volumes of 15% bleach for 1-minute, 6% for 5-minutes or 3% for 20-minutes. Bleach-sedimentation significantly decreased the number of acid-fast bacilli visualised compared with conventional smears (geometric mean of acid-fast bacilli per 100 microscopy fields 166, 95%CI 68-406, versus 346, 95%CI 139-862, respectively; p = 0.02). Bleach-sedimentation diluted paucibacillary specimens less than specimens with higher concentrations of visible acid-fast bacilli (p = 0.02). Smears made from bleach-sedimented sputum were read more rapidly than conventional smears (9.6 versus 11.2 minutes, respectively, p = 0.03). Counting conventional acid-fast bacilli had high reliability (inter-observer agreement, r = 0.991) that was significantly reduced (p = 0.03) by bleach-sedimentation (to r = 0.707) because occasional strongly positive bleach-sedimented smears were misread as negative.. Gravity bleach-sedimentation improved laboratory safety by sterilising sputum but decreased the concentration of acid-fast bacilli visible on microscopy, especially for sputum specimens containing high concentrations of M. tuberculosis. Bleach-sedimentation allowed examination of more of each specimen in the time available but decreased the inter-observer reliability with which slides were read. Thus bleach-sedimentation effects vary depending upon specimen characteristics and whether microscopy was done for a specified time, or until a specified number of microscopy fields had been read. These findings provide an explanation for the contradictory results of previous studies.

Topics: Bacteriological Techniques; Centrifugation; Disinfectants; Disinfection; Humans; Microscopy; Mycobacterium tuberculosis; Observer Variation; Reproducibility of Results; Sensitivity and Specificity; Sodium Hypochlorite; Specimen Handling; Sputum; Time Factors; Tuberculosis

Diagnostic options for pulmonary tuberculosis in resource-poor settings are commonly limited to smear microscopy. We investigated whether bleach concentration by sedimentation and sputum cytology analysis (SCA) increased the positivity rate of smear microscopy for smear-positive tuberculosis.. We did a prospective diagnostic study in a Médecins Sans Frontières-supported hospital in Mindouli, Republic of Congo. Three sputum samples were obtained from 280 consecutive pulmonary tuberculosis suspects, and were processed according to WHO guidelines for direct smear microscopy. The remainder of each sputum sample was homogenised with 2.6% bleach, sedimented overnight, smeared, and examined blinded to the direct smear result for acid-fast bacilli (AFB). All direct smears were assessed for quality by SCA. If a patient produced fewer than three good-quality sputum samples, further samples were requested. Sediment smear examination was performed independently of SCA result on the corresponding direct smear. Positivity rates were compared using McNemar's test.. Excluding SCA, 43.2% of all patients were diagnosed as positive on direct microscopy of up to three samples. 47.9% were diagnosed on sediment microscopy, with 48.2% being diagnosed on direct microscopy, sediment microscopy, or both. The positivity rate increased from 43.2% to 47.9% with a case definition of one positive smear (≥1 AFB/100 high power fields) of three, and from 42.1% to 43.9% with two positive smears. SCA resulted in 87.9% of patients producing at least two good-quality sputum samples, with 75.7% producing three or more. Using a case definition of one positive smear, the incremental yield of bleach sedimentation was 14/121, or 11.6% (95% CI 6.5-18.6, p = 0.001) and in combination with SCA was 15/121, or 12.4% (95% CI 7.1-19.6, p = 0.002). Incremental yields with two positive smears were 5/118, or 4.2% (95% CI 1.4-9.6, p = 0.062) and 7/118, or 5.9% (95% CI 2.4-11.8, p = 0.016), respectively.. The combination of bleach sedimentation and SCA resulted in significantly increased microscopy positivity rates with a case definition of either one or two positive smears. Implementation of bleach sedimentation led to a significant increase in the diagnosis of smear-positive patients. Implementation of SCA did not result in significantly increased diagnosis of tuberculosis, but did result in improved sample quality. Requesting extra sputum samples based on SCA results, combined with bleach sedimentation, could significantly increase the detection of smear-positive patients if routinely implemented in resource-limited settings where gold standard techniques are not available. We recommend that a pilot phase is undertaken before routine implementation to determine the impact in a particular context.

Topics: Adult; Bacteriological Techniques; Congo; Developing Countries; Disinfectants; Female; Humans; Male; Microscopy; Middle Aged; Prospective Studies; Sensitivity and Specificity; Sodium Hypochlorite; Specimen Handling; Sputum; Tuberculosis

Bleach sputum concentration and fluorescence microscopy (FM) are reportedly more sensitive than direct Ziehl-Neelsen (ZN) sputum smears for tuberculosis detection, and might be particularly valuable for human immunodeficiency virus (HIV)-positive patients excreting fewer bacilli. This study, implemented in Yaoundé, Cameroon, determined the yield from both direct and bleach-concentrated FM and ZN duplicate smears against culture on Löwenstein-Jensen medium, with HIV testing from the sputa. From 418 HIV-positive and 518 HIV-negative tuberculosis suspects, 185 (44.3%) and 243 (46.9%) cultures, respectively, grew Mycobacterium tuberculosis. Direct ZN was positive for, respectively, 87 (47.0%) and 202 (83.1%) of the culture-positive cases. Proportional incremental yield over direct ZN from ZN and FM bleach smears was 14.9% (P < 10(-3)) and 17.2% (P < 10(-4)) for HIV-positive versus 4.9% (P < 10(-2)) and 2.0% (non-significant) for HIV-negative cases. There was no gain from direct FM. Bleach FM showed 2% excess false positives. The bleach concentration, therefore, increases the yield of ZN and FM, particularly from HIV-positive patients, but with a higher risk for false positives with bleach FM. With excellent baseline direct ZN, the gain remains modest. Field studies under real-life conditions are needed to determine whether it is worth the risks and operational challenges in HIV high-prevalence populations. FM was not more sensitive than ZN in this study, probably because of sub-optimal objective power and background staining. Culture on solid media with sparing laurylsulfate decontamination was clearly superior for HIV-positives, but it remains to be seen if culture also leads to more cases started on treatment routinely.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Centrifugation; Chi-Square Distribution; Child; Child, Preschool; Enzyme-Linked Immunosorbent Assay; Female; HIV Infections; Humans; Male; Microscopy; Middle Aged; Mycobacterium tuberculosis; Rosaniline Dyes; Sensitivity and Specificity; Sodium Hypochlorite; Sputum; Tuberculosis; Young Adult

Although treatment of sputum using bleach has shown increased sensitivity in smear microscopy, the safety aspect of sodium hypochlorite (NaOCl) has not been addressed.. To determine the biocidal effect of NaOCl on Mycobacterium tuberculosis when used as a safety measure in direct sputum smear microscopy for the diagnosis of tuberculosis (TB).. A total of 156 smear- and culture-positive pooled sputum specimens were assessed for the viability of M. tuberculosis after treatment with NaOCl. Each specimen was divided into 11 equal portions. One portion was directly cultured. Each of five portions was treated with 3.5% NaOCl and the other five with 5% NaOCl. Specimens were then cultured at intervals of 15 min and 1, 3, 15 and 24 h.. A total of 18 (11.5%) specimens showed growth after treatment with NaOCl. Of these, 5 (3.2%) showed growth upon treatment with 3.5% NaOCl at 15 min and 15 and 24 h. Ten (6.4%) showed growth after treatment with 5% NaOCl between 15 min and 3 h, but none showed growth after 15 h. Three (1.9%) specimens showed growth in both concentrations.. Our preliminary findings show that exposure of sputum to 5% NaOCl for at least 15 h may be a safety procedure in smear microscopy for the diagnosis of TB. However, further studies are warranted.

Topics: Bacteriological Techniques; Culture Media, Conditioned; Humans; Microscopy; Mycobacterium tuberculosis; Safety; Sampling Studies; Sensitivity and Specificity; Sodium Hypochlorite; Sputum; Tuberculosis

Bacteriological diagnosis of tuberculosis (TB) is largely dependent on Ziehl-Neelsen (ZN) microscopy. This method has a low sensitivity. Although concentration of sputum with sodium hypochlorite (NaOC1) followed by sedimentation increases the sensitivity of direct smear microscopy, no study has focused on the effect of NaOC1 on smear negative sputum specimens.. To establish whether 3.5% NaOC1 sedimentation method specifically improves the diagnosis of Ziehl-Neelsen smear negative tuberculosis.. A prospective study.. Mbagathi District Hospital and Center for Respiratory Diseases Research, Kenya Medical Research Institute.. Two hundred and thirty confirmed direct ZN smear negative sputum specimens from new TB suspects were analysed.. Seventy (30.4%) specimens were culture positive. Of these, 19 were ZN smear positive. The ZN sensitivity, specificity, positive and negative predictive values were 27.1%, 99%, 95% and 76%, respectively, after sedimentation with 3.5% NaOC1.. Overnight sedimentation using 3.5% NaOC1 significantly improves diagnosis of ZN smear negative TB. This technique has potential to improve diagnosis in TB diagnostic services especially in settings with high burden of dual TB/HIV infection.

Topics: Clinical Laboratory Techniques; Humans; Prospective Studies; Sodium Hypochlorite; Time Factors; Tuberculosis

The reliability of direct smear microscopy for diagnosis of tuberculosis has frequently been questioned due to low sensitivity. Treatment of sputum with sodium hypochlorite (NaOCI) has been used to increase sensitivity in many settings. However, no study has established the effect of NaOCI on fluorescent microscopy.. To establish whether NaOCI concentration method enhances positivity of fluorescent microscopy smear negative sputum for diagnosis of tuberculosis.. A prospective study.. Mbagathi District Hospital and Centre for Respiratory Diseases Research, Kenya Medical Research Institute.. Forty five (22%) specimens were culture positive. Fluorescent microscopy sensitivity was 28.9% and 22.2% after centrifugation and sedimentation with 3.5% NaOCI, respectively (P > 0.05). Sensitivity was 24.4% and 17.8% after centrifugation and sedimentation with 5% NaOCI, respectively (P > 0.05). Although there was no statistical significance difference between the two NaOCI concentration methods, 3.5% NaOCI with centrifugation indicated a higher yield.. Use of NaOCI significantly enhances positivity of smear negative sputum for diagnosis of tuberculosis when used with fluorescent microscopy. This approach could be recommended for screening all tuberculosis suspects especially in settings with potential smear negative tuberculosis.

Topics: Centrifugation; Clinical Laboratory Techniques; Humans; Mass Screening; Microscopy, Fluorescence; Prospective Studies; Quality Control; Sodium Hypochlorite; Sputum; Tuberculosis

Extra pulmonary tuberculosis (TB) comprises 15% of the total tuberculosis cases. Bleach concentration method for demonstration of Acid fast bacilli (AFB) has been recently described for sputum. The aim of this study is to apply this method for demonstration of AFB in material obtained from extra-pulmonary sites and to correlate with cytology and conventional Ziehl Neelsen (Z N) staining. A total of 55 samples were studied from clinically suspected cases of extrapulmonary TB which included, FNA lymph nodes (17), abscesses drained from various body parts (18), body fluids (18) and skin scrapping (2). All the samples were processed for routine cytology, conventional ZN staining and bleach method followed by ZN staining. Out of 55 samples, 24(43.40%) were indicative of tuberculosis on cytology, 12(21.8%) were positive for AFB on conventional ZN staining, while the positivity increased to 39(70.90%) by Bleach method. Bleach solution is inexpensive and readily available in hospitals and its application has been proved in pulmonary tuberculosis. However to the best of our knowledge this is a pioneer study applied to the extra-pulmonary samples and the results of the present study shows improved detection of AFB.

Topics: Adolescent; Adult; Bacteriological Techniques; Child; Female; Humans; India; Male; Middle Aged; Mycobacterium tuberculosis; Sodium Hypochlorite; Tuberculosis

In order to try to improve the results of direct smear microscopy, we used the mucus-digesting quality of chitin in tuberculosis (TB) laboratories. For this purpose, a total of 430 sputum specimens were processed by the N-acetyl-L-cysteine concentration, sodium hypochlorite (NaOCl) liquefaction, chitin sedimentation, and direct microscopy methods. Then, the smear sensitivity for acid-fast bacillus detection by chitin-treated sputum was compared with the sensitivity of smears prepared by other methods. Our results showed that the chitin solution took less time to completely homogenize the mucoid sputum than did the N-acetyl-L-cysteine and NaOCl methods. The N-acetyl-L-cysteine concentration method demonstrated sensitivity and specificity levels of 83 and 97%, respectively. In comparison, the sensitivity of chitin sedimentation was 80%, with a specificity of 96.7%. The NaOCl liquefaction method showed a sensitivity of 78%, with a specificity of 96%. Finally, the sensitivity of direct microscopy was lower than those of the other tested methods and was only 46%, with a specificity of 90%. The chitin and NaOCl liquefaction methods are both easy to perform, and they do not require additional equipment (centrifuges). Also, our results demonstrated that the chitin method is less time-consuming than the NaOCl method, since only 30 min of incubation is required to bring complete sedimentation of bacilli in chitin-treated sputum whereas the NaOCl method needs 10 to 12 h to give the same results in the same sputum specimens. Therefore, the chitin liquefaction and sedimentation method may provide better results in TB laboratories of developing countries than the N-acetyl-L-cysteine concentration, NaOCl overnight sedimentation, and direct smear microscopy methods.

Topics: Acetylcysteine; Centrifugation; Chitin; Humans; Microbiological Techniques; Microscopy; Mycobacterium tuberculosis; Sensitivity and Specificity; Sodium Hypochlorite; Sputum; Tuberculosis

Topics: Animals; Bacteriological Techniques; Chloramines; Equipment and Supplies; Mycobacterium; Mycobacterium bovis; Mycobacterium tuberculosis; Phosphates; Sodium Hypochlorite; Specimen Handling; Tuberculosis; Veterinary Medicine

Topics: Bacillus; Humans; Sodium Hypochlorite; Tuberculosis; Tuberculosis, Pulmonary