Biocidal effect of bleach on Mycobacterium tuberculosis: a safety measure.

BACKGROUND: Although treatment of sputum using bleach has shown increased sensitivity in smear microscopy, the safety aspect of sodium hypochlorite (NaOCl) has not been addressed. OBJECTIVE: 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). METHOD: 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. RESULTS: 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. CONCLUSION: 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.

Improved diagnosis of Ziehl-Neelsen smear negative tuberculosis using sodium hypochlorite sedimentation method.

BACKGROUND: 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. OBJECTIVE: To establish whether 3.5% NaOC1 sedimentation method specifically improves the diagnosis of Ziehl-Neelsen smear negative tuberculosis. DESIGN: A prospective study. SETTING: Mbagathi District Hospital and Center for Respiratory Diseases Research, Kenya Medical Research Institute. SUBJECTS: Two hundred and thirty confirmed direct ZN smear negative sputum specimens from new TB suspects were analysed. RESULTS: 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. CONCLUSION: 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.

Combination of bleach and flourescent microscopy: a milestone in the diagnosis of smear negative tuberculosis.

BACKGROUND: 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. OBJECTIVE: To establish whether NaOCI concentration method enhances positivity of fluorescent microscopy smear negative sputum for diagnosis of tuberculosis. DESIGN: A prospective study. SETTING: Mbagathi District Hospital and Centre for Respiratory Diseases Research, Kenya Medical Research Institute. RESULTS: 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. CONCLUSION: 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.

Isolation of multidrug-resistant tuberculosis strains in patients from private and public health care facilities in Nairobi, Kenya.

SETTING: Health care facilities in Nairobi, Kenya. OBJECTIVE: To document the presence of multidrug-resistant tuberculosis (MDR-TB) strains in patients from Nairobi between September 1999 and October 2001. DESIGN: Descriptive study. RESULTS: Of the 983 referred patients who submitted sputum for culture and drug susceptibility testing (DST), 59% were males. Two hundred and nine (21.3%) patients had a positive culture, of whom 15.2% had a request for DST against isoniazid, rifampicin, streptomycin and ethambutol. Of these, 65 (43.6%) had an isolate resistant to one or more drugs, while 17 (11.4%) had MDR-TB. Ten (59.0%) cases were referred from public health care facilities while seven (41%) were from the private sector. Sixteen isolates were resistant to all four drugs. All MDR-TB cases but one were from Nairobi. CONCLUSION: The emergence of MDR-TB in Nairobi is a cause for concern. An outbreak would be catastrophic, creating not only increased morbidity and mortality but also a tremendous strain on already limited health care resources. Lack of policies for the treatment and management of MDR-TB and the unavailability of appropriate diagnostic facilities may increase its spread. Efforts to prevent outbreaks of MDR-TB should be emphasised.

Identification of MDR-TB Beijing/W and other Mycobacterium tuberculosis genotypes in Nairobi, Kenya.

SETTING: Suspected tuberculosis (TB) patients in Nairobi, Kenya. OBJECTIVE: To identify the presence of multidrug-resistant (MDR) Beijing/W type and other genotypes of Mycobacterium tuberculosis. METHODS: Thirty-three isolates resistant to one or more drugs (resistance ratio method), including 15 MDR isolates and 40 susceptible isolates selected at random, were analysed by dot-blot hybridisation for mutations associated with resistance to isoniazid, rifampicin, streptomycin and ethambutol. All strains were genotypically classified using spoligotyping. RESULTS: Of the 33 drug-resistant isolates, 21 (64%) were from males and 12 (36%) were from females. Mutations associated with resistance to isoniazid (katG 315) and rifampicin (rpoB526, 531) were confirmed in 83.3% and 100% of the isolates, respectively, and in 87% of the MDR isolates. Mutations were detected in 25% and 71.5% of the isolates resistant to streptomycin (rpsL43) and ethambutol (embB306), respectively. No mutations were detected in drug-susceptible isolates. Spoligotyping grouped the isolates into 25 groups. Ten of these groups corresponded to previously identified strain groups, including seven families in the international database. One of these families (CAS1) comprised six (40%) of the 15 MDR isolates. Another family (Beijing) had six (8.3%) isolates, of which two (33.3%) were MDR (Beijing/W). CONCLUSION: This study is the first in Kenya and the second in sub-Saharan Africa to report the presence of MDR Beijing/W type and other possible drug-resistant outbreak strains. Application of the molecular techniques and markers will allow us to monitor the spread of existing drug-resistant strains and the appearance of new ones.

Surveillance of drug-resistant tuberculosis and molecular evaluation of transmission of resistant strains in refugee and non-refugee populations in North-Eastern Kenya.

SETTING: Three refugee camp complex clinics and an adjacent non-refugee treatment centre in North-Eastern Kenya. OBJECTIVES: To use conventional and molecular epidemiology tools to determine: 1) the prevalence of drug resistance in newly diagnosed patients with smear-positive pulmonary tuberculosis in refugee and non-refugee populations; 2) risk factors for resistance in the two populations; and 3) whether IS6110 restriction fragment length polymorphism (RFLP) and spoligotyping showed similarities in DNA fingerprinting patterns of drug-resistant isolates that could infer transmission within and between the two populations. RESULTS: Of 241 isolates from the camps, 44 (18.3%) were resistant to one or more drugs, seven of which (2.9%) were multidrug-resistant TB (MDR-TB). Of 88 isolates from the non-refugees, five (5.7%) were resistant to one or more drugs without MDR-TB. Drug resistance was higher in the camps than in the non-refugee population (OR = 3.7; 95%CI 1.42-9.68; P < 0.007). Resistance was significantly higher in one camp compared with the other two, despite a comparable ethnic distribution. Unusually, females were more associated with drug resistance than their male counterparts in both populations (OR = 2.3; 95%CI 1.2-4.8; P = 0.008). There was evidence of transmission of streptomycin-resistant strains in the refugee population. DNA fingerprints of resistant strains from the non-refugee population were unique and different from those in the refugee camps. CONCLUSION: The observed high levels of drug resistance and MDR-TB, combined with evidence of transmission of strains resistant to streptomycin in the refugee population, suggest a need for strengthened TB control programmes in settings with a high risk of developing drug-resistant strains.

Maternal mortality occurring at the Rift Valley Provincial General Hospital, Nakuru.

OBJECTIVE: To evaluate the factors that led to maternal mortality over a five-year period at the Rift Valley Provincial General Hospital, Nakuru. DESIGN: A retrospective survey based on secondary data. SETTING: Department of Obstetrics and Gynaecology, Rift Valley Provincial General Hospital, Nakuru, between 1994 and 1998. SUBJECTS: Evaluation of 109 deaths where records were available. RESULTS: The average maternal mortality was 3.16/1000 live births. Twenty eight (26%) occurred in teenagers. Sixty two deaths (58%) accounted for those aged (14-24 years). Thirty four per cent of sixty two deaths occurred in nulliparous and primiparous patients. The commonest cause of death was haemorrhage and ruptured uterus (44%). Most deaths (53%) occurred within 24 hours of admission. Of these, 36 (62%) came directly from home in labour or ill or where deliveries were being attempted by non-medical persons. CONCLUSION: Most of the deaths evaluated were preventable. Management of the major causes of death include blood (haemorrhage and ruptured uteri) transfusion. With poor blood transfusion services and the risk of HIV, this calls for alternatives to direct blood transfusion. Referral systems need to be overhauled and referring health institutions should get properly staffed. Despite good antenatal clinic attendance and availability of resources most patients still deliver at home.

Antituberculosis drug resistance surveillance in Kenya, 1995.

SETTING: Twenty-two of the 42 administrative districts in Kenya. OBJECTIVE: To determine the prevalence of drug resistance in newly diagnosed patients with pulmonary tuberculosis, to determine possible risk factors associated with resistance, and to establish standard routine surveillance of drug resistance. DESIGN: Cross-sectional study. METHODS: Sputum samples from newly diagnosed patients with smear-positive pulmonary tuberculosis were analysed using standard procedures. RESULTS: Of 638 patients, 85% were culture positive for Mycobacterium tuberculosis. Of 491 patients tested for susceptibility to isoniazid, streptomycin, rifampicin and ethambutol, 90.8% had fully sensitive strains and 9.2% had a strain resistant to one or more drugs. Of 445 patients with no history of previous chemotherapy, 6.3% had a resistant strain. Of 46 patients with a history of previous chemotherapy, 37% had a resistant strain. No resistance to either rifampicin or ethambutol was detected. There was a strong association between previous chemotherapy and resistance. Resistance was not associated with age or sex. High concordance between Kenya's results and those of the Mycobacterium Reference Unit in the UK on both drug-sensitive and drug-resistant strains indicates that clinically significant and comparable data can be obtained from laboratories employing unsophisticated and inexpensive standard procedures. CONCLUSION: Rates of initial drug resistance are still low in Kenya. The increase in acquired resistance to isoniazid requires monitoring.

Isoniazid preventive therapy for tuberculosis in HIV-1-infected adults: results of a randomized controlled trial.

OBJECTIVES: To determine the efficacy of isoniazid 300 mg daily for 6 months in the prevention of tuberculosis in HIV-1-infected adults and to determine whether tuberculosis preventive therapy prolongs survival in HIV-1-infected adults. DESIGN AND SETTING: Randomized, double-blind, placebo-controlled trial in Nairobi, Kenya. SUBJECTS: Six hundred and eighty-four HIV-1-infected adults. MAIN OUTCOME MEASURES: Development of tuberculosis and death. RESULTS: Three hundred and forty-two subjects received isoniazid and 342 received placebo. The median CD4 lymphocyte counts at enrolment were 322 and 346 x 10(6)/l in the isoniazid and placebo groups, respectively. The overall median follow-up from enrolment was 1.83 years (range, 0-3.4 years). The incidence of tuberculosis in the isoniazid group was 4.29 per 100 person-years (PY) of observation [95% confidence interval (CI) 2.78-6.33] and 3.86 per 100 PY of observation (95% CI, 2.45-5.79) in the placebo group, giving an adjusted rate ratio for isoniazid versus placebo of 0.92 (95% CI, 0.49-1.71). The adjusted rate ratio for tuberculosis for isoniazid versus placebo for tuberculin skin test (TST)-positive subjects was 0.60 (95% CI, 0.23-1.60) and for the TST-negative subjects, 1.23 (95% CI, 0.55-2.76). The overall adjusted mortality rate ratio for isoniazid versus placebo was 1.18 (95% CI, 0.79-1.75). Stratifying by TST reactivity gave an adjusted mortality rate ratio in those who were TST-positive of 0.33 (95% CI, 0.09-1.23) and for TST-negative subjects, 1.39 (95% CI, 0.90-2.12). CONCLUSIONS: Overall there was no statistically significant protective effect of daily isoniazid for 6 months in the prevention of tuberculosis. In the TST-positive subjects, where reactivation is likely to be the more important pathogenetic mechanism, there was some protection and some reduction in mortality, although this was not statistically significant. The small number of individuals in this subgroup made the power to detect a statistically significant difference in this subgroup low. Other influences that may have diluted the efficacy of isoniazid include a high rate of transmission of new infection and rapid progression to disease or insufficient duration of isoniazid in subjects with relatively advanced immunosuppression. The rate of drug resistance observed in subjects who received isoniazid and subsequently developed tuberculosis was low.

No increased prevalence of adrenocortical insufficiency in human immunodeficiency virus-associated tuberculosis.

SETTING: Acute medical wards, Kenyatta National Hospital, Nairobi, Kenya. OBJECTIVE: To determine the prevalence of adrenocortical insufficiency in human immunodeficiency virus (HIV)-1 infected and non-infected patients with tuberculosis. DESIGN: One hundred and seventy-four patients with proven tuberculosis (90 HIV-1 positive and 84 HIV-1 negative) were assessed for adrenocortical insufficiency with a 30 min synacthen stimulation test. RESULTS: Fifty-one percent of those with pulmonary tuberculosis and 56% of those with extra-pulmonary tuberculosis had a subnormal cortisol response. However there was no statistically significant difference between the HIV-1 infected and non-infected patients in either group. CONCLUSION: While an impaired cortisol response is common in tuberculosis, it is no more prevalent in HIV-1 infected patients than non-infected patients with tuberculosis.

A comparative study on the reliability of the fluorescence microscopy and Ziehl-Neelsen method in the diagnosis of pulmonary tuberculosis.

Pulmonary tuberculosis (PTB) is the most common presentation of tuberculosis (TB) in Kenya. For the diagnosis of PTB the sputum smear is used because it is technically simple, non-invasive and cheap. The reliability of direct smear examination for the diagnosis of TB has however frequently been questioned. To address this problem, a study comparing the reliability of fluorescence microscopy (FM) and Ziehl-Neelsen (ZN) staining method for examination of direct smear in the diagnosis of PTB was carried out at the Respiratory Disease Research Unit Laboratory, Nairobi, Kenya. A total of 1480 sputum specimens collected from patients with suspected PTB were analyzed. Two direct smears were prepared from each specimen, one stained using FM and the other using the ZN method. Culture results were used as the gold standard for assessment. Specificity was 97% and 96% for FM and ZN methods, respectively. The sensitivity of the FM method was 80% and that of the ZN method 65% (p < 0.001). Overall agreement was 86.8%. Positive smears which were missed on the ZN stained smears (15%) contained low density bacilli on both FM stained smears and on culture. The use of FM greatly improves the diagnostic value of the sputum smear especially in patients with a low density of bacilli who are likely to be missed on ZN stained smears. The method is economical in both time and expense and is recommended for laboratories handling large numbers of sputum specimens.

Cohort study of HIV-positive and HIV-negative tuberculosis, Nairobi, Kenya: comparison of bacteriological results.

We have set up a cohort of human immunodeficiency virus (HIV) positive and negative patients with tuberculosis in order to address the problems associated with HIV-related tuberculosis. We present here the results of sputum smear microscopy, culture, mycobacterial identification tests and drug susceptibility assays from specimens taken at presentation. In this selected population of largely pulmonary tuberculosis cases, HIV infection is not associated with significant differences in sputum smear positivity rate, culture positivity rate or initial drug resistance. No atypical mycobacteria were found. Direct sputum smear examination remains specific for the diagnosis of tuberculosis in Kenya in spite of the presence of HIV. HIV infection was not associated with an increase in the proportion of pulmonary cases still culture-positive at 6 months. However a significant increase in the proportion of cases still culture-positive at 6 months was seen in those with initially resistant strains and also in those treated with standard regimen (streptomycin, thiacetazone and isoniazid for 1 month followed by thiacetazone and isoniazid for 11 months, 1STH/11TH) rather than a short-course, rifampicin-containing regimen (rifampicin, pyrazinamide and isoniazid for 2 months, together with streptomycin for the first month and followed by 6 months of thiacetazone and isoniazid, SHRZ/6TH).

PIP: A study of HIV-related tuberculosis (TB) in Kenya was conducted by analyzing all cases of new suspected TB presenting from April 1989 through May 1990 at the Infectious Disease Hospital and the Ngaira Avenue Chest Clinic. There were 355 cases, 117 of whom were HIV+. No atypical mycobacteria were isolated. There were no significant differences between HIV+ and HIV-patients as to colony counts, grading of sputum smears or prevalence of drug-resistant strains initially. HIV infection was not associated with a significantly increased proportion of pulmonary TB cases still culture positive at 6 months. However, there was a significant increase in the proportion of cases still culture-positive at 6 months among those who initially had drug resistant strains and who were treated with the standard drug regimen (streptomycin, thiacetazone and isoniazid for 1 month, then thiacetazone and isoniazid for 11 months). The discussion enumerated several factors potentially effecting selection of the high proportion of pulmonary TB patients in this series. Direct sputum smear remains specific for diagnosis of TB in Kenya in spite of HIV. These results still suggest that HIV+ pulmonary TB responds well to standard anti-TB therapy.

Use of common reagent containers to stain sputum smears for acid fast bacilli using ziehl-neelsen technque

The use of common reagent containers for mass staining of sputum smears for acid fast bacilli (AFB) by the Ziehl-Neelsen (ZN:) Technique has previously been discouraged for fear of cross contamination. We undertook the present study to ascertain whether this fear has any justification. Out of 1296 smears prepared from 198 specimens including 9 known negative controls; 108 stainings were performed. 70 specimens and all positive controls were repeatedly positive on different stainings. 69 (99) cases and all positive controls were confirmed by culture. No carry over of AFB from positive to negative smears was experienced. Time; cost and staining mess was significantly reduced during mass eestaining. The use of a common container for staining of sputum smears for AFB by the ZN technique is not only economical but also ideal for laboratories where space is insufficient and larg numbers of sputum specimens are involved