The difficulties of childhood tuberculosis diagnosis.

AIMS AND OBJECTIVES: The WHO estimate that 8.8millions new cases of tuberculosis occurred in 2010, (between 8.5 and 9.2millions) linked to 1.45millions death cases. The load of children tuberculosis is estimated at 10-15 per cent of the total load. In 2014 more than one million children have developed the disease. The children just as adults are exposed to contract and develop the multi resistant forms of the tuberculosis, constituting a major issue for the disease control. The children less than 5years of age are the most exposed to present the most serious and more often deadly forms of the illness. Further, in many developing countries, the lack of pediatric forms of the tuberculosis drugs makes it difficult to control the problem. The tuberculosis diagnosis among the children is based on a set of arguments: the presence of a tuberculous person excreting bacillus, exposition and receptivity conditions of the child (the level of his immunity, the level of under nutrition, associated pathologies etc…). The diagnosis is also based on the research of the symptoms and other signs suggestive of tuberculosis: tuberculin skin test, thoracic radiography, interferon-gamma test. The aim of this study, is to describe and analyze the features and difficulties of the biological diagnosis of tuberculosis among the children and to find a strategy for the improvement of the results. METHODS: It's a retrospective study from 2002 to 2015, dealing with pediatric patients' records from whom a bacteriological diagnosis was requested. We took advantage of the methods used on the laboratory to establish a diagnosis: microscopy, culture, study of the susceptibility to the tuberculosis drugs in solid medium and molecular biology. RESULTS: From 2002 to 2015, only 207 strains were isolated from the children samples, aged from 0 to 15years predominantly female sex (sex ratio is 0.53) with an average age of 9years. The detailed results of the diagnosis methods of tuberculosis and the drugs resistance will be presented. CONCLUSIONS: Tuberculosis in children is often undiagnosed or difficult to diagnose, most developing countries still using ancient methods which can recognize only the developed tuberculosis. It's necessary to evaluate the issue's importance in order to improve the diagnosis conditions (systematic culture and susceptibility test in children), and to ensure the availability of the effective treatment (the pediatric formulation of the essential drugs).

Rifampicin-resistant Mycobacterium tuberculosis: susceptibility to isoniazid and other anti-tuberculosis drugs.

Based on data from 14 Supranational Tuberculosis (TB) Reference Laboratories worldwide, the proportion of rifampicin (RMP) resistant isolates that were isoniazid (INH) susceptible by phenotypic drug susceptibility testing varied widely (0.5-11.6%). RMP-resistant isolates that were INH-susceptible had significantly lower rates of resistance to other first- and second-line anti-tuberculosis drugs (except rifabutin) compared to multidrug-resistant isolates. RMP resistance is not always a good proxy for a presumptive diagnosis of multidrug-resistant TB, which has implications for use of molecular assays that identify only RMP resistance-associated DNA mutations.

Global trends in resistance to antituberculosis drugs. World Health Organization-International Union against Tuberculosis and Lung Disease Working Group on Anti-Tuberculosis Drug Resistance Surveillance.

BACKGROUND: Data on global trends in resistance to antituberculosis drugs are lacking. METHODS: We expanded the survey conducted by the World Health Organization and the International Union against Tuberculosis and Lung Disease to assess trends in resistance to antituberculosis drugs in countries on six continents. We obtained data using standard protocols from ongoing surveillance or from surveys of representative samples of all patients with tuberculosis. The standard sampling techniques distinguished between new and previously treated patients, and laboratory performance was checked by means of an international program of quality assurance. RESULTS: Between 1996 and 1999, patients in 58 geographic sites were surveyed; 28 sites provided data for at least two years. For patients with newly diagnosed tuberculosis, the frequency of resistance to at least one antituberculosis drug ranged from 1.7 percent in Uruguay to 36.9 percent in Estonia (median, 10.7 percent). The prevalence increased in Estonia, from 28.2 percent in 1994 to 36.9 percent in 1998 (P=0.01), and in Denmark, from 9.9 percent in 1995 to 13.1 percent in 1998 (P=0.04). The median prevalence of multidrug resistance among new cases of tuberculosis was only 1.0 percent, but the prevalence was much higherin Estonia (14.1 percent), Henan Province in China (10.8 percent), Latvia (9.0 percent), the Russian oblasts of Ivanovo (9.0 percent) and Tomsk (6.5 percent), Iran (5.0 percent), and Zhejiang Province in China (4.5 percent). There were significant decreases in multidrug resistance in France and the United States. In Estonia, the prevalence in all cases increased from 11.7 percent in 1994 to 18.1 percent in 1998 (P<0.001). CONCLUSIONS: Multidrug-resistant tuberculosis continues to be a serious problem, particularly among some countries of eastern Europe. Our survey also identified areas with a high prevalence of multidrug-resistant tuberculosis in such countries as China and Iran.

Genetic diversity of Mycobacterium africanum clinical isolates based on IS6110-restriction fragment length polymorphism analysis, spoligotyping, and variable number of tandem DNA repeats.

A collection of 105 clinical isolates originally identified as Mycobacterium africanum were characterized using both phenotypic and genotyping methods. The phenotypic methods included routine determination of cultural properties and biochemical tests used to discriminate among the members of the M. tuberculosis complex, whereas genotypic characterization was based on IS6110-restriction fragment length polymorphism (IS6110-RFLP) analysis, IS1081-RFLP analysis, direct repeat-based spacer oligonucleotide typing (spoligotyping), variable number of tandem DNA repeats (VNTR), and the polymorphism of the oxyR, pncA, and mtp40 loci. The results obtained showed that a majority of M. africanum isolates were characterized by a specific spoligotyping pattern that was intermediate between those of M. tuberculosis and M. bovis, which do not hybridize with spacers 33 to 36 and spacers 39 to 43, respectively. A tentative M. africanum-specific spoligotyping signature appeared to be absence of spacers 8, 9, and 39. Based on spoligotyping, as well as the polymorphism of oxyR and pncA, a total of 24 isolates were excluded from the final study (19 were identified as M. tuberculosis, 2 were identified as M. canetti, and 3 were identified as M. bovis). The remaining 81 M. africanum isolates were efficiently subtyped in three distinct subtypes (A1 to A3) by IS6110-RFLP analysis and spoligotyping. The A1 and A2 subgroups were relatively more homogeneous upon spoligotyping than A3. Further analysis of the three subtypes by VNTR corroborated the highly homogeneous nature of the A2 subtype but showed significant variations for subtypes A1 and A3. A phylogenetic tree based on a selection of isolates representing the three subtypes using VNTR and spoligotyping alone or in combination confirmed the subtypes described as well as the heterogeneity of subtype A3.

Mycobacterial growth indicator tube versus the proportion method on Löwenstein-Jensen medium for antibiotic susceptibility testing of Mycobacterium tuberculosis.

The Mycobacterial Growth Indicator Tube, a reliable system for detection of mycobacterial growth, was compared with the reference proportion method on Löwenstein-Jensen medium for antibiotic susceptibility testing of Mycobacterium tuberculosis. A total of 62 clinical strains and four reference strains of Mycobacterium tuberculosis were tested for susceptibility to streptomycin, isoniazid, rifampicin and ethambutol. Of these, 36 were susceptible to all four antibiotics and 30 were resistant to at least one of them. Tests were repeated in cases of discrepant results. When each drug/strain combination was considered separately, the overall agreement between the two methods was 96.5% (98.4% for streptomycin, 95.3% for isoniazid, 96.9% for rifampicin and 95.3% for ethambutol) with regard to the initial testing and 98.8% (100, 98.5, 98.5 and 98.4%, respectively) after repeated testing. When the results were considered strain by strain, the agreement was 86% after the initial testing and 95% after repeated testing. The results were obtained after a mean time of 9.5 days. These results suggest that the Mycobacterial Growth Indicator Tube is a reliable method for testing susceptibility of mycobacterial strains to first-line antituberculous drugs.

A national survey of human Mycobacterium bovis infection in France. Network of Microbiology Laboratories in France.

OBJECTIVE: To evaluate the role of Mycobacterium bovis in the epidemiology of human tuberculosis in France. DESIGN: A national survey in France in 1995 using a questionnaire mailed to all French microbiological laboratories performing mycobacteria cultures. RESULTS: M. bovis was isolated in 38 out of 7075 cases of bacteriologically confirmed tuberculosis (0.5%) notified to the National Reference Centre (CNR) in 1995, resulting in an incidence of 0.07 per 100,000 population. Incidence rates increased with age, and were the highest among patients of 75 years or more (range 0.02-0.33/100,000). Two cases of tuberculosis due to M. bovis were reported in foreign-born children who had come to France for treatment of their disease. No cases were reported among French-born children. The site of tuberculosis was pulmonary in 17 cases, extra-pulmonary in 14, both pulmonary and extra-pulmonary in one, and unknown in six. Extra-pulmonary sites were more frequent in older patients, and pulmonary sites more frequent in younger patients. Two patients were coinfected with the human immunodeficiency virus. Occupational exposure was identified in 13 cases and ingestion of non pasteurised milk in three. In addition, 11 patients had a possible risk of exposure related to their country of birth, family contact or occupation. CONCLUSION: In France, the 0.5% proportion of human tuberculosis due to M. bovis is similar to that of other developed countries. The higher incidence of the disease among older people is likely to reflect the efficacy of the control measures for tuberculosis in cattle.

Prevalence of primary and acquired resistance of Mycobacterium tuberculosis to antituberculosis drugs in Benin after 12 years of short-course chemotherapy.

SETTING: Benin National Tuberculosis Programme, West Africa. OBJECTIVE: To measure the prevalence of primary and acquired resistance of Mycobacterium tuberculosis to the antituberculosis drugs isoniazid, rifampicin, ethambutol and streptomycin in Benin from 1994-1995, after 12 years of short-course chemotherapy regimens. METHODS: Prospective study by cluster sampling according to the methodology recommended by the International Union Against Tuberculosis and Lung Disease (IUATLD) and the World Health Organization (WHO). RESULTS: The survey of primary resistance included 333 strains, of which 28 (8.4%) were drug-resistant, one to both rifampicin and isoniazid (multidrug-resistant). For acquired resistance, out of 57 strains tested 26 (45.6%) were resistant, six of which (11%) were multidrug-resistant. CONCLUSION: Despite the considerable increase in the number of tuberculosis cases observed in recent years (52% between 1987 and 1995), direct observation of patients taking their antituberculosis drugs during the intensive phase of treatment has limited the development of drug resistance in Benin.

The prevalence of Mycobacterium tuberculosis drug resistance in New Caledonia, 1995-1996.

SETTING: Study of the susceptibility to anti-tuberculosis drugs of Mycobacterium tuberculosis strains isolated in New Caledonia, a French South Pacific Territory, where tuberculosis continues to be a public health problem. OBJECTIVE: To assess the stability of this susceptibility in order to justify both non-systematic susceptibility testing and the implementation of simplified chemotherapy regimens. METHODS: Over a period of nearly 2 years (1995-1996), every new case of tuberculosis confirmed by the laboratory was included in the study. A total of 105 strains were tested against five anti-tuberculosis drugs: isoniazid, rifampicin, ethambutol, pyrazinamide and streptomycin. RESULTS: No primary drug resistance was detected for the main drugs. One strain with acquired resistance to isoniazid and streptomycin was isolated from one of the 12 patients suffering a relapse of the disease. CONCLUSIONS: The results of this exhaustive study justify the non-systematic approach to susceptibility testing for new patients. However, for strains isolated from patients suffering from relapse or therapeutic failure, or who belong to a high risk population, drug susceptibility testing should be performed. This kind of management will aid in the detection of possible isoniazid and streptomycin resistance, thus avoiding the selection and possible emergence of strains resistant to rifampicin. The results of the study argue for the use of a fixed dose regimen using triple combination tablets of isoniazid, rifampicin and pyrazinamide (HRZ) for 2 months, followed by dual drug therapy (HR) for 4 months.

Use of two methods of analysis to estimate the annual rate of tuberculosis infection in Southern Algeria.

SETTING: Analysis of tuberculin skin test surveys. OBJECTIVE: To estimate the annual tuberculous infection rate in Tamanrasset (southern Algeria) by applying on the one hand the classical method of the Tuberculosis Surveillance Research Unit (TSRU) of the International Union Against Tuberculosis, and on the other the study of variations of allergy published by Raj Narain et al. METHODS: 3675 pupils aged 5-18 years were tested in October 1982 or November 1983; 1240 of them were tested on both occasions. The technique of the World Health Organization, with 2 TU PPD RT23 tuberculin was used by trained testers. The TSRU method was applied to the 863 children aged 5-15 years without bacille Calmette-Guerin (BCG) scar, according to three estimates of the prevalence of infection (cut-off points of the distribution of reactions of 10 mm, of 14 mm corrected by dividing by 0.82, and mirror technique with a mode of 17 mm), and to three hypotheses of the decrease in infection risk (1%, 3% and 5%). The study of variations of allergy was used in the 1240 pupils tested twice (BCG-vaccinated or not). RESULTS: The average annual risk of tuberculous infection estimated by the TSRU method was lower than 1% (0.5%-0.6% according to the most stringent criteria). By the method of variations of allergy, the estimated annual infection rate was 0.9%. For the latter, our results are consistent with those obtained by other researchers. CONCLUSIONS: These findings suggest that the study of variations of allergy might be a good alternative approach to estimate the annual infection rate in countries where children are BCG-vaccinated at birth, which is the case for most developing countries.

Surveillance of drug resistance for tuberculosis control: why and how?

The resistance of Mycobacterium tuberculosis to antibiotics, which reflects the quality of the chemotherapy applied in the community, is one of the elements of epidemiological surveillance used in national tuberculosis programmes. Measurement of drug resistance poses problems for biologists in standardization of laboratory methods and quality control. The definition of rates of acquired and primary drug resistance also necessitates standardization in the methods used to collect information transmitted by clinicians. Finally, the significance of the rates calculated depends on the choice of the patients sample on which sensitivity tests have been performed. National surveys of drug resistance therefore require multidisciplinary participation in order to select the only useful indicators: rates of primary resistance and of acquired resistance. These indicators, gathered in representative groups of patients over a long period, are a measurement of the impact of modern chemotherapy regimens on bacterial ecology.

[Clinical trial of a combination of three drugs in fixed proportions in the treatment of tuberculosis. Groupe de Travail sur la Chimiothérapie de la Tuberculose]. / Essai clinique d'une combinaison en proportions fixes de trois medicaments dans le traitement de la tuberculose. Groupe de Travail sur la Chimiothérapie de la Tuberculose.

SETTING: The Matiben Chest Clinic at the West Algiers University Teaching Hospital, and 3 outpatient clinics specializing in tuberculosis and lung disease in Algiers. OBJECTIVE: To determine the tolerance and efficacy of a fixed proportion combination of 3 antituberculosis drugs (per tablet: 50 mg isoniazid + 120 mg rifampicin + 300 mg pyrazinamide) given during the first 2 months of a daily 6-month chemotherapy regimen. DESIGN: Random prospective treatment trial comparing a group of 124 patients receiving the triple combination with another group of 126 patients receiving the 3 drugs separately during the initial treatment phase. The continuation phase was identical for the 2 groups. Comparison of tolerance in the first 2 months, and of the failure and relapse rates (respectively at the end of treatment and 24 months after the end of treatment). RESULTS: During the first 2 months side-effects were significantly more common in the group receiving the drugs separately. At the end of treatment and during the following 24 months there were no significant differences in the cumulative rates of observed failures and relapses (2% and 1%). CONCLUSION: The triple combination studied could replace the separate drugs in the initial treatment phase in countries where the bioavailability of the drugs used has been proven.

[A therapeutic trial of a combination of 3 essential drugs in a short course of chemotherapy in tuberculosis. Results 6 months after the end of treatment]. / Essai thérapeutique d'une combinaison de trois médicaments essentiels dans la chimiothérapie courte de la tuberculose. Résultats six mois après la fin du traitement.

250 patients suffering from pulmonary tuberculosis who were smear positive received a chemotherapy regime for 6 months combining Rifampicin and Isoniazid every day with a daily supplement of Pyrazinamide for the first 8 weeks. The three drugs given in the initial phase of treatment were administered either separately or in combined preparations according to the 2 controlled randomised groups. During the maintenance phase the drugs were given in a combined form in fixed proportions in the 2 groups. 6 months after the end of treatment the bacteriological results were similar in the 2 groups in the 144 cases which were analysed. Amongst 137 cases with bacilli which were initially sensitive to Isoniazid (68 cases with separate medicines at the beginning and 69 with combined drugs) there was no failure at 6 months, nor any relapse during the course of the first period of surveillance. Amongst 7 cases with bacilli which were originally resistant to Isoniazid (4 and 3 respectively), there were 2 failures at 6 months one in each group with acquired resistance to Rifampicin observed at the time of the failure. There was no difference in the therapeutic results observed whatever the presentation of drugs used during the initial phase of treatment.

[A controlled survey comparing 2 short-term therapeutic regimes in lymph node tuberculosis]. / Enquête contrôlée comparant deux régimes thérapeutiques de courte durée dans la tuberculose ganglionnaire.

To study the efficacy of short term chemotherapy in the treatment of peripheral glandular tuberculosis a controlled trial was carried out in Algiers in March 1982 comparing two therapeutic regimes. All the patients admitted to the study presented with glandular tuberculosis which was proved either histologically or bacteriologically. They were recruited in three clinics serving the Algerian population and were required to live in Algiers. The two anti-tuberculous regimes used consisted of an initial phase of four drugs: Rifampicin (R), Isoniazid (H), Streptomycin (S) and Pyrazinamide (Z) every day for two months. This initial phase was followed by R.H. every day for four months in regime A, making six months treatment in all and for seven months in regime B making nine months treatment in all. 141 patients were thus admitted to the study, of whom 117 could be used for analysis at the end of treatment. Of these 68 were female making up 58% of the total. 12 patients or 10% were under 15 years of age. After two years of review following the end of treatment there were nine therapeutic failures (7.7%) of whom five were in regime A and four were in regime B (no significant difference). Amongst the failures large volume nodes persisted in three patients and two patients presented with new nodes. The lymph nodes increased in volume at the end of treatment in two cases; and finally two patients presented again with fistulae at the end of treatment. There were eight unfavourable outcomes in nine patients under observation during treatment or at the end of chemotherapy. There was only one failure noted some time after the finish of treatment at the end of two years of follow up.

[Therapeutic trial of a combination of isoniazid, rifampicin and pyrazinamide in the first 2 months of treatment of pulmonary tuberculosis]. / Essai thérapeutique d'une combinaison d'isoniazide, de rifampicine et de pyrazinamide au cours des deux premiers mois du traitement de la tuberculose pulmonaire.

250 patients suffering from smear positive pulmonary tuberculosis who had never been treated before, received a six-month regimen combining isoniazid and rifampicin every day with a supplement of pyrazinamide for the first eight weeks of treatment. The three drugs given during the initial phase of therapy were administered in two different forms: either as separate tablets or in a form combining all three drugs in fixed proportions (each tablet contained 50 mg of isoniazid, 120 mg of rifampicin and 300 mg of pyrazinamide). The patients were randomised into two groups of 125 receiving one or other combination of medications with the dose adapted for their weight. An analysis of the results at the end of the second month of treatment was carried out on 240 cases who could be analysed for tolerance and acceptability and in 193 cases who were analysed for the efficacy of therapy as judged by negative cultures. The treatment results were excellent in both groups, as was patient acceptability. The level of side effects was significantly lower in the group receiving combined medication (p less than 0.02). Both groups gave a similar proportion of negative cultures at the end of two months. The combined medicaments studied are thus well tolerated and at least as effective as individual drugs. They may thus replace them.