Elsevier

Journal of Infection

Volume 49, Issue 2, August 2004, Pages 94-101
Journal of Infection

Evaluation of tuberculosis transmission in Tehran: using RFLP and spoloigotyping methods

https://doi.org/10.1016/j.jinf.2003.11.015Get rights and content

Abstract

Objectives. To evaluate the DNA polymorphism among Mycobacterium tuberculosis (MTB) strains isolated from new smear positive tuberculosis (TB) patients residing in Tehran capital city of Iran, during the year 2001.

Methods. IS6110-restriction fragment length polymorphism (RFLP) and spoligotyping analyses were performed on 129 M. tuberculosis strains. Additional patient's information was collected for further epidemiological analyses. Patients whose isolates had identical RFLP and spoligotyping patterns were considered a cluster.

Results. The results show that the IS6110 were polymorphic and the strains with 8 or 9 IS6110 copy number were more frequently defected (42%). Out of 129 available isolates, 56 (43%) belonged to clusters and 72 (57%) did not. The risk factors like age, sex, family history or close contact and intravenous drug abuse were associated with clustering. Whereas, unemployment (61%) and poor living conditions (83%) contributed to diseases development in both groups. Spoligotyping of M. tuberculosis strains resulted in 46 different patterns, out of which 38 patterns were unique and reported for the first time. We found one M. tuberculosis strains with a pattern characteristic of the Beijing family.

Conclusion. In the studied time period both reactivation (57%) and recent transmission (43%) were contributing to annual new TB cases in Tehran.

Introduction

According to East Mediterranean Regional office (EMRO) records, the estimated incidence rate of tuberculosis in Tehran (with approximately 12 million population) was 12–16 per 100 000.1 Whereas, the notification rate was less than 70%.2 This means considerable numbers of TB patients are missed in the early stages of diseases. Each of these patients soon develop more extensive disease and themselves become new vectors of transmission. Therefore, the low case detection system might increase the risk of TB infection through active transmission. Previous studies3., 4. showed that 90% of active cases of tuberculosis in Iran resulted from reactivation of an infection contracted years before and that recently transmitted diseases had a minor role. In Iran, the TB surveillance is generally performed by conventional methods.5 The sum of information about sex/age/foreign/non-foreign/part of body involved/pervious history/close contact and microbiological confirmation tests are collected from various public health offices at the district level and send to the statistical office of the TB control manager.5 Further, analysing the data would determine the percentage of exposure, infection and death from diseases inside the country. The method has limited practical value because it cannot identify the source and the route of infection and cannot discriminate between new and old infection. Thereby, for identifying the M. tuberculosis strains which are responsible for multiple infection, presumably through recent transmission as well as the strains found in only one TB patient, presumably representing reactivation of latent infection the IS6110-RFLP and spoligotyping were performed. The IS6110-RFLP is based on the observation that RFLP patterns among non-epidemiologically related isolates show a high degree of variation, whereas epidemiological related M. tuberculosis strains show identical or similar fingerprint patterns.6., 7., 8. However, since RFLP analysis with IS6110 alone may be inconclusive for strains carrying few copies of IS6110,9., 10. we also used an alternative PCR-based technique called spoligotyping.11 The technique detects various non-repetitive spacer sequences located between small repetitive units (direct repeat DR) in the chromosome of M. tuberculosis complex. The spacer sequences are a source of diversity that can be use for detecting polymorphism among M. tuberculosis complex.12 In this study, we focused on Iran-born TB patients who were residing in Tehran and had no prior history of tuberculosis treatment.

Section snippets

Data collection

The study was conducted from January to December 2001. Case data were collected by trained technicians using standard questionnaires. Information was obtained on sex, age, country of birth, close contact, previous TB history, present address and associated medical data, such as human immunodeficiency virus (HIV) infection, tuberculin skin test and chest radiography finding.

Bacterial strains

Primary isolation and culturing of Mycobacterium isolates from sputum specimens were followed in accordance to procedures

Characteristics of the Patients

Of the initial 218 patients with pulmonary TB, 89 (40.8%) were excluded because the patients were from retreatment cases or they were Afghan-born immigrants. The study group, therefore, consisted of 129 Iranian cases (59%) with no prior history of tuberculosis. All of them were incidents of in Tehran during the last 10 years. The patients were between 18 and 95 years old. The male to female ratio was 1.2:1. One hundred and eleven patients had (86%) in ratios susceptible to all four drugs

Discussion

The major consideration of the usefulness of the IS6110-RFLP typing method is its specificity6., 8. which is depends on the number of bands obtained. Previously,3 we found 38% of M. tuberculosis strains with low or no copies of IS6110. These strains were collected from Khorasan province which is 930 km in north east of Tehran. In the present study, only seven strains (5.4%) were found with low or no copies of IS6110. Whereas, the remaining isolates (94.5%) contained more than six copies.

Acknowledgements

This work was supported by a grant from UNESCO-IUMS Mircens-SGM short-term fellowship We gratefully thanks Center of National de Reference des Mycobateries, Institute Pasteur, Paris (France) for providing all chemicals and Laboratory facilities.

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