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Sobenna George
TOWARDS A MORE EFFICIENT METHOD FOR DETECTING M. tb INFECTION: A
PRELIMINARY SOUTH AFRICAN STUDY
Sobenna George, Dr. Gerhard Walzl and Ms. Laurianne Van der Merwe and Dr. Anna Mandalakas
Division of Immunology in the Department of Biomedical Sciences
Case Western Reserve University and Stellenbosch University, South Africa
In 2006, approximately 9.2 million people contracted and 1.7 million people died from Mycobacterium
Tuberculosis (M. tb) worldwide. The tuberculosis burden was especially felt in resource-limited countries as 12
African countries made the top 15 list of countries with the highest incidence rates of the disease (1).
Early detection and treatment of M. tb infection are integral to the effective control of the disease. Until recently,
one of the main tools used to detect M.tb infection was the tuberculin skin test (TST). The major drawbacks of this
test are its limited specificity and sensitivity (2). T-cell based Interferon (IFN)-γ release assays (IGRAs) are
relatively newer methods of detecting M. tb infection that provide some of the solutions to the problems of the skin
test. However, limited research has been done on the performance of IGRAs in children less than 17 years old (3).
It is possible that IGRA outcomes may be affected by INF- γ gene expression. Since gene expression investigations
are dependent on ribonucleic acid (RNA) extraction, this preliminary study focused on identifying a suitable
method for RNA extraction from the limited blood samples that can be taken from children. One aim of this study
was to test the reproducibility of the method of RNA extraction outlined by Carrol et al. (4) on pediatric blood
samples, using the PAXgene TM Blood RNA System (PreAnalytiX, QIAGEN, Germany). The second aim of the
study was to test another easily accessible RNA extraction kit, the QIAamp ® RNA blood mini kit (QIAgen Ltd.),
also on pediatric blood samples
Neither extraction kit was optimized for the small blood volumes used as they each yielded very small quantities
of RNA. Although the RNA yields from the Carrol et al. (4) study were not reproduced, there may still be some
merit in optimizing the method for the PAXgene TM Blood RNA System (PreAnalytiX, QIAGEN, Germany) since
this kit consistently extracted larger quantities of RNA than the QIAamp ® RNA blood mini kit (QIAgen Ltd.).
REFERENCES:
1. World Health Organization. Global Tuberculosis Control 2008: Surveillance, planning, financing [cited
September 1, 2008]; Available from: http://www.who.int/tb/publications/global_report/en/
2. Lalvani, A. Diagnosing tuberculosis infection in the 21 st century: New tools to tackle an old enemy. Chest,
2007. 131(6).
3. Powell, D., and Hunt, G. Tubereculosis in children:uAn Update. Advances in Pediatrics, 2006. 53: p. 279-
322.
4. Carrol, E.D., Salway, F., Pepper, S.D., Saunders, E., Mankhmabo, L.A., Ollier, W.E., Hart, C.A., and Day,
P. Successful downstream application of the Paxgene Blood RNA system from small blood samples in
paediatric patients for quantitative PCR analysis. BMC Immunol., 2007;8: p. 20
This study was funded in part by the Minority HIV Training Program of The Center for AIDS of Case Western
Reserve University/ University Hospitals of Cleveland and by the Crile Summer Research Fellowships
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