Comparison between D29 Phage Method, rpoB Gene and Proportion Method for the Detection of Multi Drug Resistant Mycobacterium tuberculosis in Kassala State

Abstract

Tuberculosis is still one of the major health problems worldwide, with high mortality and morbidity. While third of the world population is infected with TB bacilli, the situation is getting worse by the rising emergence of drug resistant strains of Mycobacterium tuberculosis. Conventional method for drug susceptibility testing requires months before results can be reported. However, rapid methods such as phage assay have been developed and recorded as useful tool for quick diagnosis. This study is descriptive cross-sectional laboratory based study which aimed to evaluate the usefulness of phage assay compared to proportional method and PCR in the diagnosis of MDR TB, to estimate the prevalence of MDR TB among tuberculosis patients and to detect the presence of rpoB gene among multidrug resistant isolates. This study was conducted in Kassala State during the period from August 2009 to January 2012. Sputum specimens were collected from ninety acid fast bacilli consented patients (54 males and 36 females). Sputum specimens were processed for direct D29 and culture. All successful cultured isolates were subjected to biochemical tests for phenotypic characterization and further genotypic confirmation was made by amplification of IS 6110. For drug susceptibility testing, proportional method was adopted followed by both indirect D29 and amplification of rpoB gene. The results showed that both males and females in different age groups were infected with TB and those between 21-50 are of the highest infection rate. 21 (23.3%) of the specimens were categorized as rifampicin resistant by direct D29 method, 75/90 (83.3%) of the specimens showed growth on LJ medium similar to MTB complex colonies while 5/90 (3.3%) were identified as rapid growers.60 out of the 75 slow growers (80%) were confirmed as MTB complex members depending on their biochemical characters (PNB, catalase and nitrate reduction). DST result for the 60 MTB isolates showed that 31/60 were drug resistant and that isoniazid compose for the highest percentage of resistance (20/31), followed by rifampicin (19/31) while MDR was detected in 18/60 of the isolates. All the 60 slow growers were confirmed as MTB by their positive IS 6110 results and 15/60 were rpoB positive. In conclusion, the study highlighted the high prevalence of MDR TB in Kassala State. Moreover D29 phage method in its first trial of application in Sudan revealed high sensitivity and specificity, which when combined to its major character of time saving (3 days compared to 70 days in DST) makes it a promising method for rapid uncostly diagnosis of MDR TB.