International Journal of Mycobacteriology (Jan 2015)
Comparative study between routine and molecular diagnosis of Mycobacterium species isolated from humans and bovines
Abstract
Background: Tuberculosis (TB) is one of the most widespread, infectious, zoonotic diseases and continues to be a leading cause of death. TB is a predominant public health problem in the world. Mycobacterium tuberculosis (MTB) is the most common cause of human TB. M. bovis is the causative agent of this disease in cattle. In this study the human results revealed that from a total 1244 cases, 123 (10%) of the specimens were positive upon direct examination, 173 (14%) by cultivation on LJ medium, and the rest were negative. Fifty positive samples were chosen for direct smear and culturing and were followed-up from the first step of diagnosis to molecular assay; these specimens were taken from 33 (66%) males and 17 (34%) females; fifty positive cases were detected by direct smear. Furthermore, 50 positive cases were also noticed using culturing on Lowenstein–Jensen (LJ) media and 23 positive cases by PCR. The bovine milk sample results showed that 3 positive cases were detected by direct smear and 7 positive cases were observed using culturing on LJ media with sodium pyruvate; the results of culturing methods were confirmed by PCR. Objective: Diagnosis of TB based on phenotypic and genotypic methods using routine methods and PCR by amplification of nucleic acid of MTB. Material and Methods: During the study period of February 2009 to June 2009, the Institute of Chest and Respiratory Diseases – Baghdad received 1244 suspected cases of TB from different Iraqi governorates; 759 (61%) males and 485 (39%) females ranging in age from 2 months to 90 years. Only 50 positive sputum samples were chosen from 173 positive cases for this study and were detected by routine and molecular methods, including polymerase chain reaction. In addition, 68 bovine milk samples were taken from Al-Fthelia – Baghdad during the study period. The results of routine methods were confirmed by PCR. Results: The human samples: The results revealed that from a total 1244 cases, 123 (10%) of the specimens were positive by direct examination and 173 (14%) by cultivation; the rest were negative. The age range of patients was 2 months to 90 years. Distribution of patients among age groups showed that 32 (13.3%) patients were in the 60 years age group. It is obvious that positive male cases had the highest number (113/173) throughout the study period. Fifty positive samples for direct smear and culturing were chosen and followed-up from the first step of diagnosis to molecular assay. These specimens were taken from 33 (66%) males and 17 (34%) females with a range of ages in both sexes; such a selection aimed to represent a variety of backgrounds. The results indicated that all the 50 samples were positive for direct smear (100%) and 50 positive in culture (100%), while 23 (46%) cases were positive for PCR and 27 (54%) cases were negative for PCR. The bovine samples: From a total of 68 milk samples, the positive rate for direct smear was 3 (4.4%), while the positive rates for culture were 7 (10.2%). The positive samples were cultured on LJ slant for 3–6 weeks. The appearance of colonies was typical cream-colored, buff and rough colonies against the green egg-based medium. The isolated colonies were identified as M. bovis by five biochemical tests.
