Study On Mycobacterium Tuberculosis And KatG Gene In Sputum

Objective To rapidly identify the category of mycobacterium in sputum with acid-fast positive in luzhou area and to investigate the mutations of katG gene of mycobacterium tuberculosis (MTB). Methods 1.The sputa of 120 patients of pulmonary tuberculosis with positive acid-fast staining were studied in this paper, which come from 18 tuberculosis patients with complications and 102 patients with no complications.2. Amplified the insertion sequence 6110(IS6110) that was specific differentiated non-tuberculosis mycobacteria (NTM) from the mycobacterium tuberculosis complex (MTC) groups.3. katG gene were amplified by PCR reaction and detected with agarose gel by electrophoresed. Compared the katG gene sequence of samples with that of the standard strains of mycobacterium tuberculosis H37RV. Results 1. IS6110 gene was not detected in 13(10.83%,13/120) samples with positive acid-fast staining.There are 6 cases and 7 cases in initial treatment and retreatment group respectively. The ratio of detection of IS6110 gene is no significant difference in two groups. The ratio of detection of IS6110 gene of group with complications is higher than that of group without complications.2. katG gene mutations were detected in 25 samples in 107 IS6110 positive sputa. There is no significant difference of ratio of katG mutations between initial treatment group and retreatment group, but the ratio of katG mutations is higher in group of taking INH over 6 months than that of taking INH within 6 months.3. There are 24 single nucleotide mutation including 14 cases point mutation of 315 codon (9AGC→ACC,3AGC→ATC,2AGC→AAC),5 cases point mutation of 317(ATC→GTC),2 cases point mutation of 304 (GAC→GAG),3 cases point mutation of 270 (CAC→CAT) in this paper, and double nucleotide substitution (270,271 ACT→GTT) were detected in one case; All mutations were located in the isoniazid (INH) resistance mutable gene region. Conclusion 1.Specific IS6110 of mycobacterium tuberculosis was not amplified in 13 samples (10.83%,13/120) of patients with clinical pulmonary tuberculosis, and which suggested some people may be infected by non-tuberculous mycobacteria in this area.2. The ratio of NTM infection of patients with low immunity or long-term use of immunosuppressant is higher than that of tuberculosis patients without complications.3. The mutations of katG gene were found in 25(23.36%, 25/120) patients'sputa with pulmonary tuberculosis in this region, and all mutations occurred in the isoniazid mutable region, which suggested some MTB may be resistance to isoniazid in this region. The ratio of mutation is higher in patients with long-time (>6 months) isoniazid therapy than that of patients with short-time (<6 months). Clinicians should pay more attention to the time of isoniazid administration, and adjusted chemotherapy in time. 4.315 mutation of katG gene is the main mechanism of MTB resistance to INH in luzhou.5. Detection of MTB by molecular biology techniques in sputum specimens is a simple and rapid method which can be achieved rapid diagnosis and help us to choose chemotherapy drug and prevent from spread of resistance in Mycobacterium tuberculosis.