Deciphering the sequential events during in vivo acquisition of drug resistance in Mycobacterium tuberculosis

ABSTRACT

Tuberculosis [TB] is caused by Mycobacterium tuberculosis [MTB] and the disease has remained a major health problem in most of the developing countries, particularly after the emergence of multidrug-resistant TB [MDR-TB]. The MDR-TB is an intriguing subject and very little is known about the in vivo processes which take place during the acquisition of MDR. This study describes a unique case of pulmonary TB [PTB] from which four sequential isolates of MTB could be isolated while the patient was on anti-tubercular treatment. The first baseline isolate was sensitive to all drugs, but the subsequent three isolates acquired resistance to multiple drugs and finally the patient died after 27 months post-diagnosis when his fourth isolate became resistant to isoniazid, rifampicin, ethambutol and kanamycin. All sequential cultures were identified as MTB using conventional and molecular methods, including 16s RNA sequencing and the spoligotyping. Spoligotyping followed by comparison with SITVITWEB database revealed that all the isolates belonged to the family of the Central Asian Strain Delhi [CAS1_Delhi, ST26] genotype, and no cross or mixed infections were observed. The drug resistance was further characterized at the molecular level by sequencing the target genes [katG, inhA, rpoB, embB, eis promoter region and rrs]. The results revealed mutated alleles associated with resistance to the respective drugs. This unique case indicates that it is possible to isolate MTB during treatment if the strain is acquiring resistance. The data presented from four sequential isolates provides an insight into what sequential genetic and proteomic changes occur in the bacteria during the in vivo acquisition of MDR.