Simple Assay for Detection of the Central Asia Outbreak Clade of the Mycobacterium tuberculosis Beijing Genotype

Abstract

The Central Asia outbreak (CAO) clade is a branch of the Mycobacterium tuberculosis Beijing genotype that is associated with multidrug resistance, increased transmissibility, and epidemic spread in parts of the former Soviet Union. Furthermore, migration flows bring these strains far beyond their areas of origin. We aimed to find a specific molecular marker of the Beijing CAO clade and develop a simple and affordable method for its detection. Based on the bioinformatics analysis of the large M. tuberculosis whole-genome sequencing (WGS) data set (n = 1,398), we identified an IS6110 insertion in the Rv1359-Rv1360 intergenic region as a specific molecular marker of the CAO clade. We further designed and optimized a multiplex PCR method to detect this insertion. The method was validated in silico with the recently published WGS data set from Central Asia (n = 277) and experimentally with M. tuberculosis isolates from European and Asian parts of Russia, the former Soviet Union, and East Asia (n = 319). The developed molecular assay may be recommended for rapid screening of retrospective collections and for prospective surveillance when comprehensive but expensive WGS is not available or practical. The assay may be especially useful in high multidrug-resistant tuberculosis (MDR-TB) burden countries of the former Soviet Union and in countries with respective immigrant communities.

Comparative Genomic Analysis of Mycobacteriaceae Reveals Horizontal Gene Transfer-Mediated Evolution of the CRISPR-Cas System in the Mycobacterium tuberculosis Complex

“…A frequent cause of genomic deletions in bacteria is related to the movement of mobile genetic elements and although insertion of IS 6110 in M. tuberculosis CRISPR-Cas locus has been observed ( 19 ), its impact on the CRISPR-Cas system is poorly understood. Apart from implication of IS 6110 transposition in host adaptation ( 19 ), studying differential insertion sites in diverse strains also has potential use as molecular markers for identifying strain-specific outbreaks, as seen with the Central Asia outbreak (CAO) clade, where a specific IS 6110 insertion was detected unique to this major epidemic clade of Beijing genotype ( 42 ).…”

Genetic Variation Putatively Associated with Mycobacterium tuberculosis Resistance to Perchlozone, a New Thiosemicarbazone: Clues from Whole Genome Sequencing and Implications for Treatment of Multidrug-Resistant Tuberculosis

“…M. tuberculosis isolates (genomes) were assigned to the Beijing genotype and its specific subtypes based on the assessment of the previously described SNP markers and genomic deletions [ 26 , 27 , 28 , 29 , 30 ]. The short sequencing reads (fastq files) were subjected to in silico spoligotyping using TGS-TB online tool at [ 31 ] to deduce their spoligoprofile.…”

Genotyping, Assessment of Virulence and Antibacterial Resistance of the Rostov Strain of Mycobacterium tuberculosis Attributed to the Central Asia Outbreak Clade

“…Spoligotyping and 24 MIRU-VNTR typing were performed as described in references [32,33], respectively. Verification of the presence of SNP in the sigA gene and CAO-specific IS6110 insertion in the Rv1359-Rv1360 intergenic region was performed by PCR as described previously [18,34].…”