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OP-3
IDENTIFICATION OF THE INSERTION ELEMENT IS6110 IN PHOP
PROMOTER IN A HIGH TRANSMISSION Mycobacterium
tuberculosis STRAIN: A CLUE TO PHENOTYPIC VARIATION
Andrea Sandoval 1,2 , Andrés Cubillos 1,2 , Alejandro Reyes 3 , Nidia Correa 2,4 , Jaime Robledo 2,4 , Maria Mercedes Zambrano 1 ,
and Patricia Del Portillo 1,2
1 - Corporación CorpoGen, Bogotá, Colombia.
2 - Centro Colombiano de Investigación en Tuberculosis CCITB, Bogotá, Colombia.
3 - Center for Genome Sciences, Washington University School of Medicine, St. Louis, Missouri, USA
4 - Corporación para Investigaciones Biológicas CIB, Medellín, Colombia
Aim
The insertion element IS6110 can mediate genetic diversity in Mycobacterium tuberculosis (MTB) strains due to its capacity
to move and cause rearrangements, deletions and insertions. Transposition of these elements can therefore affect
gene expression and alter the phenotype of MTB. In this study we analyzed the insertion sites of IS6110 in two clinical
MTB Haarlem genotype strains that present differences in transmissibility as demonstrated in a cohort of patients and
household contacts from Colombia.
Methods
Restriction Fragment Length Polymorphism (IS6110-RFLP) was performed and revealed genomic differences between
the two strains. DNA was isolated, digested with XmaI and ligated to specific adapters designed for this purpose. Ligation
Mediated PCR (LM-PCR) was carried out to amplify the regions flanking IS6110 insertion sites. A library containing the
amplified products was constructed and sequenced clones were mapped against the annotated sequenced genomes. PCR
was used to confirm the sites of insertion.
Results
Twelve different insertions were identified; nine were common to both strains (Rv2336, Rv1754c, Rv0963c, Rv0403c,
Rv1358, Rv2813/DR, Rv2254c, Rv0795 and PPE34), two were specific for the high transmission strain (DR region and
transcriptional regulator phoP), and one was found just in the low transmission strain (PPE46).
Conclusions
LM-PCR allowed us to identify IS6110 flanking regions and localized the genomic differences between two strains with
contrasting transmission pattern. Most of the insertions occur in conserved hypothetical proteins, followed by proteins
involved in cell wall synthesis and cell processes, regulatory proteins and members of the PE/PPE family. The DR region,
considered a hotspot for insertions, had three insertions, two common to both strains and one in the high transmission
strain. Since the phoP gene regulates several functions implicated in virulence, it is possible that the IS6110 insertion in
the phoP promoter could enhance transmission by acting as a portable promoter and inducing gene expression, as has
been reported before in M. bovis.
Acknowledgment
Consorcio Colombiano de Investigación en Tuberculosis, CCITB. 4312004
European Society of Mycobacteriology | 30 th Annual Congress | July 2009 | Porto - Portugal
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