Whole genome sequencing-based analysis of tuberculosis (TB) in migrants: Rapid tools for crossborder surveillance and to distinguish between recent transmission in the host country and new importations

Background: The analysis of transmission of tuberculosis (TB) is challenging in areas with a large migrant population. Standard genotyping may fail to differentiate transmission within the host country from new importations, which is key from an epidemiological perspective. Aim: To propose a new strategy to simplify and optimise cross-border surveillance of tuberculosis and to distinguish between recent transmission in the host country and new importations. Methods: We selected 10 clusters, defined by 24-locus mycobacterial interspersed repetitive unitvariable number tandem repeat (MIRU-VNTR), from a population in Spain rich in migrants from eastern Europe, north Africa and west Africa and reanalysed 66 isolates by whole-genome sequencing (WGS). A multiplex-allele-specific PCR was designed to target strain-specific marker single nucleotide polymorphisms (SNPs), identified from WGS data, to optimise the surveillance of the most complex cluster. Results: In five of 10 clusters not all isolates showed the short genetic distances expected for recent transmission and revealed a higher number of SNPs, thus suggesting independent importations of prevalent strains in the country of origin. In the most complex cluster, rich in Moroccan cases, a multiplex allele-specific oligonucleotide-PCR (ASO-PCR) targeting the marker SNPs for the transmission subcluster enabled us to prospectively identify new secondary cases. The ASOPCR- based strategy was transferred and applied in Morocco, demonstrating that the strain was prevalent in the country. Conclusion: We provide a new model for optimising the analysis of cross-border surveillance of TB transmission in the scenario of global migration.