Reducing delays in the genomic epidemiology of tuberculosis: a flexible and decentralized analysis of each incident case

Abstract

Background: Genomic analysis has markedly improved our knowledge of the transmission dynamics of M. tuberculosis. The high-throughput analysis provided by short-read platforms in reference laboratories ensures coverage of all TB cases in a population, guaranteeing the identification of complete transmission chains. However, this strategy inevitably leads to delays until genomic data are available at local level. We evaluated an alternative approach based on a case-by-case genomic analysis of primary cultures from each incident case. Methods: The strategy was evaluated in 23 consecutive stain-positive cases diagnosed in Almeria, Spain over a 4-month period (March-July 2023). DNA was purified from primary cultures on LJ medium. Nanopore sequencing and reusage of flow cells was performed. Results: In 52% of cases, sufficient culture growth was obtained to prepare sequencing libraries <20 days after inoculation. Up to two isolates were loaded per run to ensure rapid availability of results. Seven flow cells in all were needed, which were reused up to 6 times. In 61% of isolates, optimal genome coverage (>90% at >20X) was achieved in less than 2.5 hours of sequencing, which enabled new cases to be labeled as clustered (39%), or orphans (61%) on the same day. Nanopore data correlated with those ultimately obtained using high-throughput short-read sequencing. Conclusions: Our strategy could offer a flexible and more rapid decentralized alternative for local settings, accelerating the information on clusters needed to tailor control interventions.