generating-whole-bacterial-genomes-from-clinical-samples-using-target

Generating Whole Bacterial Genomes from Clinical Samples using a Target Enrichment Workflow

Obtaining complete genomes of bacterial sexually transmitted infections (STIs) directly from clinical samples is challenging due to the presence of human ...

University of Zürich

Whole-genome analysis has provided insights into the evolution of  and, recently, into circulating strains that cause lymphogranuloma venereum (LGV). A large LGV outbreak of a new -genotype, L2b, was first reported in Europe in the early 2000s, primarily affecting men who have sex with men (MSM), and then expanded globally. More recent work shows that this outbreak is diversifying into variants of described -genotypes, with the same L2b genomic backbone. This study extends the investigation of LGV cases to Argentina and Finland. In 2017, an LGV outbreak was described in Argentina characterized by distinct genomic features shown by both -genotyping and Multi-Locus Sequence Typing (MLST) analysis. We have obtained whole-genome sequences from cultured isolates and clinical samples via SureSelect (Agilent) target enrichment. Based on  and phylogenetic analyses, we describe further diversity within the -genotype L2b clade, illustrating the transmission dynamics in Argentina and Finland. A key finding is that of a novel clade of Argentinian samples, characterized by a proposed new -genotype L4. Additionally, we present the genome sequence of a non-LGV strain associated with anorectal proctitis. These findings contribute to the investigation of LGV evolution, particularly with the presence of the novel L4 lineage, and provide insights into genomic diversity and transmission dynamics of .

 genomes from rectal samples: description of a new clade comprising -genotype L4 from Argentina.

Rates of bacterial sexually transmitted infections (STIs) are rising, and accessing their genomes provides information on strain evolution, circulating strains and encoded antimicrobial resistance (AMR). Notable pathogens include  (CT),  (NG) and  (TP), globally the most common bacterial STIs.  (formerly )  (MG) is also a bacterial STI that is of concern due to AMR development. These bacteria are also fastidious or hard to culture, and standard sampling methods lyse bacteria, completely preventing pathogen culture. Clinical samples contain large amounts of human and other microbiota DNA. These factors hinder the sequencing of bacterial STI genomes. We aimed to overcome these challenges in obtaining whole-genome sequences and evaluated four approaches using clinical samples from Argentina (39), and Switzerland (14), and cultured samples from Finland (2) and Argentina (1). First, direct genome sequencing from swab samples was attempted through Illumina deep metagenomic sequencing, showing extremely low levels of target DNA, with under 0.01% of the sequenced reads being from the target pathogens. Second, host DNA depletion followed by Illumina sequencing was not found to produce enrichment in these very low-load samples. Third, we tried a selective long-read approach with the new adaptive sequencing from Oxford Nanopore Technologies, which also did not improve enrichment sufficiently to provide genomic information. Finally, target enrichment using a novel pan-genome set of custom SureSelect probes targeting CT, NG, TP and MG followed by Illumina sequencing was successful. We produced whole genomes from 64% of CT-positive samples, from 36% of NG-positive samples and 60% of TP-positive samples. Additionally, we enriched MG DNA to gain partial genomes from 60% of samples. This is the first publication to date to utilize a pan-genome STI panel in target enrichment. Target enrichment, though costly, proved essential for obtaining genomic data from clinical samples. These data can be utilized to examine circulating strains and genotypic resistance and guide public health strategies.

Article	Year
genomes from rectal samples: description of a new clade comprising -genotype L4 from Argentina. Microbial genomics\| PubMed ID: 39943870	2025
Evaluating methods for genome sequencing of and other sexually transmitted bacteria directly from clinical swabs. Microbial genomics\| PubMed ID: 39943872	2025

Vera Bregy

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