Biology subjects

Joseph, S. J.

Publications and source records attributed to Joseph, S. J..

3 recordsLinked to original sources

bioRxiv2021

Global emergence and dissemination of Neisseria gonorrhoeae ST-9363 isolates with reduced susceptibility to azithromycin

Neisseria gonorrhoeae multi-locus sequence type (ST) 9363 genogroup isolates have been associated with reduced azithromycin susceptibility (AZMrs) and show evidence of clonal expansion in the U.S. Here we analyze a global collection of ST-9363 genogroup genomes to shed light on the emergence and dissemination of this strain. The global population structure of ST-9363 genogroup falls into three lineages: Basal, European, and North American; with 32 clades within all lineages. Although, ST-9363 genogroup is inferred to have originated from Asia in the mid-19th century; we estimate the three modern lineages emerged from Europe in the late 1970s to early 1980s. The European lineage appears to have emerged and expanded from around 1986 to 1998, spreading into North America and Oceania in the mid-2000s with multiple introductions, along with multiple secondary reintroductions into Europe. Our results suggest two separate acquisition events of mosaic mtrR and mtrR promoter alleles: first during 2009-2011 and again during the 2012-2013 time, facilitating the clonal expansion of this genogroup with AZMrs in the U.S. By tracking phylodynamic evolutionary trajectories of clades that share distinct demography as well as population-based genomic statistics, we demonstrate how recombination and selective pressures in the mtrCDE efflux operon granted a fitness advantage to establish ST-9363 as a successful gonococcal lineage in the U.S. and elsewhere. Although it is difficult to pinpoint the exact timing and emergence of this young genogroup, it remains critically important to continue monitoring it, as it could acquire additional resistance markers.

microbiology↗

bioRxiv2021

Selective whole genome amplification as a tool to enrich specimens with low Treponema pallidum genomic DNA copies for whole genome sequencing

Downstream next generation sequencing (NGS) of the syphilis spirochete Treponema pallidum subspecies pallidum (T. pallidum) is hindered by low bacterial loads and the overwhelming presence of background metagenomic DNA in clinical specimens. In this study, we investigated selective whole genome amplification (SWGA) utilizing multiple displacement amplification (MDA) in conjunction with custom oligonucleotides with an increased specificity for the T. pallidum genome, and the capture and removal of CpG-methylated host DNA using the NEBNext(R) Microbiome DNA Enrichment Kit followed by MDA with the REPLI-g Single Cell Kit as enrichment methods to improve the yields of T. pallidum DNA in isolates and lesion specimens from syphilis patients. Sequencing was performed using the Illumina MiSeq v2 500 cycle or NovaSeq 6000 SP platform. These two enrichment methods led to 93-98% genome coverage at 5 reads/site in 5 clinical specimens from the United States and rabbit propagated isolates, containing >14 T. pallidum genomic copies/l of sample for SWGA and >129 genomic copies/l for CpG methylation capture with MDA. Variant analysis using sequencing data derived from SWGA-enriched specimens, showed that all 5 clinical strains had the A2058G mutation associated with azithromycin resistance. SWGA is a robust method that allows direct whole genome sequencing (WGS) of specimens containing very low numbers of T. pallidum, which have been challenging until now. ImportanceSyphilis is a sexually transmitted, disseminated acute and chronic infection caused by the bacterial pathogen Treponema pallidum subspecies pallidum. Primary syphilis typically presents as single or multiple mucocutaneous lesions, and if left untreated, can progress through multiple stages with varied clinical manifestations. Molecular studies often rely on direct amplification of DNA sequences from clinical specimens; however, this can be impacted by inadequate samples due to disease progression or timing of patients seeking clinical care. While genotyping has provided important data on circulating strains over the past two decades, WGS data is needed to better understand strain diversity, perform evolutionary tracing, and monitor antimicrobial resistance markers. The significance of our research is the development of a SWGA DNA enrichment method that expands the range of clinical specimens that can be directly sequenced to include samples with low numbers of T. pallidum.

genomics↗

bioRxiv2020

Whole-Genome Enrichment and Sequencing of Chlamydia trachomatis Directly from Patient Clinical Vaginal and Rectal Swabs

Chlamydia trachomatis is the most prevalent cause of bacterial sexually transmitted infections (STIs) worldwide. U.S. cases have been steadily increasing for more than a decade in both the urogenital tract and rectum. C. trachomatis is an obligate intracellular bacterium that is not easily cultured, limiting the capacity for genome studies to understand strain diversity and emergence among various patient populations globally. While Agilent SureSelectXT target-enrichment RNA bait libraries have been developed for whole-genome enrichment and sequencing of C. trachomatis directly from clinical urine, vaginal, conjunctival and rectal samples, efficiencies are only 60-80% for [≥]95-100% genome coverage. We therefore re-designed and expanded the RNA bait library to augment enrichment of the organism from clinical samples to improve efficiency. We describe the expanded library, the limit of detection for C. trachomatis genome copy input, and the 100% efficiency and high-resolution of generated genomes where genomic recombination among paired vaginal and rectal specimens from four patients was identified. This workflow provides a robust approach for discerning genomic diversity and advancing our understanding of the molecular epidemiology of contemporary C. trachomatis STIs across sample types, among geographic populations, sexual networks, and outbreaks associated with proctitis/proctocolitis among women and men who have sex with men. ImportanceChlamydia trachomatis is an obligate intracellular bacterium that is not easily cultured, and there is limited information on rectal C. trachomatis transmission and its impact on morbidity. To improve efficiency of previous studies involving whole genome target enrichment and sequencing of C. trachomatis directly from clinical urine, vaginal, conjunctival, and rectal specimens, we expanded the RNA bait library to augment enrichment of the organism from clinical samples. We demonstrate an increased efficiency in the percentage of reads mapping to C. trachomatis. We show the new system is sensitive for near identical genomes of C. trachomatis from two body sites in four women. Further, we provide a robust genomic epidemiologic approach to advance our understanding of C. trachomatis strains causing ocular, urogenital and rectal infections, and to explore geo-sexual networks, outbreaks of colorectal infections among women and men who have sex with men, and the role of these strains in morbidity.

genomics↗