Metagenomic surveillance for bacterial tick-borne pathogens using nanopore adaptive sampling

Kipp, Evan J.; Lindsey, Laramie L.; Khoo, Benedict; Faulk, Christopher; Oliver, Jonathan D.; Larsen, Peter A.

doi:10.1038/s41598-023-37134-9

Cited by 3 publications

(2 citation statements)

References 58 publications

(61 reference statements)

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“…By sequencing three clinical blood samples originating from human patients showing parasitemia levels between 0.09% and 0.60%, we validated the observed enrichment levels (3.9-, 5.8-, and 2.7-fold) which were consistent with the enrichment observed for the benchmarking samples diluted with 0.1%–8.4% of P. falciparum DNA. Overall, the degree of enrichment for all samples was similar to previous reports where adaptive sampling enriched target DNA two- up to six-fold in various sample types ( 11 , 23 – 25 ).…”

Section: Discussionsupporting

confidence: 87%

Selective whole-genome sequencing of Plasmodium parasites directly from blood samples by nanopore adaptive sampling

De Meulenaere,

Cuypers,

Gauglitz

et al. 2024

mBio

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Whole-genome sequencing of Plasmodium is becoming an increasingly important tool for genomic surveillance of malaria. Due to the predominance of human DNA in a patient blood sample, time-consuming laboratory procedures are required to deplete human DNA or enrich Plasmodium DNA. Here, we investigated the potential of nanopore adaptive sampling to enrich Plasmodium falciparum reads while sequencing unenriched patient blood samples. To compare adaptive sampling versus regular sequencing on a MinION device, a dilution series consisting of 0%–84% P . falciparum DNA in human DNA was sequenced. Half of the flow cell channels were run in adaptive sampling mode, enriching for the P. falciparum reference genome, resulting in a three- to five-fold enrichment of P. falciparum bases in samples containing 0.1%–8.4% P. falciparum DNA. This finding was confirmed by sequencing three P. falciparum patient blood samples with common levels of parasitemia, that is, 0.1%, 0.2%, and 0.6% in adaptive mode. Their estimated enrichment was 5.8, 3.9, and 2.7, respectively, which was sufficient to cover at least 97% of the P. falciparum reference genome at a median depth of 5 (lowest parasitemia) or 355 (highest parasitemia). In all, 38 drug resistance loci were compared to Sanger sequencing results, showing high concordance, which suggests that the obtained sequencing data are of sufficient quality to address common clinical research questions for patients with parasitemias of 0.1% and higher. Overall, our results indicate that adaptive nanopore sequencing has the potential to replace more time-consuming Plasmodium enrichment protocols in the future. IMPORTANCE Malaria is caused by parasites of the genus Plasmodium , and reached a global disease burden of 247 million cases in 2021. To study drug resistance mutations and parasite population dynamics, whole-genome sequencing of patient blood samples is commonly performed. However, the predominance of human DNA in these samples imposes the need for time-consuming laboratory procedures to enrich Plasmodium DNA. We used the Oxford Nanopore Technologies’ adaptive sampling feature to circumvent this problem and enrich Plasmodium reads directly during the sequencing run. We demonstrate that adaptive nanopore sequencing efficiently enriches Plasmodium reads, which simplifies and shortens the timeline from blood collection to parasite sequencing. In addition, we show that the obtained data can be used for monitoring genetic markers, or to generate nearly complete genomes. Finally, owing to its inherent mobility, this technology can be easily applied on-site in endemic areas where patients would benefit the most from genomic surveillance.

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Section: Discussionsupporting

confidence: 87%

Selective whole-genome sequencing of Plasmodium parasites directly from blood samples by nanopore adaptive sampling

De Meulenaere,

Cuypers,

Gauglitz

et al. 2024

mBio

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“…However, with continued development and cost reductions of genomic sequencing technologies (e.g. [19] and applications thereof, such as [20]) alleviating some of the aforementioned challenges, genomic research is poised to rapidly expand our understanding of, and thus ability to respond to, wildlife disease outbreaks for the protection of ecologically and economically important species or populations thereof.…”

Section: Introductionmentioning

confidence: 99%

Metagenomic sequencing sheds light on microbes putatively associated with pneumonia-related fatalities of white-tailed deer (Odocoileus virginianus)

Prentice,

Gilbertson,

Storm

et al. 2024

Microbial Genomics

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With emerging infectious disease outbreaks in human, domestic and wild animal populations on the rise, improvements in pathogen characterization and surveillance are paramount for the protection of human and animal health, as well as the conservation of ecologically and economically important wildlife. Genomics offers a range of suitable tools to meet these goals, with metagenomic sequencing facilitating the characterization of whole microbial communities associated with emerging and endemic disease outbreaks. Here, we use metagenomic sequencing in a case-control study to identify microbes in lung tissue associated with newly observed pneumonia-related fatalities in 34 white-tailed deer (Odocoileus virginianus) in Wisconsin, USA. We identified 20 bacterial species that occurred in more than a single individual. Of these, only Clostridium novyi was found to substantially differ (in number of detections) between case and control sample groups; however, this difference was not statistically significant. We also detected several bacterial species associated with pneumonia and/or other diseases in ruminants (Mycoplasma ovipneumoniae, Trueperella pyogenes, Pasteurella multocida, Anaplasma phagocytophilum, Fusobacterium necrophorum); however, these species did not substantially differ between case and control sample groups. On average, we detected a larger number of bacterial species in case samples than controls, supporting the potential role of polymicrobial infections in this system. Importantly, we did not detect DNA of viruses or fungi, suggesting that they are not significantly associated with pneumonia in this system. Together, these results highlight the utility of metagenomic sequencing for identifying disease-associated microbes. This preliminary list of microbes will help inform future research on pneumonia-associated fatalities of white-tailed deer.

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A roadmap for biomonitoring in the 21st century: Merging methods into metrics via ecological networks

Cuff,

Deivarajan Suresh,

Dopson

et al. 2023

Advances in Ecological Research

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Metagenomic surveillance for bacterial tick-borne pathogens using nanopore adaptive sampling

Cited by 3 publications

References 58 publications

Selective whole-genome sequencing of Plasmodium parasites directly from blood samples by nanopore adaptive sampling

Selective whole-genome sequencing of Plasmodium parasites directly from blood samples by nanopore adaptive sampling

Metagenomic sequencing sheds light on microbes putatively associated with pneumonia-related fatalities of white-tailed deer (Odocoileus virginianus)

A roadmap for biomonitoring in the 21st century: Merging methods into metrics via ecological networks

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