Sequencing

11th Annual Sequencing, Finishing, and Analysis in the Future Meeting
HUMAN GUT ANTIMICROBIAL RESISTANCE: A
COMPARISON OF MICROARRAY, TARGETED
SEQUENCING AND DEEP METAGENOMICS
SEQUENCING
Wednesday, 1st June 11:00 La Fonda Ballroom Talk (OS‐2.01)
Tom Slezak 1 , Tom Brettin 2 , Dionysios Antonopoulos 2 , Sarah Owens 2 , Kenneth Frey 3 , Shea
Gardner 1 , Jonathan Allen 1 , Sam Minot 4 , Nick Greenfield 4 , Nisha Mulakken 5 , Rongsu Qi 5 ,
Chengya Liang 5 , Gary Vora 3 , Gary An 6
1 Lawrence Livermore National Laboratory, 2 Argonne National Laboratory, 3 Naval Medical
Research Center, 4 One Codex, 5 Thermo Fisher, 6 University of Chicago
Patients in Intensive care units (ICUs) are particularly vulnerable to infections from antimicrobial
resistant (AMR) organisms. These patients often receive multiple courses of broad‐spectrum
antibiotics and given the role of fecal auto‐contamination in nosocomial infections we posit that
characterizing the AMR determinants in their gut microbiomes can inform the timely construction
of antibiotic regimens to limit the emergence of clinically significant AMR organisms. We report on
an early‐stage pilot program that evaluated the ability of 3 technologies (microarray, targeted
sequencing, and deep metagenomics whole‐genome shotgun (WGS) sequencing) to detect a key set
of over 500 AMR genes in fecal microbiome samples obtained from three long‐term ICU patients
and three healthy volunteers. Microbiome structure and diversity was also characterized via 16S
rRNA‐based amplicon sequencing. We used the set of AMR genes detected by the Antimicrobial
Resistance Determinant Microarray (ARDM) developed by the Naval Research Laboratory (NRL)
and developed an equivalent AmpliSeq® targeted amplification panel (1,354 total amplicons) to test
these samples. Deep WGS (approximately 200 million reads for each healthy subject and 60 million
reads for each ICU patient) was performed as a comparison using the Illumina HiSeq and as a reference
dataset for non‐targeted sequences. The results demonstrated that the targeted sequencing
consistently detected more AMR genes than WGS, in addition to being faster and less expensive
(multiple targeted samples were run on a single Thermo Fisher Ion PGM or S5 run, compared to
using a full HiSeq lane per samples in the WGS set). A cloud‐based analysis and reporting package
was prototyped and applied to both the targeted and WGS sequence data. We will discuss the
sequencing and array results in detail and make a case for the clinical utility of targeted sequencing
for known genes present in complex tissue/body fluid samples, both to impact the treatment of
individual patients and reduce the emergence of epidemiological foci of AMR within the hospital.
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