Rapid diagnosis of bacterial meningitis by nanopore 16S amplicon sequencing: A pilot study

Jangsup Moon, Narae Kim, Tae Joon Kim, Jin Sun Jun, Han Sang Lee, Hye Rim Shin, Soon Tae Lee, Keun Hwa Jung, Kyung Il Park, Ki Young Jung, Manho Kim, Sang Kun Lee, Kon Chu

Research output: Contribution to journalArticle

Abstract

Early administration of antibiotics is crucial in the management of bacterial meningitis. Rapid pathogen identification helps to make a definite diagnosis of bacterial meningitis and enables tailored antibiotic treatment. We investigated if the 16S amplicon sequencing performed by MinION, a nanopore sequencer, was capable of rapid pathogen identification in bacterial meningitis. Six retrospective cases of confirmed bacterial meningitis and two prospective cases were included. The initial cerebrospinal fluid (CSF) samples of these patients were used for the experiments. DNA was extracted from the CSF, and PCR was performed on the 16S ribosomal DNA (16S rDNA). Sequencing libraries were prepared using the PCR products, and MinION sequencing was performed for up to 3 h. The reads were aligned to the bacterial database, and the results were compared to the conventional culture studies. Pathogenic bacteria were successfully detected from the CSF by 16S sequencing in all retrospective cases. 16S amplicon sequencing was more sensitive than conventional diagnostic tests and worked properly even in antibiotics-treated samples. MinION sequencing significantly reduced the turnaround time, and even 10 min of sequencing was sufficient for pathogen detection in certain cases. Protocol adjustment could further increase the sensitivity and reduce the turnaround time for MinION sequencing. Finally, the prospective application of MinION 16S sequencing was successful. Nanopore 16S amplicon sequencing is capable of rapid bacterial identification from the CSF of the bacterial meningitis patients. It may have many advantages over conventional diagnostic tests and should therefore be applied in a larger number of patients in the future.

Original languageEnglish
Article number151338
JournalInternational Journal of Medical Microbiology
Volume309
Issue number6
DOIs
StatePublished - 1 Sep 2019

Fingerprint

Nanopores
Bacterial Meningitides
Cerebrospinal Fluid
Anti-Bacterial Agents
Routine Diagnostic Tests
Polymerase Chain Reaction
Ribosomal DNA
Databases
Bacteria
DNA

Keywords

  • 16S rRNA gene
  • Acute bacterial meningitis
  • Amplicon sequencing
  • MinION
  • Pathogen detection

Cite this

Moon, Jangsup ; Kim, Narae ; Kim, Tae Joon ; Jun, Jin Sun ; Lee, Han Sang ; Shin, Hye Rim ; Lee, Soon Tae ; Jung, Keun Hwa ; Park, Kyung Il ; Jung, Ki Young ; Kim, Manho ; Lee, Sang Kun ; Chu, Kon. / Rapid diagnosis of bacterial meningitis by nanopore 16S amplicon sequencing : A pilot study. In: International Journal of Medical Microbiology. 2019 ; Vol. 309, No. 6.
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Rapid diagnosis of bacterial meningitis by nanopore 16S amplicon sequencing : A pilot study. / Moon, Jangsup; Kim, Narae; Kim, Tae Joon; Jun, Jin Sun; Lee, Han Sang; Shin, Hye Rim; Lee, Soon Tae; Jung, Keun Hwa; Park, Kyung Il; Jung, Ki Young; Kim, Manho; Lee, Sang Kun; Chu, Kon.

In: International Journal of Medical Microbiology, Vol. 309, No. 6, 151338, 01.09.2019.

Research output: Contribution to journalArticle

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T1 - Rapid diagnosis of bacterial meningitis by nanopore 16S amplicon sequencing

T2 - A pilot study

AU - Moon, Jangsup

AU - Kim, Narae

AU - Kim, Tae Joon

AU - Jun, Jin Sun

AU - Lee, Han Sang

AU - Shin, Hye Rim

AU - Lee, Soon Tae

AU - Jung, Keun Hwa

AU - Park, Kyung Il

AU - Jung, Ki Young

AU - Kim, Manho

AU - Lee, Sang Kun

AU - Chu, Kon

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AB - Early administration of antibiotics is crucial in the management of bacterial meningitis. Rapid pathogen identification helps to make a definite diagnosis of bacterial meningitis and enables tailored antibiotic treatment. We investigated if the 16S amplicon sequencing performed by MinION, a nanopore sequencer, was capable of rapid pathogen identification in bacterial meningitis. Six retrospective cases of confirmed bacterial meningitis and two prospective cases were included. The initial cerebrospinal fluid (CSF) samples of these patients were used for the experiments. DNA was extracted from the CSF, and PCR was performed on the 16S ribosomal DNA (16S rDNA). Sequencing libraries were prepared using the PCR products, and MinION sequencing was performed for up to 3 h. The reads were aligned to the bacterial database, and the results were compared to the conventional culture studies. Pathogenic bacteria were successfully detected from the CSF by 16S sequencing in all retrospective cases. 16S amplicon sequencing was more sensitive than conventional diagnostic tests and worked properly even in antibiotics-treated samples. MinION sequencing significantly reduced the turnaround time, and even 10 min of sequencing was sufficient for pathogen detection in certain cases. Protocol adjustment could further increase the sensitivity and reduce the turnaround time for MinION sequencing. Finally, the prospective application of MinION 16S sequencing was successful. Nanopore 16S amplicon sequencing is capable of rapid bacterial identification from the CSF of the bacterial meningitis patients. It may have many advantages over conventional diagnostic tests and should therefore be applied in a larger number of patients in the future.

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KW - Acute bacterial meningitis

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KW - MinION

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