Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection

Pneumonic plague, caused by <i>Yersinia pestis,</i> is a rapidly progressing lethal infection. The various phases of pneumonic plague are yet to be fully understood. A well-established way to address the pathology of infectious diseases in general, and pneumonic plague in particular, is...

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Main Authors: Ofir Israeli, Inbar Cohen-Gihon, Moshe Aftalion, David Gur, Yaron Vagima, Ayelet Zauberman, Yinon Levy, Anat Zvi, Theodor Chitlaru, Emanuelle Mamroud, Avital Tidhar
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Language:English
Published: MDPI AG 2021-10-01
Series:Microorganisms
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Online Access:https://www.mdpi.com/2076-2607/9/10/2166
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author Ofir Israeli
Inbar Cohen-Gihon
Moshe Aftalion
David Gur
Yaron Vagima
Ayelet Zauberman
Yinon Levy
Anat Zvi
Theodor Chitlaru
Emanuelle Mamroud
Avital Tidhar
author_facet Ofir Israeli
Inbar Cohen-Gihon
Moshe Aftalion
David Gur
Yaron Vagima
Ayelet Zauberman
Yinon Levy
Anat Zvi
Theodor Chitlaru
Emanuelle Mamroud
Avital Tidhar
author_sort Ofir Israeli
collection DOAJ
description Pneumonic plague, caused by <i>Yersinia pestis,</i> is a rapidly progressing lethal infection. The various phases of pneumonic plague are yet to be fully understood. A well-established way to address the pathology of infectious diseases in general, and pneumonic plague in particular, is to conduct concomitant transcriptomic analysis of the bacteria and the host. The analysis of dual RNA by RNA sequencing technology is challenging, due the difficulties of extracting bacterial RNA, which is overwhelmingly outnumbered by the host RNA, especially at the critical early time points post-infection (prior to 48 h). Here, we describe a novel technique that employed the infusion of an RNA preserving reagent (RNAlater) into the lungs of the animals, through the trachea, under deep anesthesia. This method enabled the isolation of stable dual mRNA from the lungs of mice infected with <i>Y. pestis</i>, as early as 24 h post-infection. The RNA was used for transcriptomic analysis, which provided a comprehensive gene expression profile of both the host and the pathogen.
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spelling doaj.art-a797869f328d426e8f385e653ac08a352023-11-22T19:15:25ZengMDPI AGMicroorganisms2076-26072021-10-01910216610.3390/microorganisms9102166Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary InfectionOfir Israeli0Inbar Cohen-Gihon1Moshe Aftalion2David Gur3Yaron Vagima4Ayelet Zauberman5Yinon Levy6Anat Zvi7Theodor Chitlaru8Emanuelle Mamroud9Avital Tidhar10Department of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelDepartment of Biochemistry and Molecular Genetics, Israel Institute for Biological Research, Ness-Ziona 74100, IsraelPneumonic plague, caused by <i>Yersinia pestis,</i> is a rapidly progressing lethal infection. The various phases of pneumonic plague are yet to be fully understood. A well-established way to address the pathology of infectious diseases in general, and pneumonic plague in particular, is to conduct concomitant transcriptomic analysis of the bacteria and the host. The analysis of dual RNA by RNA sequencing technology is challenging, due the difficulties of extracting bacterial RNA, which is overwhelmingly outnumbered by the host RNA, especially at the critical early time points post-infection (prior to 48 h). Here, we describe a novel technique that employed the infusion of an RNA preserving reagent (RNAlater) into the lungs of the animals, through the trachea, under deep anesthesia. This method enabled the isolation of stable dual mRNA from the lungs of mice infected with <i>Y. pestis</i>, as early as 24 h post-infection. The RNA was used for transcriptomic analysis, which provided a comprehensive gene expression profile of both the host and the pathogen.https://www.mdpi.com/2076-2607/9/10/2166dual RNA-seqpneumonic plague<i>Y. pestis</i>mouse modelinfection
spellingShingle Ofir Israeli
Inbar Cohen-Gihon
Moshe Aftalion
David Gur
Yaron Vagima
Ayelet Zauberman
Yinon Levy
Anat Zvi
Theodor Chitlaru
Emanuelle Mamroud
Avital Tidhar
Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
Microorganisms
dual RNA-seq
pneumonic plague
<i>Y. pestis</i>
mouse model
infection
title Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
title_full Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
title_fullStr Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
title_full_unstemmed Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
title_short Novel RNA Extraction Method for Dual RNA-seq Analysis of Pathogen and Host in the Early Stages of <i>Yersinia pestis</i> Pulmonary Infection
title_sort novel rna extraction method for dual rna seq analysis of pathogen and host in the early stages of i yersinia pestis i pulmonary infection
topic dual RNA-seq
pneumonic plague
<i>Y. pestis</i>
mouse model
infection
url https://www.mdpi.com/2076-2607/9/10/2166
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