Abstract
Pneumonic plague, caused by Yersinia pestis, 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 Y. pestis, 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.
Highlights
Bacteria have successfully evolved sophisticated mechanisms that allow them to sense, cope with, and adapt to varying conditions in their immediate surroundings
We present a novel method for isolating bacterial RNA from the lungs of infected mice during the early stages of pneumonic plague, enabling a comprehensive dual RNA-seq analysis for the subsequent study of host–pathogen interactions
Y. pestis bacteria were grown on BHIA plates (BD, Franklin Lakes, MD, USA) for 48 h at 28 ◦ C, and several colonies were suspended in HIB (BD, Franklin Lakes, MD, USA)
Summary
Bacteria have successfully evolved sophisticated mechanisms that allow them to sense, cope with, and adapt to varying conditions in their immediate surroundings. Often, these mechanisms are essential for the manifestation of virulence in bacterial pathogens [1]. The Gram-negative pathogen Yersinia pestis, the causative agent of plague, has been responsible, in the past, for major pandemics, which caused the death of over 200 million people and still represent a significant public health concern in endemic regions, in the third world. According to the report of the World. Health Organization (WHO), since 2010, there have been hundreds of people that have died of plague. In 2017, 2348 cases of plague were reported in Madagascar, including
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