Introduction

Understanding the mechanisms by which microbes survive in their host intracellular environments is an essential precursor to developing novel intervention strategies that might exploit the interplay between host and microbe. Transcriptional profiling allows us a glimpse of these molecular interactions and promises to reveal much of this dialogue in both pathogenic and symbiotic settings. Such studies will underpin the development of antivirulence strategies for new therapeutics. This chapter aims to describe the challenges of studying microbial RNA expression from intracellular and in vivo scenarios, and to detail a fraction of the insights acquired from this rapidly expanding field, driven by the recent availability of microbe whole-genome sequence information.

Microbial intracellular gene expression patterns may be used not only to recognize mechanisms vital for survival during infection, but also as a bioprobe to define the host microenvironments encountered. Furthermore, intracellular RNA profiling may decipher how these interactions change over time or with different genetic backgrounds of both host and microbe. This nonreductionist approach of profiling host-microbe crosstalk at the RNA, protein and metabolic levels will contribute to a systems biology view of microbial colonization. The major methodological bottlenecks and applications of these expression datasets are summarized in Figure 9.1, with each step broadly translating into a section of this chapter. We present examples of the use of bacterial and protozoan mRNA expression studies in commensal, mutualistic and parasitic scenarios, with a particular focus on the pathogen Mycobacterium tuberculosis, most familiar to the authors.

Figure 9.1 A cartoon illustrating the bottlenecks and applications of RNA profiling from intracellular microbes. The gene expression pattern of microbes intracellularly or in vivo may be interrogated (1) to identify gene products or metabolic states necessary for successful

microbial infection; (2) as a bioprobe, to define the conditions confronting microbes during infection; or (3) to investigate the role of individual or multiple genes in the changing relationship between microbe and host.

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