Abstract
Recent advances in culture-independent microbiological analyses have greatly expanded our understanding of the diversity of unculturable microbes. However, human pathogenic bacteria differing significantly from known taxa have rarely been discovered. Here, we present the complete genome sequence of an uncultured bacterium detected in human respiratory tract named IOLA, which was determined by developing a protocol to selectively amplify extremely AT-rich genomes. The IOLA genome is 303,838 bp in size with a 20.7% GC content, making it the smallest and most AT-rich genome among known human-associated bacterial genomes to our best knowledge and comparable to those of insect endosymbionts. While IOLA belongs to order Rickettsiales (mostly intracellular parasites), the gene content suggests an epicellular parasitic lifestyle. Surveillance of clinical samples provides evidence that IOLA can be predominantly detected in patients with respiratory bacterial infections and can persist for at least 15 months in the respiratory tract, suggesting that IOLA is a human respiratory tract-associated bacterium.
Highlights
Recent advances in culture-independent microbiological analyses have greatly expanded our understanding of the diversity of unculturable microbes
To reconstruct the whole genome sequence (WGS) of IOLA, we established a protocol for metagenomic DNA preparation enriched with IOLA DNA (Fig. 1)
By developing a protocol to selectively amplify extremely AT-rich bacterial genomes, we successfully reconstructed the complete IOLA genome sequence and showed that, to the best of our knowledge, IOLA has the smallest genome (304 kb in size) among the human-associated bacterial genomes sequenced to date, followed by M. genitalium (580 kb)[36]
Summary
Recent advances in culture-independent microbiological analyses have greatly expanded our understanding of the diversity of unculturable microbes. Only a few 16S rRNA sequences similar to that of IOLA (>96% identity) have been deposited in the NCBI database (https://www.ncbi.nlm.nih.gov), they were all identified in specimens from patients with respiratory tract disorders, including the first registered IOLA-like 16S rRNA sequence detected in a BALF sample from a cystic fibrosis patient[10]. These data suggest that IOLA may be a previously unknown human respiratory pathogen. The results of a survey of respiratory disease patients using IOLA-specific PCR and an analysis of multiple samples obtained in the survey are presented, which suggest that IOLA can inhabit and persist in the respiratory tract of patients with respiratory disorders
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