Methylation analysis of Klebsiella pneumoniae from Portuguese hospitals

• Published in: Scientific reports 2021-03, Vol.11 (1), p.6491-10, Article 6491
• Main Authors: Spadar, Anton, Perdigão, João, Phelan, Jody, Charleston, James, Modesto, Ana, Elias, Rita, de Sessions, Paola Florez, Hibberd, Martin L., Campino, Susana, Duarte, Aida, Clark, Taane G.
• Format: Article
• Language: English
• Subjects: 631/208/177; 631/326/22/1434; 631/326/41/2530; Adenine; Antimicrobial resistance; Bacterial Proteins - genetics; beta-Lactam Resistance; Carbapenemase; Chromosomes; Deoxyribonucleic acid; DNA; DNA Methylation; DNA Modification Methylases - genetics; DNA sequencing; Epigenetics; Epigenome; Fosfomycin; Gene transfer; Genomes; Horizontal transfer; Humanities and Social Sciences; Humans; Klebsiella Infections - microbiology; Klebsiella pneumoniae; Klebsiella pneumoniae - drug effects; Klebsiella pneumoniae - genetics; Klebsiella pneumoniae - isolation & purification; Klebsiella pneumoniae - pathogenicity; Methylase; multidisciplinary; N6-methyladenosine; Nosocomial infection; Plasmids; Science; Science (multidisciplinary); Site-specific DNA methyltransferase (adenine-specific); Transposase; β Lactamase
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-021-85724-2

Klebsiella pneumoniae is an important nosocomial infectious agent with a high antimicrobial resistance (AMR) burden. The application of long read sequencing technologies is providing insights into bacterial chromosomal and putative extra-chromosomal genetic elements (PEGEs) associated with AMR, but also epigenetic DNA methylation, which is thought to play a role in cleavage of foreign DNA and expression regulation. Here, we apply the PacBio sequencing platform to eight Portuguese hospital isolates, including one carbapenemase producing isolate, to identify methylation motifs. The resulting assembled chromosomes were between 5.2 and 5.5Mbp in length, and twenty-six PEGEs were found. Four of our eight samples carry bla CTX-M-15 , a dominant Extended Spectrum Beta Lactamase in Europe. We identified methylation motifs that control Restriction–Modification systems, including GATC of the DNA adenine methylase (Dam), which methylates N6-methyladenine (m6A) across all our K. pneumoniae assemblies. There was a consistent lack of methylation by Dam of the GATC motif downstream of two genes: fosA , a locus associated with low level fosfomycin resistance, and tnpB transposase on IncFIB(K) plasmids. Overall, we have constructed eight high quality reference genomes of K. pneumoniae , with insights into horizontal gene transfer and methylation m6A motifs.