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. 2021 Oct 26;10(11):1301.
doi: 10.3390/antibiotics10111301.

CRISPR Element Patterns vs. Pathoadaptability of Clinical Pseudomonas aeruginosa Isolates from a Medical Center in Moscow, Russia

Marina Tyumentseva  1 , Yulia Mikhaylova  1 , Anna Prelovskaya  1 , Konstantin Karbyshev  1 , Aleksandr Tyumentsev  1 , Lyudmila Petrova  2 , Anna Mironova  2 , Mikhail Zamyatin  2 , Andrey Shelenkov  1 , Vasiliy Akimkin  1
Affiliations

CRISPR Element Patterns vs. Pathoadaptability of Clinical Pseudomonas aeruginosa Isolates from a Medical Center in Moscow, Russia

Marina Tyumentseva et al. Antibiotics (Basel). .

Abstract

Pseudomonas aeruginosa is a member of the ESKAPE opportunistic pathogen group, which includes six species of the most dangerous microbes. This pathogen is characterized by the rapid acquisition of antimicrobial resistance, thus causing major healthcare concerns. This study presents a comprehensive analysis of clinical P. aeruginosa isolates based on whole-genome sequencing data. The isolate collection studied was characterized by a variety of clonal lineages with a domination of high-risk epidemic clones and different CRISPR/Cas element patterns. This is the first report on the coexistence of two and even three different types of CRISPR/Cas systems simultaneously in Russian clinical strains of P. aeruginosa. The data include molecular typing and genotypic antibiotic resistance determination, as well as the phylogenetic analysis of the full-length cas gene and anti-CRISPR genes sequences, predicted prophage sequences, and conducted a detailed CRISPR array analysis. The differences between the isolates carrying different types and quantities of CRISPR/Cas systems were investigated. The pattern of virulence factors in P. aeruginosa isolates lacking putative CRISPR/Cas systems significantly differed from that of samples with single or multiple putative CRISPR/Cas systems. We found significant correlations between the numbers of prophage sequences, antibiotic resistance genes, and virulence genes in P. aeruginosa isolates with different patterns of CRISPR/Cas-elements. We believe that the data presented will contribute to further investigations in the field of bacterial pathoadaptability, including antimicrobial resistance and the role of CRISPR/Cas systems in the plasticity of the P. aeruginosa genome.

Keywords: Pseudomonas aeruginosa; WGS; antibiotic resistance; multiple CRISPR/Cas systems; pathoadaptability; virulence factors.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Figures

Figure 1
Figure 1
Venn diagrams of CRISPR/Cas element distribution for CriePir P. aeruginosa isolates sequenced by us (left) and reference isolates available from Pseudomonas genome DB (right).
Figure 2
Figure 2
Venn diagrams of CRISPR/Cas system type distribution for CriePir (left) and reference (right) P. aeruginosa isolates.
Figure 3
Figure 3
Maximum likelihood phylogenetic tree of full-length csy3 gene sequences of clinical CriePir P. aeruginosa isolates and reference isolates from Pseudomonas genome database. Bootstrap test (1000 replicates) was applied, and bootstrap values are shown at the branch nodes. The ST654 sequences identified in this study are marked with black circle. Sequences identified in this study are indicated by the isolate names and the reference sequences, by GenBank accession number.
Figure 4
Figure 4
Anti-CRISPRs among the clinical CriePir P. aeruginosa isolates from a Moscow medical center with different CRISPR/Cas systems. * indicates the level of significance p < 0.05.
Figure 5
Figure 5
Comparison of the number of AR genes in CriePir and reference (Ref) P. aeruginosa isolates with and without antiCRISPRs. The numbers are given for CriePir vs. reference isolates with antiCRISPRs (A) and without antiCRISPRs (B); with vs. without antiCRISPRs for reference isolates (C) and CriePir isolates (D); *** indicates the level of significance p < 0.001.
Figure 5
Figure 5
Comparison of the number of AR genes in CriePir and reference (Ref) P. aeruginosa isolates with and without antiCRISPRs. The numbers are given for CriePir vs. reference isolates with antiCRISPRs (A) and without antiCRISPRs (B); with vs. without antiCRISPRs for reference isolates (C) and CriePir isolates (D); *** indicates the level of significance p < 0.001.
Figure 6
Figure 6
Comparison of CriePir P. aeruginosa isolates having different CRISPR/Cas systems by the number of CRISPR4 arrays. **** indicates the level of significance p < 0.0001.
Figure 7
Figure 7
The number of active and ambiguous prophages among CriePir P. aeruginosa isolates having no cas cassettes. * indicates the level of significance p < 0.05.
Figure 8
Figure 8
CRISPR array length (A) and spacers count (B) among CriePir P. aeruginosa isolates. ** and **** indicate the increasing levels of significance p < 0.01 and p < 0.0001, respectively.
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