Fluctuating Bacteriophage-induced galU Deficiency Region is Involved in Trade-off Effects on the Phage and Fluoroquinolone Sensitivity in Pseudomonas aeruginosa

Keisuke Nakamura; Jumpei Fujiki; Tomohiro Nakamura; Takaaki Furusawa; Satoshi Gondaira; Masaru Usui; Hidetoshi Higuchi; Yutaka Tamura; Hidetomo Iwano

doi:10.1016/j.virusres.2021.198596

Fluctuating Bacteriophage-induced galU Deficiency Region is Involved in Trade-off Effects on the Phage and Fluoroquinolone Sensitivity in Pseudomonas aeruginosa

Virus Res. 2021 Dec:306:198596. doi: 10.1016/j.virusres.2021.198596. Epub 2021 Oct 12.

Authors

Keisuke Nakamura¹, Jumpei Fujiki², Tomohiro Nakamura¹, Takaaki Furusawa¹, Satoshi Gondaira³, Masaru Usui⁴, Hidetoshi Higuchi³, Yutaka Tamura⁴, Hidetomo Iwano¹

Affiliations

¹ Laboratory of Veterinary Biochemistry, Department of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan.
² Laboratory of Veterinary Biochemistry, Department of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan. Electronic address: j-fujiki@rakuno.ac.jp.
³ Laboratory of Animal Health, Rakuno Gakuen University School of Veterinary Medicine, Ebetsu, Hokkaido, Japan.
⁴ Laboratory of Food Microbiology and Food Safety, Rakuno Gakuen University School of Veterinary Medicine, Ebetsu, Hokkaido, Japan.

PMID: 34648885
DOI: 10.1016/j.virusres.2021.198596

Abstract

Pseudomonas aeruginosa, which causes chronic infections, has demonstrated rapid acquisition of antimicrobial resistance (AMR). Therefore, bacteriophages have received significant attention as promising antimicrobial agents; however, previous trials have reported the occurrence of phage-resistant variants. P. aeruginosa has lost large chromosomal fragments via evolutionary selection by MutL. Mutants lacking galU and hmgA, located in close proximity, exhibit phage resistance and brown color phenotype since hmgA encodes a homogentisic acid metabolic enzyme and deletion of galU results in a lack of O-antigen polysaccharide and absence of the phage receptor. In the present study, we evaluated this mechanism for controlling phage resistance in P. aeruginosa veterinary isolate Pa12. Phage-resistant Pa12 brown mutants (brmts) with galU and hmgA deletions were isolated. Whole-genome sequencing of the brmts revealed that regions 148-27 kbp upstream and 261-110 kbp downstream of galU were largely deleted from the Pa12 parental chromosome. Furthermore, all of these fluctuating deleted sequences in Pa12 brmts, tentatively designated bacteriophage-induced galU deficiency (BigD) regions, harbor multi-drug efflux system genes (mexXY). Minimum inhibitory concentration (MIC) assays demonstrated that brmts altered sensitivity to antibiotics and exhibited increased levofloxacin sensitivity compared with the Pa12 parent. Orbifloxacin and enrofloxacin also effectively suppressed growth of the Pa12 brmts, suggesting that MexXY, which mediates quinolone efflux and is located in the BigD region, might be associated with restoration of fluoroquinolone sensitivity. Our findings indicate that AMR-related genes in the BigD region could produce trade-off effects between phages and drug sensitivity and thereby contribute to a potential strategy to control and prevent phage-resistant variants in phage therapy.

Keywords: Bacteriophage; Fitness cost; Phage resistance; Phage therapy; Pseudomonas aeruginosa; Trade-off; galU.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Anti-Bacterial Agents / pharmacology
Bacteriophages* / genetics
Fluoroquinolones / metabolism
Fluoroquinolones / pharmacology
HMGA Proteins* / metabolism
Phage Therapy*
Pseudomonas aeruginosa / genetics

Substances

Anti-Bacterial Agents
Fluoroquinolones
HMGA Proteins