Pseudomonas aeruginosa MutL promotes large chromosomal deletions through non-homologous end joining to prevent bacteriophage predation

Shen, Mengyu; Zhang, Huidong; Shen, Wei; Zou, Zhirong; Lu, Shuguang; Li, Gang; He, Xuesong; Agnello, Melissa; Shi, Wenyuan; Hu, Fuquan; Le, Shuai

doi:10.1093/nar/gky160

Cited by 56 publications

(63 citation statements)

References 60 publications

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“…Additionally, pyomelanin-producing mutants are regularly isolated from infections; up to 13% of CF patients harbour pyomelanin-producing mutants (52), likely because the production of pyomelanin increases resistance to oxidative stress and persistence in chronic lung infections (22). Recent work has also shown that these mutations can be selected to prevent bacteriophage predation (53). Notably, melanogenic clinical isolates of P. aeruginosa present large chromosomal deletions, similar to those reported in the present work (54).…”

Section: Discussionmentioning

confidence: 99%

Mutation-driven evolution of Pseudomonas aeruginosa in the presence of either ceftazidime or ceftazidime/avibactam

Sanz-García

Hernando-Amado

Martínez

2018

Preprint

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12Ceftazidime/avibactam is a combination of beta-lactam/beta-lactamases inhibitor, which 13 use is restricted to some clinical cases including cystic fibrosis patients infected with 14 multidrug resistant Pseudomonas aeruginosa, in which mutation is the main driver of 15 resistance. This study aims to predict the mechanisms of mutation-driven resistance that 16 are selected for when P. aeruginosa is challenged with either ceftazidime or 17 ceftazidime/avibactam. For this purpose, P. aeruginosa PA14 was submitted to 18 experimental evolution in the absence of antibiotics and in the presence of increasing 19 concentrations of ceftazidime or ceftazidime/avibactam for 30 consecutive days. Final 20 populations were analysed by whole-genome sequencing. All evolved populations 21 reached similar levels of ceftazidime resistance. Besides, all of them were more 22 susceptible to amikacin and produced pyomelanin. A first event in the evolution was the 23 selection of large chromosomal deletions containing hmgA (involved in pyomelanin 24 production), galU (involved in β-lactams resistance) and mexXY-oprM (involved in 25 aminoglycoside resistance). Besides mutations in mpl and dacB that regulate β-26 lactamase expression, mutations related to MexAB-OprM overexpression were 27 prevalent. Ceftazidime/avibactam challenge selected mutants in the putative efflux 28 pump PA14_45890-45910 and in a two-component system (PA14_45870-45880), likely 29 regulating its expression. All populations produce pyomelanin and were more 30 susceptible to aminoglycosides likely due to the selection of large chromosomal 31 deletions. Since pyomelanin-producing mutants, presenting similar deletions are 32 regularly isolated from infections, the potential aminoglycosides hyper-susceptiblity and 33 reduced β-lactams susceptibility of pyomelanin-producing P. aeruginosa should be 34 taken into consideration for treating infections by these isolates. 35 42 including those encoding multidrug (MDR) efflux pumps (4) and beta-lactamases. In 43 addition, an increasing number of P. aeruginosa isolates has acquired several resistance 44 genes through horizontal gene transfer (HGT), including different classes of 45carbapenemases. Finally, P. aeruginosa is able to develop resistance through mutation, 46 particularly when causing chronic infections, to nearly any available antibiotic. Under 47 this situation, the emergence and spread of MDR resistant global clones is of special 48 concern (5). 49The use of β-lactam/β-lactamase inhibitor combinations, such as amoxicillin/clavulanic 50 acid or ceftolozane/tazobactam, has proven to be effective against class A β-lactamases 51 (which include narrow and extended-spectrum β-lactamases and some 52 carbapenemases); whereas effective combinations against class B, C (extended 53 spectrum cephalosporinases) and D β-lactamases (6-8) have not been available until 54 recently. One of them is the ceftazidime/avibactam combination, whose use was 55 approved in 2015 by the FDA (9). 56 Avibactam, formerly known as NXL104, be...

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Section: Discussionmentioning

confidence: 99%

Mutation-driven evolution of Pseudomonas aeruginosa in the presence of either ceftazidime or ceftazidime/avibactam

Sanz-García

Hernando-Amado

Martínez

2018

Preprint

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“…Pseudomonas aeruginosa develops phage resistance through several mechanisms, including modification or loss of the O-antigen component of lipopolysaccharide (LPS), or glycosylation of its type IV pilus (Harvey et al, 2018;Shen et al, 2018). Previously, we identified two types of phageresistant P. aeruginosa mutants following infection with the dsDNA phages PaP1 or PaoP5, based upon colony pigmentation ( Figure 1A).…”

Section: Introductionmentioning

confidence: 99%

“…Previously, we identified two types of phageresistant P. aeruginosa mutants following infection with the dsDNA phages PaP1 or PaoP5, based upon colony pigmentation ( Figure 1A). The mutant designated PAO1r-1, with a brown colony phenotype, contained a large chromosomal deletion including the genes galU and hmgA, which respectively, resulted in the complete loss of O-antigen and the accumulation of characteristic brown-colored homogentisic acids, while conferring phage resistance (Le et al, 2014;Shen et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

Development of a Bacteriophage Cocktail to Constrain the Emergence of Phage-Resistant Pseudomonas aeruginosa

Shen²,

et al. 2020

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With the emergence of multidrug-resistant and extensively drug-resistant bacterial pathogens, phage therapy and other alternative or additional therapeutic modalities are receiving resurgent attention. One of the major obstacles in developing effective phage therapies is the evolution of phage resistance in the bacterial host. When Pseudomonas aeruginosa was infected with a phage that uses O-antigen as receptor, phage resistances typically achieved through changing or loss of O-antigen structure. In this study, we showed that dsRNA phage phiYY uses core lipopolysaccharide as receptor and therefore efficiently kills the O-antigen deletion mutants. Furthermore, by phage training, we obtained PaoP5-m1, a derivative of dsDNA phage PaoP5, which is able to infect mutants with truncated O-antigen. We then generated a cocktail by mixing phiYY and PaoP5-m1 with additional three wide host range P. aeruginosa phages. The phage cocktail was effective against a diverse selection of clinical isolates of P. aeruginosa, and in the short-term constrained the appearance of the phage-resistant mutants that had beleaguered the effectiveness of single phage. Resistance to the 5phage cocktail emerges after several days, and requires mutations in both wzy and migA Thus, this study provides an alternative strategy for designing phage cocktail and phage therapy.

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“…The PA5oct-resistant isolates subjected to typing with a panel of phages recognising different receptors, showed that the phenotypic changes under the influence of PA5oct pressure went beyond the modulation of a single receptor. PA5oct-resistant isolates showed a cross-resistance to phages belonging to different taxonomic units, and the typing patterns were diverse, suggesting genomic rearrangements in the bacterial genome which impact multiple receptors (Le et al , 2014; Shen et al , 2018). These changes in the genome of the host are currently under investigation by our team.…”

Section: Discussionmentioning

confidence: 99%

Pseudomonas aeruginosa PA5oct jumbo phage impacts planktonic and biofilm population and reduces its host virulence

Olszak

Danis-Wlodarczyk

Arabski

et al. 2018

Preprint

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SummaryIn this work we analyzed the impact of jumbo phage PA5oct on the planktonic, cell line adhered, and biofilm population of P. aeruginosa. PA5oct has a broad host-range, able to infect up to 40% of our clinical P. aeruginosa Cystic Fibrosis (CF) collection. In the airway surface liquid (ASL) model, the infection of PA5oct effectively reduced the bacterial population both adhered to epithelial cells, mucus entrapped, and dispersed. The explanation for its infectivity can also be linked to the sensitization of infected bacteria to the innate immune mechanisms and pro-inflammatory effect. Interferometry of a 72-hour old biofilm highlighted the contribution of PA5oct in biofilm matrix degradation. Interestingly, two virion-associated proteins, gp162 and gp205, have been found as putative enzymes that can degrade matrix exopolysaccharides. Two third of biofilm clones developed PA5oct phage-resistance and the cross-resistance to both LPS- and pili-dependent phages. Simultaneously, all clones resistant to phage PA5oct maintain the phage DNA within the population, strongly reducing bacterial virulence in vivo. These properties can be considered as key parameters for the application of this bacterial virus in phage therapy settings.Originality-Significance StatementThe emergence of phage-resistant mutants is a key aspect of lytic phages-bacteria interaction and the main driver for the co-evolution between both organisms. However, this fundamental property also has implications for bacterial eradication in phage therapy settings. Here, we analyze the impact of PA5oct jumbo phage treatment of planktonic/cell line associated and sessile P. aeruginosa population in a preclinical evaluation of this phage for therapeutic applications. Besides its broad-spectrum activity and efficient bacteria reduction in both airway surface liquid (ASL) model, and biofilm matrix degradation, PA5oct appears to persist in most of phage-resistant clones. Indeed, a high percentage of resistance (20/30 clones) to PA5oct is accompanied by the presence of phage DNA within bacterial culture. Moreover, the maintenance of this phage in the bacterial population is correlated to reduced P. aeruginosa virulence, coupled with a sensitization to innate immune mechanisms, and a significantly reduced growth rate. We observed rather unusual consequences of PA5oct infection causing an increased inflammatory response of monocytes to P. aeruginosa. This, phenomenon combined with the loss or modification of the phage receptor makes most of the phage-resistant clones significantly less pathogenic in in vivo model. During phage therapy treatment, phage-resistance is considered as an adverse effect, but our results indicate that it leads to diminished bacterial virulence and increased clearance of the infected host. These findings provide new insights into the general knowledge of giant phages biology and the impact of their application in phage therapy.

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Pseudomonas aeruginosa MutL promotes large chromosomal deletions through non-homologous end joining to prevent bacteriophage predation

Cited by 56 publications

References 60 publications

Mutation-driven evolution of Pseudomonas aeruginosa in the presence of either ceftazidime or ceftazidime/avibactam

Mutation-driven evolution of Pseudomonas aeruginosa in the presence of either ceftazidime or ceftazidime/avibactam

Development of a Bacteriophage Cocktail to Constrain the Emergence of Phage-Resistant Pseudomonas aeruginosa

Pseudomonas aeruginosa PA5oct jumbo phage impacts planktonic and biofilm population and reduces its host virulence

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