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Brief Report

Genome-wide analyses reveal antibiotic resistance genes and mechanisms in pathogenic Pseudomonas bacteria

[version 1; peer review: 1 approved, 1 not approved]
PUBLISHED 04 Aug 2020
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This article is included in the Pathogens gateway.

This article is included in the Antimicrobial Resistance collection.

Abstract

Background: The global emergence and re-emergence of antibiotic resistance among the Pseudomonas pathogens causes great problems to patients undergoing chemotherapy. However, there is limited comparative information on the antibiotic resistance genes (ARGs) and mechanisms across the Pseudomonas pathogenic groups.
Methods: The complete genomes of five Pseudomonas pathogen groups, P. aeruginosa, P. fluorescens, P. putida, P. stutzeri and P. syringae, were analyzed for ARGs.
Results: A significant number of ARGs were identified in the P. aeruginosa genome compared to the other Pseudomonas pathogens. The opportunistic pathogens P. stutzeri and P. putida were shown to be the closest to P. aeruginosa with an average nucleotide identity (%) of 80.30 and 79.52.  The pathogen genome with the least hit was P. stutzeri. The four major antibiotic resistance mechanisms that include the efflux, inactivation, target alteration and efflux::target alteration were reported.
Conclusion: The findings of this brief report could be useful in understanding the chemotherapeutics against antibiotic resistance strains of Pseudomonas pathogens

Keywords

Pseudomonas, Antibiotics Resistance, bacterial pathogen, P. aeruginosa

Introduction

The emergence of Gram-negative bacterial antibiotics resistance is a growing threat to antibiotic therapy. The bacterial pathogens in the genus Pseudomonas are mostly opportunistic that cause great damage and loss of life (Breidenstein et al., 2011; Evans et al., 2008; Kawai, 1974; Lalucat et al., 2006). These Pseudomonas pathogens are pervasive that is able to infect, survive and proliferate in a wide range of biotic and abiotic environments (Azam & Khan, 2019; Silby et al., 2011). Presently, there are seven groups in the genus Pseudomonas, with P. aeruginosa being the most pathogenic (Barbier et al., 2013) and which causes high morbidity and mortality in cystic fibrosis patients and immunocompromised individuals (Sadikot et al., 2005). The other Pseudomonas pathogenic groups include P. fluorescens (Biaggini et al., 2015), P. putida (Fernández et al., 2015), P. stutzeri (Lalucat et al., 2006) and P. syringae (Xin et al., 2018). P. aeruginosa is the major cause of infections in developed countries due to its highly evolved resistance to a wide variety of antibiotics (Hancock & Speert, 2000) making it very difficult to treat and limiting therapeutics (Breidenstein et al., 2011). Even though P. aeruginosa is the most studied, there are some other Pseudomonas species that exhibits opportunistic pathogenic behavior to animals (P. fluorescens, P. putida and P. stutzeri) (Azam & Khan, 2019) and plants (P. syringae) (Xin et al., 2018). Most of the known antimicrobial resistance (AMR) gene and mechanistic studies are focused on P. aeruginosa with little attention directed to other Pseudomonas pathogens. Hence, there is need to investigate different Pseudomonas pathogens genomes for diverse antibiotic resistance genes (ARGs) and mechanism. Therefore, this brief report concisely revealed the ARGs, mechanisms and drugs in P. aeruginosa in comparison to other Pseudomonas pathogens.

Methods

The complete genomes of the five groups of Pseudomonas pathogens, which included the P. aeruginosa (NC_002516.2), P. fluorescens (NC_016830.1), P. putida (NC_002947.4), P. stutzeri (NC_015740.1) and P. syringae (NC_007005.1) fasta file sequences, were downloaded from The National Center for Biotechnology Information Genome database. The five Pseudomonas pathogen genomes were selected to represent the Pseudomonas groups. The fasta file format of the genome sequence of bacteria were thoroughly analyzed for ARGs on the bulk analysis Resistance Gene Identifier (RGI) 5.1.0, CARD 3.0.9 Platform (Alcock et al., 2020) to extract AMR Genes, AMR Gene Family, Drug Class and Resistance Mechanism data. Default select criteria, which identified gene based on strict or perfect only was used. On the RGI platform, each genome sequence file was uploaded and all settings were left at default. The resistance genes, mechanism and drugs obtained from RGI platform were further analyzed using Prism 8 for number of ARG hits, gene family, mechanism and drug class per Pseudomonas species.

Results

The five Pseudomonas pathogens show significant genome similarity (Table 1). P. stutzeri and P. putida, which have been reported to be opportunist pathogens to humans, were shown to be the closest relations to P. aeruginosa, with an average nucleotide identity (%) of 80.30 and 79.52. P. syringae, which infects plants, had the lowest average nucleotide identity (%) of 78.67. A number of antimicrobial resistance hits and genes were identified across the five Pseudomonas pathogens, but the highest hits were seen in the P. aeruginosa genome. The pathogen genome with the least hit was P. stutzeri (Figure 1A, B). The four major antibiotic resistance mechanisms identified were the efflux, inactivation, target alteration and efflux::target alterations in the P. aeruginosa genome. The efflux, inactivation and efflux::target alterations were identified in the P. fluorescens genome, while only the efflux and inactivation alterations were identified in the P. putida, P. stutzeri and P. syringae genomes (Figure 2A). The number of drug classes that P. aeruginosa was shown to be resistant to is also shown in Figure 2B.

Table 1. Pseudomonas pathogen genomes used in this study.

S/NPseudomonas
groups
PathogenicGenome accession No.Average nucleotide
identity (%)
1P. aeruginosaPathogenic to plants and
animals (Azam & Khan, 2019)
NC_002516.2RG
2P. fluorescensopportunistic human pathogens
(Biaggini et al., 2015)
NC_016830.179.25
3P. putidaopportunistic human pathogens
(Fernández et al., 2015)
NC_002947.479.52
4P. stutzeriopportunistic human pathogens
(Lalucat et al., 2006)
NC_015740.180.30
5P. syringaePathogenic to plants (Xin et al., 2018)NC_007005.178.67
41399484-e76c-4c59-8dfd-9f4f54e58ccd_figure1.gif

Figure 1. P. aeruginosa exhibited more Antibiotics Resistance Genes (ARGs).

(A) The number of hit per each Pseudomonas pathogen genome (B). Number of ARGs per each Pseudomonas pathogen genome.

41399484-e76c-4c59-8dfd-9f4f54e58ccd_figure2.gif

Figure 2. Antibiotic efflux resistance mechanism is widely spread across the Pseudomonas pathogen genome and highly abundant in P. aeruginosa.

(A) Number of resistance mechanism per each Pseudomonas pathogen genome. (B) P. aeruginosa resistance drug class. Drug class key: A= aminoglycoside antibiotic; B= fluoroquinolone antibiotic, diaminopyrimidine antibiotic, phenicol antibiotic; C= macrolide antibiotic, carbapenem, tetracycline antibiotic, acridine dye, diaminopyrimidine antibiotic, phenicol antibiotic; D= macrolide antibiotic, fluoroquinolone antibiotic, aminoglycoside antibiotic, carbapenem, cephalosporin, cephamycin, penam, tetracycline antibiotic, acridine dye, phenicol antibiotic; E= macrolide antibiotic, fluoroquinolone antibiotic, aminoglycoside antibiotic, cephalosporin, penam, tetracycline antibiotic, aminocoumarin antibiotic, diaminopyrimidine antibiotic, phenicol antibiotic; F= macrolide antibiotic, fluoroquinolone antibiotic, cephalosporin, penam, tetracycline antibiotic, aminocoumarin antibiotic, diaminopyrimidine antibiotic, phenicol antibiotic; G= macrolide antibiotic, fluoroquinolone antibiotic, monobactam, carbapenem, cephalosporin, cephamycin, penam, tetracycline antibiotic, peptide antibiotic, aminocoumarin antibiotic, diaminopyrimidine antibiotic, sulfonamide antibiotic, phenicol antibiotic, penem; H= peptide antibiotic; I = phenicol antibiotic; J = sulfonamide antibiotic.

Discussion

There is an increasing interest and focus in the antibiotic resistance in pathogenic Pseudomonas (Blair et al., 2014; Blanco et al., 2016; Li et al., 2015; Soto, 2013; Webber & Piddock, 2003). Hence, this report took advantage of the available Pseudomonas pathogens genomes and analysed for antibiotic resistance genes and mechanisms against different available drugs.

The significant number of ARGs and mechanisms were identified in the genome of P. aeruginosa, which is more virulent and well-studied compared to other species. P. aeruginosa also infects a wide range of plants and animals, including humans (Azam & Khan, 2019; Hancock & Speert, 2000; Sadikot et al., 2005). P. aeruginosa is of great medical importance due to its exhibition of multidrug resistance and its association with serious illnesses (Breidenstein et al., 2011; Evans et al., 2008; Gonzalez et al., 2019).

The most common resistance mechanism of Pseudomonas pathogens is the antibiotic efflux pump mechanism. Although this mechanism is most often seen in P. aeruginosa, it is also found in other Pseudomonas pathogen genomes. It has long been known that the antibiotic efflux pump is a key mechanism of resistance in Gram-negative bacterial pathogens (Blair et al., 2014; Blanco et al., 2016; Soto, 2013; Webber & Piddock, 2003). An antibiotic resistance strain’s efflux pumps allow it to regulate itself by excluding toxic substances, including antimicrobial drugs (Blair et al., 2014; Li et al., 2015; Soto, 2013; Webber & Piddock, 2003).

Conclusion

The different ARGs and mechanisms against known drugs in P. aeruginosa in comparison to other Pseudomonas pathogen were investigated and concisely reported in this brief report. The findings in this report could be useful in understanding the use of chemotherapeutics against antibiotic-resistant strains of Pseudomonas pathogens.

Data availability

Source data

Table 1 lists the NCBI Genome accession numbers used in this study.

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Benson O. Genome-wide analyses reveal antibiotic resistance genes and mechanisms in pathogenic Pseudomonas bacteria [version 1; peer review: 1 approved, 1 not approved]. F1000Research 2020, 9:903 (https://doi.org/10.12688/f1000research.25391.1)
NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article.
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ApprovedThe paper is scientifically sound in its current form and only minor, if any, improvements are suggested
Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.
Not approvedFundamental flaws in the paper seriously undermine the findings and conclusions
Version 1
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PUBLISHED 04 Aug 2020
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Reviewer Report 16 Nov 2020
Paula Blanco, Molecular Basis of Adaptation Laboratory, Departamento de Sanidad Animal, Facultad de Veterinaria, Universidad Complutense de Madrid, Madrid, Spain 
Not Approved
VIEWS 18
This brief report deals with the comparison of five Pseudomonas species' genomes in order to get information about the different antibiotic resistance genes and mechanisms. However, the findings described here lack novelty and interest, since the mechanisms of resistance and ... Continue reading
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CITE
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Blanco P. Reviewer Report For: Genome-wide analyses reveal antibiotic resistance genes and mechanisms in pathogenic Pseudomonas bacteria [version 1; peer review: 1 approved, 1 not approved]. F1000Research 2020, 9:903 (https://doi.org/10.5256/f1000research.28015.r74062)
NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article.
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Reviewer Report 19 Aug 2020
Asad U. Khan, Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, Uttar Pradesh, India 
Approved
VIEWS 18
Short communication focuses on antibiotic resistance caused by efflux pump overexpression and the use of cryo electron microscopy for protein protein or protein ligand interaction. It also describes the limitations of x ray crystallography. The author also extends the cryo ... Continue reading
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CITE
HOW TO CITE THIS REPORT
Khan AU. Reviewer Report For: Genome-wide analyses reveal antibiotic resistance genes and mechanisms in pathogenic Pseudomonas bacteria [version 1; peer review: 1 approved, 1 not approved]. F1000Research 2020, 9:903 (https://doi.org/10.5256/f1000research.28015.r68814)
NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article.

Comments on this article Comments (0)

Version 1
VERSION 1 PUBLISHED 04 Aug 2020
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Alongside their report, reviewers assign a status to the article:
Approved - the paper is scientifically sound in its current form and only minor, if any, improvements are suggested
Approved with reservations - A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.
Not approved - fundamental flaws in the paper seriously undermine the findings and conclusions
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