The Acquisition of Resistance to Carbapenem and Macrolide-mediated Quorum Sensing Inhibition byPseudomonas aeruginosavia a Novel Integrative and Conjugative Element ICETn43716385

AbstractPseudomonas aeruginosacan cause persistant and life-threatening infections in immunocompromised patients. Carbapenems are the first-line agents to treatP. aeruginosainfections; therefore, the emergence of carbapenem-resistantP. aeruginosastrains has greatly challenged effective antibiotic therapy. In this study, we characterised the full-length genomes of two carbapenem resistantP. aeruginosaclinical isolates that produce the carbapebemase New Delhi metallo-β-lactamase-1 (NDM-1). We found that theblaNDM-1gene is encoded by a novel intergrative and conjugative element (ICE) ICETn43716385, which also carries the macrolide resistance genemsr(E)and the florfenicol resistance genefloR. Themsr(E)gene has rarely been described inP. aeruginosagenomes. To investigate the functional roles ofmsr(E)inP. aeruginosa, we exogeneously expressed this gene inP. aeruginosalaboratory strains and found that the acquisition ofmsr(E)could abolish the azithromycin-mediated quorum sensing inhibitionin vitroand the anti-Pseudomonas effect of azithromycinin vivo. In addition, the expression ofmsr(E)almost completely restored the azithromycin-affectedP. aeruginosatranscriptome, as shown by our RNA sequencing analysis. We present the first evidence ofblaNDM-1to be carried by intergrative and conjugative elements, and the first evidence of co-transfer of carbapenem resistance and the resistance to macrolide-mediated quorum sensing inhibition intoP. aeruginosagenomes.ImportanceCarbapenem resistantP. aeruginosahas recently been listed as the top three most dangerous superbugs by World Health Organisation. The transmission ofblaNDM-1gene intoP. aeruginosacan cause extreme resistance to carbapenems and fourth generation cephalosporins, which greatly compromises the effectiveness of these antibiotics against Pseudomonas infections. However, the lack of complete genome sequence of NDM-1-producingP. aeruginosahas limited our understanding of the transmisibility ofblaNDM-1in this organism. Here we showed the co-transfer ofblaNDM-1andmsr(E)intoP. aeruginosagenome by a novel integrative and conjugative element (ICE). The acquisition of these two genes confersP. aeruginosawith resistance to carbapenem and macrolide-mediated quorum sensing inhibition, both of which are important treatment stretagies forP. aeruginosainfections. Our findings highlight the potential of ICEs in transmitting carbapenem resistance, and that the anti-virulence treatment ofP. aeruginosainfections by macrolides can be challenged by horizontal gene transfer.

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Integration of the blaNDM-1 carbapenemase gene into a novel SXT/R391 integrative and conjugative element in Proteus vulgaris

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkaa068 ◽

2020 ◽

Vol 75 (6) ◽

pp. 1439-1442

Author(s):

Ling-Han Kong ◽

Rong Xiang ◽

Yu-Long Wang ◽

Shun-Kang Wu ◽

Chang-Wei Lei ◽

...

Keyword(s):

Resistance Gene ◽

Resistance Genes ◽

Bioinformatics Analysis ◽

Carbapenem Resistance ◽

Resistance Determinant ◽

Proteus Vulgaris ◽

Genetic Environment ◽

Carbapenem Resistant ◽

Carbapenemase Gene ◽

Integrative And Conjugative Element

Abstract Objectives To characterize the genetic environment of the carbapenem resistance determinant in Proteus vulgaris of swine origin. Methods The carbapenem-resistant P. vulgaris strain BC22 was isolated from a faecal swab from a diseased pig with diarrhoea in Sichuan Province of China in 2018. The presence of carbapenemase genes was screened by PCR. WGS and bioinformatics analysis were performed to analyse the genetic environment of the carbapenem resistance determinant. Results P. vulgaris strain BC22 was found to harbour the carbapenemase gene blaNDM-1. WGS data revealed that blaNDM-1 was located in a truncated ISAba125 composite transposon. The carbapenem resistance gene blaNDM-1 and 20 other resistance genes, including the multiresistance gene cfr and the bifunctional aminoglycoside/quinolone resistance gene aac(6′)-lb-cr, were located in a novel SXT/R391 integrative and conjugative element (ICE). This new SXT/R391 ICE of 148.7 kb was chromosomally located, and could be transferred to Escherichia coli. Conclusions Here, we report a carbapenemase gene, blaNDM-1, integrated into an SXT/R391 ICE. Our study highlights that this SXT/R391 ICE may facilitate the dissemination of clinically important resistance genes such as blaNDM-1, cfr and aac(6′)-lb-cr.

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OmpK26, a Novel Porin Associated with Carbapenem Resistance in Klebsiella pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00309-11 ◽

2011 ◽

Vol 55 (10) ◽

pp. 4742-4747 ◽

Cited By ~ 29

Author(s):

Laura García-Sureda ◽

Antonio Doménech-Sánchez ◽

Mariette Barbier ◽

Carlos Juan ◽

Joan Gascó ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Outer Membrane Proteins ◽

Systemic Infection ◽

Carbapenem Resistance ◽

Resistant Isolate ◽

Identical Result ◽

Sequencing Analysis ◽

Content Type ◽

Carbapenem Resistant

ABSTRACTClinical isolates ofKlebsiella pneumoniaeresistant to carbapenems are being isolated with increasing frequency. Loss of the expression of the major nonspecific porins OmpK35/36 is a frequent feature in these isolates. In this study, we looked for porins that could compensate for the loss of the major porins in carbapenem-resistant organisms. Comparison of the outer membrane proteins from twoK. pneumoniaeclinical isogenic isolates that are susceptible (KpCS-1) and resistant (KpCR-1) to carbapenems revealed the absence of OmpK35/36 and the presence of a new 26-kDa protein in the resistant isolate. An identical result was obtained when another pair of isogenic isolates that are homoresistant (Kpn-3) and heteroresistant (Kpn-17) to carbapenems were compared. Mass spectrometry and DNA sequencing analysis demonstrated that this new protein, designated OmpK26, is a small monomeric oligogalacturonate-specific porin that belongs to the KdgM family of porins. Insertion-duplication mutagenesis of the OmpK26 coding gene,yjhA, in the carbapenem-resistant, porin-deficient isolate KpCR-1 caused the expression of OmpK36 and the reversion to the carbapenem-susceptible phenotype, suggesting that OmpK26 is indispensable for KpCR-1 to lose OmpK36 and become resistant to these antibiotics. Moreover, loss of the major porin and expression of OmpK26 reducedin vitrofitness and attenuated virulence in a murine model of acute systemic infection. Altogether, these results indicate that expression of the oligogalacturonate-specific porin OmpK26 compensates for the absence of OmpK35/36 and allows carbapenem resistance inK. pneumoniaebut cannot restore the fitness of the microorganism.

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1348. In vitro Activity of Plazomicin, a Next-Generation Aminoglycoside, Against Carbapenemase-Producing Klebsiella pneumoniae

Open Forum Infectious Diseases ◽

10.1093/ofid/ofy210.1179 ◽

2018 ◽

Vol 5 (suppl_1) ◽

pp. S412-S413

Author(s):

Michael R Jacobs ◽

Caryn E Good ◽

Ayman M Abdelhamed ◽

Daniel D Rhoads ◽

Kristine M Hujer ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Carbapenem Resistance ◽

Vitro Activity ◽

In Vitro Activity ◽

Next Generation ◽

Aminoglycoside Resistance ◽

Carbapenem Resistant ◽

Carbapenemase Gene ◽

Research Grant

Abstract Background Plazomicin is a next-generation aminoglycoside with in vitro activity against multidrug-resistant Gram-negative species, including carbapenem-resistant isolates. The Consortium on Resistance Against Carbapenems in Klebsiella and other Enterobacteriaceae (CRACKLE) is a federally funded, prospective multicenter consortium of 20 hospitals from nine US healthcare systems to track carbapenem-resistant Enterobacteriaceae. Methods Minimum inhibitory concentrations (MICs) of plazomicin were determined by broth microdilution according to current CLSI guidelines against a collection of 697 carbapenem-resistant Klebsiella pneumoniae with defined carbapenem resistance mechanisms, including KPC and OXA carbapenemases. Isolates were submitted by participating CRACKLE centers. Results Carbapenemases present in study isolates included KPC-2 (n = 323), KPC-3 (n = 364), KPC-4 (n = 2), OXA-48 like (n = 7), and NDM (n = 1). Plazomicin MICs ranged from ≤0.12 to >32 mg/L, with MIC50 and MIC90 values of 0.25 and 1 mg/L, respectively (figure). MICs of 689 (98.8%) isolates were ≤4 mg/L, while MICs of the remaining eight isolates were >32 mg/L. Plazomicin MICs were related to specific carbapenemases present in isolates: of eight isolates with MICs >32 mg/L, seven contained OXA-48 like and one contained KPC-3, suggesting that these isolates possess an aminoglycoside-resistance mechanism on the same plasmid as their carbapenemase gene, such as a 16S ribosomal RNA methyltransferase, against which plazomicin is not active. Conclusion Plazomicin has good in vitro potency against a collection of carbapenemase-producing K. pneumoniae, with MIC90 value of 1 mg/L and MICs of ≤4 mg/L for 98.9% of isolates. Disclosures M. R. Jacobs, Achaogen: Investigator, Research grant. Shionogi: Investigator, Research grant. L. Connolly, Achaogen, Inc.: Consultant, Consulting fee. K. M. Krause, Achaogen: Employee, Salary. S. S. Richter, bioMerieux: Grant Investigator, Research grant. BD Diagnostics: Grant Investigator, Research grant. Roche: Grant Investigator, Research grant. Hologic: Grant Investigator, Research grant. Diasorin: Grant Investigator, Research grant. Accelerate: Grant Investigator, Research grant. Biofire: Grant Investigator, Research grant. D. Van Duin, achaogen: Scientific Advisor, Consulting fee. shionogi: Scientific Advisor, Consulting fee. Allergan: Scientific Advisor, Consulting fee. Astellas: Scientific Advisor, Consulting fee. Neumedicine: Scientific Advisor, Consulting fee. Roche: Scientific Advisor, Consulting fee. T2 Biosystems: Scientific Advisor, Consulting fee.

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Exemplifying the next generation of antibiotic susceptibility intensifiers of phytochemicals by LasR-mediated quorum sensing inhibition

Scientific Reports ◽

10.1038/s41598-021-01845-8 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Arpit Shukla ◽

Gaurav Shukla ◽

Paritosh Parmar ◽

Baldev Patel ◽

Dweipayan Goswami ◽

...

Keyword(s):

Quorum Sensing ◽

Metabolic Pathways ◽

Single Unit ◽

Antimicrobial Properties ◽

Well Being ◽

Natural Compounds ◽

Human Pathogens ◽

Quorum Sensing Inhibition ◽

In Silico Studies

AbstractThere persists a constant threat from multidrug resistance being acquired by all human pathogens that challenges the well-being of humans. This phenomenon is predominantly led by Pseudomonas aeruginosa which is already resistant to the current generations of antibiotic by altering its metabolic pathways to survive. Specifically for this microbe the phenomenon of quorum sensing (QS) plays a crucial role in acquiring virulence and pathogenicity. QS is simply the cross talk between the bacterial community driven by signals that bind to receptors, enabling the entire bacterial microcosm to function as a single unit which has led to control P. aeruginosa cumbersome even in presence of antibiotics. Inhibition of QS can, therefore, be of a significant importance to curb such virulent and pathogenic strains of P. aeruginosa. Natural compounds are well known for their antimicrobial properties, of which, information on their mode of action is scarce. There can be many antimicrobial phytochemicals that act by hindering QS-pathways. The rationale of the current study is to identify such natural compounds that can inhibit QS in P. aeruginosa driven by LasR, PhzR, and RhlR dependent pathways. To achieve this rationale, in silico studies were first performed to identify such natural compounds which were then validated by in vitro experiments. Gingerol and Curcumin were identified as QS-antagonists (QSA) which could further suppress the production of biofilm, EPS, pyocyanin, and rhamnolipid along with improving the susceptibility to antibiotics.

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Multicity Outbreak of Carbapenem-Resistant Acinetobacter baumannii Isolates Producing the Carbapenemase OXA-40

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00116-06 ◽

2006 ◽

Vol 50 (9) ◽

pp. 2941-2945 ◽

Cited By ~ 127

Author(s):

Karen Lolans ◽

Thomas W. Rice ◽

L. Silvia Munoz-Price ◽

John P. Quinn

Keyword(s):

Acinetobacter Baumannii ◽

Carbapenem Resistance ◽

The United States ◽

Nucleotide Sequencing ◽

Sequencing Analysis ◽

Carbapenem Resistant ◽

Extended Care ◽

Care Facilities ◽

High Level ◽

First Time

ABSTRACT During 2005 we detected a multicity outbreak of infections or colonization due to high-level imipenem-resistant Acinetobacter baumannii (MIC, 64 μg/ml). One hundred isolates from diverse sources were obtained from seven acute-care hospitals and two extended-care facilities; 97% of the isolates belonged to one clone. Susceptibility testing of the first 42 isolates (January to April 2005) revealed broad resistance profiles. Half of the isolates were susceptible to ceftazidime, with many isolates susceptible only to colistin. The level of AmpC β-lactamase expression was stronger in isolates resistant to ceftazidime. PCR and subsequent nucleotide sequencing analysis identified bla OXA-40. The presence of an OXA-40 β-lactamase in these isolates correlated with the carbapenem resistance. By Southern blot analysis, a bla OXA-40-specific probe revealed that the gene was both plasmid and chromosomally located. This is the first time in the United States that such carbapenem resistance in A. baumannii has been attributable to a carbapenemase.

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The carbapenem resistance gene blaOXA-23 is disseminated by a conjugative plasmid containing the novel transposon Tn6681 in Acinetobacter johnsonii M19

Antimicrobial Resistance and Infection Control ◽

10.1186/s13756-020-00832-4 ◽

2020 ◽

Vol 9 (1) ◽

Author(s):

Gongli Zong ◽

Chuanqing Zhong ◽

Jiafang Fu ◽

Yu Zhang ◽

Peipei Zhang ◽

...

Keyword(s):

Molecular Docking ◽

Resistance Gene ◽

Carbapenem Resistance ◽

Conjugative Plasmid ◽

The Novel ◽

Illumina Hiseq ◽

Discovery Studio ◽

Acinetobacter Johnsonii ◽

Type Iv ◽

Carbapenem Resistant

Abstract Background Carbapenem resistant Acinetobacter species have caused great difficulties in clinical therapy in the worldwide. Here we describe an Acinetobacter johnsonii M19 with a novel blaOXA-23 containing transposon Tn6681 on the conjugative plasmid pFM-M19 and the ability to transferand carbapenem resistance. Methods A. johnsonii M19 was isolated under selection with 8 mg/L meropenem from hospital sewage, and the minimum inhibitory concentrations (MICs) for the representative carbapenems imipenem, meropenem and ertapenem were determined. The genome of A. johnsonii M19 was sequenced by PacBio RS II and Illumina HiSeq 4000 platforms. A homologous model of OXA-23 was generated, and molecular docking models with imipenem, meropenem and ertapenem were constructed by Discovery Studio 2.0. Type IV secretion system and conjugation elements were identified by the Pathosystems Resource Integration Center (PATRIC) server and the oriTfinder. Mating experiments were performed to evaluate transfer of OXA-23 to Escherichia coli 25DN. Results MICs of A. johnsonii M19 for imipenem, meropenem and ertapenem were 128 mg/L, 48 mg/L and 24 mg/L, respectively. Genome sequencing identified plasmid pFM-M19, which harbours the carbapenem resistance gene blaOXA-23 within the novel transposon Tn6681. Molecular docking analysis indicated that the elongated hydrophobic tunnel of OXA-23 provides a hydrophobic environment and that Lys-216, Thr-217, Met-221 and Arg-259 were the conserved amino acids bound to imipenem, meropenem and ertapenem. Furthermore, pFM-M19 could transfer blaOXA-23 to E. coli 25DN by conjugation, resulting in carbapenem-resistant transconjugants. Conclusions Our investigation showed that A. johnsonii M19 is a source and disseminator of blaOXA-23 and carbapenem resistance. The ability to transfer blaOXA-23 to other species by the conjugative plasmid pFM-M19 raises the risk of spread of carbapenem resistance. Graphic abstract The carbapenem resistance gene blaOXA-23 is disseminated by a conjugative plasmid containing the novel transposon Tn6681 in Acinetobacter johnsonii M19.

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Tridecaptin M, a New Variant Discovered in Mud Bacterium, Shows Activity against Colistin- and Extremely Drug-ResistantEnterobacteriaceae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00338-19 ◽

2019 ◽

Vol 63 (6) ◽

Cited By ~ 8

Author(s):

Manoj Jangra ◽

Manpreet Kaur ◽

Rushikesh Tambat ◽

Rohit Rana ◽

Sushil K. Maurya ◽

...

Keyword(s):

Hemolytic Activity ◽

World Health ◽

Fourth Generation ◽

Infection Model ◽

Drug Resistant ◽

Content Type ◽

Extensively Drug Resistant ◽

New Variant

ABSTRACTThe World Health Organization has categorized the Gram-negative superbugs, which are inherently impervious to many antibiotics, as critical priority pathogens due to the lack of effective treatments. The breach in our last-resort antibiotic (i.e., colistin) by extensively drug-resistant and pan-drug-resistantEnterobacteriaceaestrains demands the immediate development of new therapies. In the present study, we report the discovery of tridecaptin M, a new addition to the family, and its potential against colistin-resistantEnterobacteriaceae in vitroandin vivo. Also, we performed mode-of-action studies using various fluorescent probes and studied the hemolytic activity and mammalian cytotoxicity in two cell lines. Tridecaptin M displayed strong antibacterial activity (MICs of 2 to 8 μg ml−1) against clinical strains ofKlebsiella pneumoniae(which were resistant to colistin, carbapenems, third- and fourth-generation cephalosporins, fluoroquinolones, fosfomycin, and other antibiotics) andmcr-1-positiveEscherichia colistrains. Unlike polymyxins, tridecaptin M did not permeabilize the outer membrane or cytoplasmic membrane. It blocked ATP synthesis in bacteria by dissipating the proton motive force. The compound exhibited negligible acquired resistance, lowin vitrocytotoxicity and hemolytic activity, and no significant acute toxicity in mice. It also showed promising efficacy in a thigh infection model of colistin-resistantK. pneumoniae. Altogether, these results demonstrate the future prospects of this class of antibiotics to address the unmet medical need to circumvent colistin resistance in extensively drug-resistantEnterobacteriaceaeinfections. The work also emphasizes the importance of natural products in our shrunken drug discovery pipeline.

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The antimicrobial activity of silver acetate against Acinetobacter baumannii in a Galleria mellonella infection model

PeerJ ◽

10.7717/peerj.11196 ◽

2021 ◽

Vol 9 ◽

pp. e11196

Author(s):

Eden Mannix-Fisher ◽

Samantha McLean

Keyword(s):

Acinetobacter Baumannii ◽

Bacterial Infections ◽

Galleria Mellonella ◽

World Health Organisation ◽

World Health ◽

Infection Model ◽

Silver Acetate ◽

Selective Toxicity ◽

Carbapenem Resistant

Background The increasing prevalence of bacterial infections that are resistant to antibiotic treatment has caused the scientific and medical communities to look for alternate remedies aimed at prevention and treatment. In addition to researching novel antimicrobials, there has also been much interest in revisiting some of the earliest therapies used by man. One such antimicrobial is silver; its use stretches back to the ancient Greeks but interest in its medicinal properties has increased in recent years due to the rise in antibiotic resistance. Currently antimicrobial silver is found in everything from lunch boxes to medical device implants. Though much is claimed about the antimicrobial efficacy of silver salts the research in this area is mixed. Methods Herein we investigated the efficacy of silver acetate against a carbapenem resistant strain of Acinetobacter baumannii to determine the in vitro activity of this silver salt against a World Health Organisation designated category I critical pathogen. Furthermore, we use the Galleria mellonella larvae model to assess toxicity of the compound and its efficacy in treating infections in a live host. Results We found that silver acetate can be delivered safely to Galleria at medically relevant and antimicrobial levels without detriment to the larvae and that administration of silver acetate to an infection model significantly improved survival. This demonstrates the selective toxicity of silver acetate for bacterial pathogens but also highlights the need for administration of well-defined doses of the antimicrobial to provide an efficacious treatment.

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Comparative analysis of Clinial Carbapenem- Resistant (CR), Hypermucoviscous (HM) and CR-HM Klebsiella pneumoniae isolates in China

10.21203/rs.3.rs-17188/v1 ◽

2020 ◽

Author(s):

Jun-Ying Zhu ◽

Guang-Yu Wang ◽

Qing Wei ◽

Zhen Shen ◽

Qiong Li ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Virulence Genes ◽

Carbapenem Resistance ◽

Resistance Rate ◽

Molecular Characteristics ◽

Carbapenem Resistant ◽

Recent Emergence ◽

String Test

Abstract Background: Although carbapenem-resistant Klebsiella pneumoniae (CRKP) and hypermucoviscous K. pneumoniae (HMKP) were largely non-overlapping, the recent emergence of CR-HMKP has raised great alarm in the world. We compared the molecular characteristics of CRKP, HMKP and CR-HMKP isolates.Results: 220 cases of K. pneumoniae isolates was collected and identified between Jan 2015 and Dec 2016 from Renji Hospital. Carbapenem resistance test and string test were performed to screen CRKP, HMKP and CR-HMKP isolates. All the CRKP, HMKP and CR-HMKP isolates were investigated for capsular genotyping, virulence genes and resistance genes by PCR and DNA sequencing. Multilocus sequence typing (MLST) was used to characterize isolates sequence types (STs). Serum killing assay and mouse lethality assay were respectively performed to confirm the virulence of the isolates in vitro and in vivo. Of 220 K. pneumoniae，71 HMKP, 84 CRKP and 8 CR-HMKP were identified. Resistance rate to carbapenems was significantly higher in CRKP than HMKP and CR-HMKP. For MLST and serotyping, ST23 (26.8%)，K1 (33.8%) and K2 (23.9%) serotypes were the most common in HMKP isolates while ST11 (84.5%, 100%) and K-nontypable (91.6%, 100%) were the predominant types in CRKP and CR-HMKP isolates. The existence of virulence genes rmpA, magA and iutA was significantly higher in HMKP while the prevalence of resistance gene blaKPC-2 was higher in CRKP and CR-HMKP. Virulence test in vivo and in vitro both showed the lower virulence of CRKP and CR-HMKP compared to HMKP.Conclusions: In spite of low virulence, the emergence of CR-HMKP indicates a confluence of hypermucoviscous phenotype and carbapenem resistance. Furthermore, the similar molecular characteristics between CRKP and CR-HMKP suggested that CR-HMKP might evolve from CRKP.

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