Scarless Removal of Large Resistance Island AbaR Results in Antibiotic Susceptibility and Increased Natural Transformability in Acinetobacter baumannii

ABSTRACT With a great diversity in gene composition, including multiple putative antibiotic resistance genes, AbaR islands are potential contributors to multidrug resistance in Acinetobacter baumannii. However, the effective contribution of AbaR to antibiotic resistance and bacterial physiology remains elusive. To address this, we sought to accurately remove AbaR islands and restore the integrity of their insertion site. To this end, we devised a versatile scarless genome editing strategy. We performed this genetic modification in two recent A. baumannii clinical strains: the strain AB5075 and the nosocomial strain AYE, which carry AbaR11 and AbaR1 islands of 19.7 kbp and 86.2 kbp, respectively. Antibiotic susceptibilities were then compared between the parental strains and their AbaR-cured derivatives. As anticipated by the predicted function of the open reading frame (ORF) of this island, the antibiotic resistance profiles were identical between the wild type and the AbaR11-cured AB5075 strains. In contrast, AbaR1 carries 25 ORFs, with predicted resistance to several classes of antibiotics, and the AYE AbaR1-cured derivative showed restored susceptibility to multiple classes of antibiotics. Moreover, curing of AbaRs restored high levels of natural transformability. Indeed, most AbaR islands are inserted into the comM gene involved in natural transformation. Our data indicate that AbaR insertion effectively inactivates comM and that the restored comM is functional. Curing of AbaR consistently resulted in highly transformable and therefore easily genetically tractable strains. Emendation of AbaR provides insight into the functional consequences of AbaR acquisition.

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Scarless removal of large resistance island AbaR results in antibiotic susceptibility and increased natural transformability in Acinetobacter baumannii

10.1101/2020.04.29.067926 ◽

2020 ◽

Author(s):

Anne-Sophie Godeux ◽

Elin Svedholm ◽

Agnese Lupo ◽

Marisa Haenni ◽

Maria-Halima Laaberki ◽

...

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Insertion Site ◽

Antibiotic Resistance Genes ◽

Multi Drug Resistance ◽

Bacterial Physiology ◽

Functional Consequences ◽

Resistance Island ◽

Insight Into ◽

Effective Contribution

ABSTRACTWith a great diversity in gene composition including multiple putative antibiotic-resistance genes, AbaR islands are potential contributors to multi-drug resistance in Acinetobacter baumannii. However, the effective contribution of AbaR to antibiotic resistance and bacterial physiology remains elusive. To address this, we sought to accurately remove AbaR islands and restore the integrity of their insertion site. To this end, we devised a versatile scarless genome editing strategy. We performed this genetic modification in two recent A. baumannii clinical strains: the strain AB5075 and the nosocomial strain AYE which carry AbaR11 and AbaR1 islands of 19.7 kbp and 86.2 kbp, respectively. Antibiotic susceptibilities were then compared between the parental strains and their AbaR-cured derivatives. As anticipated by the predicted function of the ORF of this island, the antibiotic resistance profiles were identical between the wild type and the AbaR11-cured AB5075 strains. In contrast, AbaR1 carries 25 ORFs with a predicted resistance to several classes of antibiotics and the AYE AbaR1-cured derivative showed restored susceptibility to multiple classes of antibiotics. Moreover, curing of AbaRs restored high levels of natural transformability. Indeed, most AbaR islands are inserted into the comM gene involved in natural transformation. Our data indicate that AbaR insertion effectively inactivates comM and that the restored comM is functional. Curing of AbaR consistently resulted in highly transformable, and therefore, easily genetically tractable strains. Emendation of AbaR provides insight into the functional consequences of AbaR acquisition.

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Genomic Analysis of the Multidrug-Resistant Acinetobacter baumannii Strain MDR-ZJ06 Widely Spread in China

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01134-10 ◽

2011 ◽

Vol 55 (10) ◽

pp. 4506-4512 ◽

Cited By ~ 77

Author(s):

Hua Zhou ◽

Tongwu Zhang ◽

Dongliang Yu ◽

Borui Pi ◽

Qing Yang ◽

...

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Genomic Analysis ◽

Multidrug Resistant ◽

Whole Genome Sequence ◽

Whole Genome ◽

Content Type ◽

Resistance Island

ABSTRACTWe previously reported that the multidrug-resistant (MDR)Acinetobacter baumanniistrain MDR-ZJ06, belonging to European clone II, was widely spread in China. In this study, we report the whole-genome sequence of this clinically important strain. A 38.6-kb AbaR-type genomic resistance island (AbaR22) was identified in MDR-ZJ06. AbaR22 has a structure similar to those of the resistance islands found inA. baumanniistrains AYE and AB0057, but it contained only a few antibiotic resistance genes. The region of resistant gene accumulation as previously described was not found in AbaR22. In the chromosome of the strain MDR-ZJ06, we identified the geneblaoxa-23in a composite transposon (Tn2009). Tn2009shared the backbone with otherA. baumanniitransponsons that harborblaoxa-23, but it was bracketed by two ISAba1elements which were transcribed in the same orientation. MDR-ZJ06 also expressed thearmAgene on its plasmid pZJ06, and this gene has the same genetic environment as thearmAgene of theEnterobacteriaceae. These results suggest variability of resistance acquisition even in closely relatedA. baumanniistrains.

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Acquisition of a genomic resistance island (AbGRI5) from global clone 2 through homologous recombination in a clinical Acinetobacter baumannii isolate

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkaa389 ◽

2020 ◽

Vol 76 (1) ◽

pp. 65-69

Author(s):

Xiaoting Hua ◽

Robert A Moran ◽

Qingye Xu ◽

Jintao He ◽

Youhong Fang ◽

...

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Whole Genome Sequence ◽

Oxford Nanopore ◽

History Of ◽

Agar Dilution ◽

Resistance Island ◽

New Location

Abstract Objectives To reconstruct the evolutionary history of the clinical Acinetobacter baumannii XH1056, which lacks the Oxford scheme allele gdhB. Methods Susceptibility testing was performed using broth microdilution and agar dilution. The whole-genome sequence of XH1056 was determined using the Illumina and Oxford Nanopore platforms. MLST was performed using the Pasteur scheme and the Oxford scheme. Antibiotic resistance genes were identified using ABRicate. Results XH1056 was resistant to all antibiotics tested, apart from colistin, tigecycline and eravacycline. MLST using the Pasteur scheme assigned XH1056 to ST256. However, XH1056 could not be typed with the Oxford MLST scheme as gdhB is not present. Comparative analyses revealed that XH1056 contains a 52 933 bp region acquired from a global clone 2 (GC2) isolate, but is otherwise closely related to the ST23 A. baumannii XH858. The acquired region in XH1056 also contains a 34 932 bp resistance island that resembles AbGRI3 and contains the armA, msrE-mphE, sul1, blaPER-1, aadA1, cmlA1, aadA2, blaCARB-2 and ere(B) resistance genes. Comparison of the XH1056 chromosome to that of GC2 isolate XH859 revealed that the island in XH1056 is in the same chromosomal region as that in XH859. As this island is not in the standard AbGRI3 position, it was named AbGRI5. Conclusions XH1056 is a hybrid isolate generated by the acquisition of a chromosomal segment from a GC2 isolate that contains a resistance island in a new location—AbGRI5. As well as generating ST256, it appears likely that a single recombination event is also responsible for the acquisition of AbGRI5 and its associated antibiotic resistance genes.

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Novel Aminoglycoside Resistance Transposons and Transposon-Derived Circular Forms Detected in Carbapenem-Resistant Acinetobacter baumannii Clinical Isolates

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02143-15 ◽

2016 ◽

Vol 60 (3) ◽

pp. 1801-1818 ◽

Cited By ~ 29

Author(s):

Nabil Karah ◽

Chinmay Kumar Dwibedi ◽

Karin Sjöström ◽

Petra Edquist ◽

Anders Johansson ◽

...

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Resistance Genes ◽

Point Mutations ◽

Antibiotic Resistance Genes ◽

Opportunistic Pathogen ◽

Clinical Isolates ◽

Aminoglycoside Resistance ◽

Content Type ◽

Carbapenem Resistant

Acinetobacter baumanniihas emerged as an important opportunistic pathogen equipped with a growing number of antibiotic resistance genes. Our study investigated the molecular epidemiology and antibiotic resistance features of 28 consecutive carbapenem-resistant clinical isolates ofA. baumanniicollected throughout Sweden in 2012 and 2013. The isolates mainly belonged to clonal complexes (CCs) with an extensive international distribution, such as CC2 (n= 16) and CC25 (n= 7). Resistance to carbapenems was related toblaOXA-23(20 isolates),blaOXA-24/40-like(6 isolates),blaOXA-467(1 isolate), and ISAba1-blaOXA-69(1 isolate). Ceftazidime resistance was associated withblaPER-7in the CC25 isolates. Two classical point mutations were responsible for resistance to quinolones in all the isolates. Isolates with high levels of resistance to aminoglycosides carried the 16S rRNA methylasearmAgene. The isolates also carried a variety of genes encoding aminoglycoside-modifying enzymes. Several novel structures involved in aminoglycoside resistance were identified, including Tn6279, ΔTn6279, Ab-ST3-aadB, and different assemblies of Tn6020and TnaphA6. Importantly, a number of circular forms related to the IS26or ISAba125composite transposons were detected. The frequent occurrence of these circular forms in the populations of several isolates indicates a potential role of these circular forms in the dissemination of antibiotic resistance genes.

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Novel Variants of AbaR Resistance Islands with a Common Backbone in Acinetobacter baumannii Isolates of European Clone II

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05678-11 ◽

2012 ◽

Vol 56 (4) ◽

pp. 1969-1973 ◽

Cited By ~ 36

Author(s):

Vaida Šeputienė ◽

Justas Povilonis ◽

Edita Sužiedėlienė

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Stress Protein ◽

Antibiotic Resistance Genes ◽

Structural Similarity ◽

Chromosomal Dna ◽

Class 1 Integron ◽

Content Type ◽

Novel Variants ◽

Class 1

ABSTRACTIn this study, the genetic organization of three novel genomic antibiotic resistance islands (AbaRs) inAcinetobacter baumanniiisolates belonging to group of European clone II (EC II)comMintegrated sequences of 18-, 21-, and 23-kb resistance islands were determined. These resistance islands carry the backbone of AbaR-type transposon structures, which are composed of the transposition module coding for potential transposition proteins and other genes coding for the intact universal stress protein (uspA), sulfate permease (sul), and proteins of unknown function. The antibiotic resistance genesstrA,strB,tetB, andtetRand insertion sequence CR2 element were found to be inserted into the AbaR transposons. GenBank homology searches indicated that they are closely related to the AbaR sequences found integrated incomMin strains of EC II (A. baumanniistrains 1656-2 and TCDC-AB0715) and AbaR4 integrated in another location ofA. baumanniiAB0057 (EC I). All of the AbaRs showed structural similarity to the previously described AbaR4 island and share a 12,008-bp backbone. AbaRs contain Tn1213, Tn2006, and the multiple fragments which could be derived from transposons Tn3, Tn10, Tn21, Tn1000, Tn5393, and Tn6020, the insertion sequences IS26, ISAba1, ISAba14, and ISCR2, and the class 1 integron. Moreover, chromosomal DNA was inserted into distinct regions of the AbaR backbone. Sequence analysis suggested that the AbaR-type transposons have evolved through insertions, deletions, and homologous recombination. AbaR islands, sharing the core structure similar to AbaR4, appeared to be distributed in isolates of EC I and EC II via integration into distinct genomic sites, i.e.,phoandcomM, respectively.

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IncM Plasmid R1215 Is the Source of Chromosomally Located Regions Containing Multiple Antibiotic Resistance Genes in the Globally Disseminated Acinetobacter baumannii GC1 and GC2 Clones

mSphere ◽

10.1128/msphere.00117-16 ◽

2016 ◽

Vol 1 (3) ◽

Cited By ~ 20

Author(s):

Grace A. Blackwell ◽

Mohammad Hamidian ◽

Ruth M. Hall

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Acquired Resistance ◽

Antibiotic Resistance Genes ◽

Modern Medicine ◽

Single Step ◽

Multiple Antibiotic Resistance ◽

Antibiotic Resistant ◽

Content Type ◽

Resistance Island

ABSTRACT Two lineages of extensively antibiotic-resistant A. baumannii currently plaguing modern medicine each acquired resistance to all of the original antibiotics (ampicillin, tetracycline, kanamycin, and sulfonamides) by the end of the 1970s and then became resistant to antibiotics from newer families after they were introduced in the 1980s. Here, we show that, in both of the dominant globally disseminated A. baumannii clones, a related set of antibiotic resistance genes was acquired together from the same resistance region that had already evolved in an IncM plasmid. In both cases, the action of IS26 was important in this process, but homologous recombination was also involved. The findings highlight the fact that complex regions carrying several resistance genes can evolve in one location or organism and all or part of the evolved region can then move to other locations and other organisms, conferring resistance to several antibiotics in a single step. Clear similarities between antibiotic resistance islands in the chromosomes of extensively antibiotic-resistant isolates from the two dominant, globally distributed Acinetobacter baumannii clones, GC1 and GC2, suggest a common origin. A close relative of the likely progenitor of both of these regions was found in R1215, a conjugative IncM plasmid from a Serratia marcescens strain isolated prior to 1980. The 37.8-kb resistance region in R1215 lies within the mucB gene and includes aacC1, aadA1, aphA1b, bla TEM, catA1, sul1, and tetA(A), genes that confer resistance to gentamicin, streptomycin and spectinomycin, kanamycin and neomycin, ampicillin, chloramphenicol, sulfamethoxazole, and tetracycline, respectively. The backbone of this region is derived from Tn1721 and is interrupted by a hybrid Tn2670 (Tn21)-Tn1696-type transposon, Tn6020, and an incomplete Tn1. After minor rearrangements, this R1215 resistance island can generate AbGRI2-0*, the predicted earliest form of the IS26-bounded AbGRI2-type resistance island of GC2 isolates, and to the multiple antibiotic resistance region (MARR) of AbaR0, the precursor of this region in AbaR-type resistance islands in the GC1 group. A 29.9-kb circle excised by IS26 has been inserted into the A. baumannii chromosome to generate AbGRI2-0*. To create the MARR of AbaR0, a different circular form, again generated by IS26 from an R1215 resistance region variant, has been opened at a different point by recombination with a copy of the sul1 gene already present in the AbaR precursor. Recent IncM plasmids related to R1215 have a variant resistance island containing a bla SHV gene in the same location. IMPORTANCE Two lineages of extensively antibiotic-resistant A. baumannii currently plaguing modern medicine each acquired resistance to all of the original antibiotics (ampicillin, tetracycline, kanamycin, and sulfonamides) by the end of the 1970s and then became resistant to antibiotics from newer families after they were introduced in the 1980s. Here, we show that, in both of the dominant globally disseminated A. baumannii clones, a related set of antibiotic resistance genes was acquired together from the same resistance region that had already evolved in an IncM plasmid. In both cases, the action of IS26 was important in this process, but homologous recombination was also involved. The findings highlight the fact that complex regions carrying several resistance genes can evolve in one location or organism and all or part of the evolved region can then move to other locations and other organisms, conferring resistance to several antibiotics in a single step.

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Complete Genome Sequence of Collection Strain Acinetobacter baumannii ATCC BAA-1790, Used as a Model To Study the Antibiotic Resistance Reversion Induced by Iodine-Containing Complexes

Microbiology Resource Announcements ◽

10.1128/mra.01467-19 ◽

2020 ◽

Vol 9 (3) ◽

Author(s):

Ilya S. Korotetskiy ◽

Monique Joubert ◽

Sade M. Magabotha ◽

Ardak B. Jumagaziyeva ◽

Sergey V. Shilov ◽

...

Keyword(s):

Antibiotic Resistance ◽

Acinetobacter Baumannii ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Genomic Islands ◽

Circular Chromosome ◽

Content Type ◽

Genome Methylation ◽

Long Read ◽

Collection Strain

The strain Acinetobacter baumannii ATCC BAA-1790 was sequenced as a model for nosocomial multidrug-resistant infections. Long-read PacBio sequencing revealed a circular chromosome of 3,963,235 bp with two horizontally transferred genomic islands and a 67,023-bp plasmid. Multiple antibiotic resistance genes and genome methylation patterns were identified.

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Evolution of AbGRI2-0, the Progenitor of the AbGRI2 Resistance Island in Global Clone 2 of Acinetobacter baumannii

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02662-15 ◽

2015 ◽

Vol 60 (3) ◽

pp. 1421-1429 ◽

Cited By ~ 24

Author(s):

Grace A. Blackwell ◽

Steven J. Nigro ◽

Ruth M. Hall

Keyword(s):

The Netherlands ◽

Acinetobacter Baumannii ◽

Reference Strain ◽

Draft Genome ◽

Insertion Site ◽

Mercury Resistance ◽

Common Source ◽

Antibiotic Resistant ◽

Content Type ◽

Resistance Island

A320, isolated in the Netherlands in 1982 and also known as RUH134, is the earliest available multiply antibiotic-resistant (MAR)Acinetobacter baumanniiisolate belonging to global clone 2 (GC2) and is the reference strain for this clone. The draft genome sequence of A320 was used to investigate the original location and configuration of the IS26-bounded AbGRI2 resistance island found in current GC2 isolates. PCR mapping and sequencing were used to order contigs composing the resistance islands. A320 contains two IS26-bounded resistance islands, AbGRI2-0a and AbGRI2-0b, of 7.8 kb and 25.4 kb, respectively. Together they containblaTEM,aacC1,aadA1,sul1,catA1, andaphA1bgenes, which confer resistance to antibiotics used clinically in the 1970s, as well as an incomplete mercury resistance module. Tracking the continuity of the chromosome and the target site duplications revealed that the two resistance islands were originally together as AbGRI2-0, an island of 32.4 kb, and were subsequently separated via an IS26-mediated intramolecular inversion that reversed the orientation of 1.54 Mb of the chromosome and duplicated an IS26. A320 contains an ancestral form of AbGRI2, and the original insertion site of the AbGRI2 island was identified. Many of the AbGRI2 versions present in the completed GC2 genomes can be derived from it via the variant AbGRI2-1. IS26-mediated inversions have also played a part in forming AbGRI2-0, and, upon reversal, large regions of AbGRI2-0 are identical to parts of AbaR0, the ancestral version of the AbaR islands present in GC1 isolates. This indicates a common source.

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Draft Genome Sequence of the Shrimp Pathogen Vibrio parahaemolyticus ST17.P5-S1, Isolated in Peninsular Malaysia

Microbiology Resource Announcements ◽

10.1128/mra.01053-18 ◽

2018 ◽

Vol 7 (11) ◽

Cited By ~ 2

Author(s):

Sridevi Devadas ◽

Subha Bhassu ◽

Tze Chiew Christie Soo ◽

Fatimah M. Yusoff ◽

Mohamed Shariff

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Vibrio Parahaemolyticus ◽

East Coast ◽

Draft Genome ◽

Antibiotic Resistance Genes ◽

Peninsular Malaysia ◽

Penaeus Vannamei ◽

Content Type ◽

Acute Hepatopancreatic Necrosis Disease

We sequenced the genome of Vibrio parahaemolyticus strain ST17.P5-S1, isolated from Penaeus vannamei cultured in the east coast of Peninsular Malaysia. The strain contains several antibiotic resistance genes and a plasmid encoding the Photorhabdus insect-related (Pir) toxin-like genes, pirAvp and pirBvp, associated with acute hepatopancreatic necrosis disease (AHPND).

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Environmental Spread of New Delhi Metallo-β-Lactamase-1-Producing Multidrug-Resistant Bacteria in Dhaka, Bangladesh

Applied and Environmental Microbiology ◽

10.1128/aem.00793-17 ◽

2017 ◽

Vol 83 (15) ◽

Cited By ~ 32

Author(s):

Mohammad Aminul Islam ◽

Moydul Islam ◽

Rashedul Hasan ◽

M. Iqbal Hossain ◽

Ashikun Nabi ◽

...

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Tap Water ◽

Antibiotic Resistance Genes ◽

Multidrug Resistant ◽

Resistant Bacteria ◽

New Delhi ◽

Multidrug Resistant Bacteria ◽

Content Type ◽

Wastewater Samples

ABSTRACT Resistance to carbapenem antibiotics through the production of New Delhi metallo-β-lactamase-1 (NDM-1) constitutes an emerging challenge in the treatment of bacterial infections. To monitor the possible source of the spread of these organisms in Dhaka, Bangladesh, we conducted a comparative analysis of wastewater samples from hospital-adjacent areas (HAR) and from community areas (COM), as well as public tap water samples, for the occurrence and characteristics of NDM-1-producing bacteria. Of 72 HAR samples tested, 51 (71%) samples were positive for NDM-1-producing bacteria, as evidenced by phenotypic tests and the presence of the bla NDM-1 gene, compared to 5 of 41 (12.1%) samples from COM samples (P < 0.001). All tap water samples were negative for NDM-1-producing bacteria. Klebsiella pneumoniae (44%) was the predominant bacterial species among bla NDM-1-positive isolates, followed by Escherichia coli (29%), Acinetobacter spp. (15%), and Enterobacter spp. (9%). These bacteria were also positive for one or more other antibiotic resistance genes, including bla CTX-M-1 (80%), bla CTX-M-15 (63%), bla TEM (76%), bla SHV (33%), bla CMY-2 (16%), bla OXA-48-like (2%), bla OXA-1 (53%), and bla OXA-47-like (60%) genes. Around 40% of the isolates contained a qnr gene, while 50% had 16S rRNA methylase genes. The majority of isolates hosted multiple plasmids, and plasmids of 30 to 50 MDa carrying bla NDM-1 were self-transmissible. Our results highlight a number of issues related to the characteristics and source of spread of multidrug-resistant bacteria as a potential public health threat. In view of the existing practice of discharging untreated liquid waste into the environment, hospitals in Dhaka city contribute to the potential dissemination of NDM-1-producing bacteria into the community. IMPORTANCE Infections caused by carbapenemase-producing Enterobacteriaceae are extremely difficult to manage due to their marked resistance to a wide range of antibiotics. NDM-1 is the most recently described carbapenemase, and the bla NDM-1 gene, which encodes NDM-1, is located on self-transmissible plasmids that also carry a considerable number of other antibiotic resistance genes. The present study shows a high prevalence of NDM-1-producing organisms in the wastewater samples from hospital-adjacent areas as a potential source for the spread of these organisms to community areas in Dhaka, Bangladesh. The study also examines the characteristics of the isolates and their potential to horizontally transmit the resistance determinants. The significance of our research is in identifying the mode of spread of multiple-antibiotic-resistant organisms, which will allow the development of containment measures, leading to broader impacts in reducing their spread to the community.

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