Distribution and expression of the aac(6')-Im (aacA16) aminoglycoside resistance gene.

Background: The aac(6')-Im (aacA16) amikacin, netilmicin and tobramycin resistance gene cassette had been circulating globally undetected for many years in a sublineage of Acinetobacter baumannii global clone 2.

Objectives: To identify sources for the aac(6')-Im fragment found in A. baumannii.

Aminoglycoside resistance genes in early members of the Acinetobacter baumannii ST78A (SMAL, Italian clone) reside in an IS26-bounded island in the chromosome.

Background: The Acinetobacter baumannii isolate called SMAL, previously used to determine the structures of capsular polysaccharide and lipooligosaccharide, was recovered in Pavia, Italy in 2002 among the collection of aminoglycoside-resistant isolates designated as SMAL type. This type was later called the Italian clone, then ST78. ST78 isolates are now widely distributed.

IS and the IS family: versatile resistance gene movers and genome reorganizers.

In Gram-negative bacteria, the insertion sequence IS is highly active in disseminating antibiotic resistance genes. IS can recruit a gene or group of genes into the mobile gene pool and support their continued dissemination to new locations by creating pseudo-compound transposons (PCTs) that can be further mobilized by the insertion sequence (IS). IS can also enhance expression of adjacent potential resistance genes.

Acinetobacter baumannii GC2 Sublineage Carrying the (')- Amikacin, Netilmicin, and Tobramycin Resistance Gene Cassette.

The aminoglycoside antibiotics amikacin, gentamicin, and tobramycin are important therapeutic options for Acinetobacter iinfections. Several genes that confer resistance to one or more of these antibiotics are prevalent in the globally distributed resistant clones of Acinetobacter baumannii, but the (')- () gene (amikacin, netilmicin, and tobramycin resistance), first reported in isolates from South Korea, has rarely been reported since. In this study, GC2 isolates (1999 to 2002) from Brisbane, Australia, carrying (')- and belonging to the ST2:ST423:KL6:OCL1 type were identified and sequenced.

The RuvABC Holliday Junction Processing System Is Not Required for IS-Mediated Targeted Conservative Cointegrate Formation.

The insertion sequence IS plays a key role in the spread of antibiotic resistance genes in Gram-negative bacteria. IS and members of the IS family are able to use two distinct mechanisms to form cointegrates made up of two DNA molecules linked via directly oriented copies of the IS. The well-known copy-in (formerly replicative) reaction occurs at very low frequency, and the more recently discovered targeted conservative reaction, which joins two molecules that already include an IS, is substantially more efficient.

Characterisation of an early South African multiply antibiotic-resistant global clone 1 Acinetobacter baumannii isolate.

Objectives: The aim of this study was to characterise an early clinical multiply antibiotic resistant Acinetobacter baumannii global clone 1 (GC1) isolate from Africa.

Methods: The draft genome sequence was determined using short-read (Illumina MiSeq) sequence data and compared to other early GC1 isolates. Resistance genes and other features were identified using various bioinformatics tools.

Identification of an Outbreak Cluster of Extensively Antibiotic-Resistant GC1 Acinetobacter baumannii Isolates in U.S. Military Treatment Facilities.

An outbreak involving an extensively antibiotic-resistant Acinetobacter baumannii strain in three military treatment facilities was identified. Fifty-nine isolates recovered from 30 patients over a 4-year period were found among a large collection of isolates using core genome multilocus sequence typing (MLST). They differed by only 0 to 18 single nucleotide polymorphisms (SNPs) and carried the same resistance determinants except that the gene was missing in 25 isolates.

Insertion sequences related to ISAjo2 target p and sites and belong to a new IS family, the IS family.

Several insertion sequences (IS) found in various species exhibit target specificity. They are found, in the same orientation, 5 bp from the XerC binding site of the p sites associated with modules in plasmids, and searches revealed they are also found near chromosomal sites of species. These IS are 1.

IS26-mediated loss of the translocatable unit from Tn4352B requires the presence of the recA1 allele.

The pseudo-compound transposon Tn4352B is unusual in that the translocatable unit (TU) consisting of one of the bounding IS26 copies and the central portion containing the aphA1a gene has been found to be readily lost in the Escherichia coli strains used as host. Rapid loss required the presence of an additional 2 G residues adjacent to the internal end of one of the IS26 that flank the central portion and an active Tnp26 transposase. However, Tn4352B was found to be stable in wild-type Klebsiella pneumoniae strains.

Mechanisms of IS-Mediated Amplification of the Gene Leading to Tobramycin Resistance in an Acinetobacter baumannii Isolate.

Enhanced levels of resistance to antibiotics arising from amplification of an antibiotic resistance gene that impact therapeutic options are increasingly observed. Amplification can also disclose novel phenotypes leading to treatment failure. However, the mechanism is poorly understood.

Complete genome of the extensively antibiotic-resistant GC1 Acinetobacter baumannii isolate MRSN 56 reveals a novel route to fluoroquinolone resistance.

Objectives: To examine the causes of antibiotic resistance in the extensively resistant global clone 1 (GC1) Acinetobacter baumannii isolate MRSN 56 recovered at a US military treatment facility.

Methods: MRSN 56 was sequenced using MinION (Oxford Nanopore) and the reads combined with available Illumina MiSeq data using Unicycler. Acquired resistance genes were identified using ABRicate and their environment examined.

Evolution of Acinetobacter baumannii plasmids carrying the oxa58 carbapenemase resistance gene via plasmid fusion, IS26-mediated events and dif module shuffling.

Acinetobacter baumannii RepAci1-RepAci10 plasmids pA388 from a global clone 1 (GC1) isolate from Greece, and pACICU1 and variant pACICU1b from an Italian GC2 isolate were found to share a common ancestor. The ancestor formed via recombination between pdif sites in the widely-distributed RepAci1 plasmid pA1-1 and in a RepAci10 plasmid carrying the oxa58 carbapenem-resistance gene in a dif module. Each plasmid includes copies of IS26 and multiple dif modules surrounded by 28 bp pdif sites resembling the chromosomal dif site, including one carrying the oxa58 gene.

Variants of Tn, a Novel Tn Family Transposon Carrying the Metallo-β-Lactamase and 14 Copies of the Amikacin Resistance Genes Found in Acinetobacter baumannii.

Carbapenem resistance in Acinetobacter baumannii is primarily due to the global spread of two main clones that carry , , and However, new carbapenem-resistant clones are emerging that are also resistant to a wide range of antibiotics. Strains belonging to ST85 (Institut Pasteur) carry the metallo-β-lactamase carbapenem resistance gene. Here, we completed the genome sequence of an ST85 strain, Cl300, recovered in 2015 in Lebanon, using a combination of Illumina MiSeq and Oxford Nanopore sequencing and a hybrid assembly approach.

Characterization of the specific DNA-binding properties of Tnp26, the transposase of insertion sequence IS26.

The bacterial insertion sequence (IS) IS26 mobilizes and disseminates antibiotic resistance genes. It differs from bacterial IS that have been studied to date as it exclusively forms cointegrates via either a copy-in (replicative) or a recently discovered targeted conservative mode. To investigate how the Tnp26 transposase recognizes the 14-bp terminal inverted repeats (TIRs) that bound the IS, amino acids in two domains in the N-terminal (amino acids M1-P56) region were replaced.

HI1 and I1 Resistance Plasmids from Serovar Typhimurium Strain SRC27 Are Epidemic.

Conjugative plasmids are a major contributor to the global spread of antibiotic resistance determinants, but the tracking of their evolutionary history is often neglected. serovar Typhimurium ( Typhimurium) strain SRC27 was isolated from an equine infection in Australia in 1999. SRC27 was known to carry conjugative HI1 and I1 resistance plasmids.

IS26 cannot move alone.

Background: IS26 plays a major role in the dissemination of antibiotic resistance determinants in Gram-negative bacteria.

Objectives: To determine whether insertion sequence IS26 is able to move alone (simple transposition) or if it exclusively forms cointegrates.

Methods: A two-step PCR using outward-facing primers was used to search for circular IS26 molecules.

Targeted Conservative Cointegrate Formation Mediated by IS Family Members Requires Sequence Identity at the Reacting End.

IS forms cointegrates using two distinct routes, a copy-in mechanism involving one insertion sequence (IS) and a target and a targeted conservative mechanism involving two ISs in different DNA molecules. In this study, the ability of IS and some close relatives, IS, IS, and a natural hybrid, IS, which are found predominantly in spp., to interact was examined.

Structures bounded by directly-oriented members of the IS26 family are pseudo-compound transposons.

Antibiotic resistance genes are often found in structures bounded by copies of IS26, IS257/IS431 or IS1216 that resemble compound (or composite) transposons. However, because of the mechanisms used by IS26 family members, namely that they form cointegrates but cannot resolve them, none of these structures can move together as a coherent single unit. Apparent transposition of these structures is possible via a 2-step process but only if the IS are in direct orientation.

IS Family Members IS and IS Also Form Cointegrates by Copy-In and Targeted Conservative Routes.

IS has been shown to form cointegrates both by a copy-in mechanism involving one insertion sequence (IS) and a target and by a targeted conservative mechanism involving two ISs. IS is the flagship of a group of 65 bacterial ISs in the recently redefined IS/IS family. Here, whether other family members can also use two mechanisms was examined using members of the IS/IS and IS isoform groups, which are associated with antibiotic resistance genes in staphylococci and enterococci, respectively.

The Complete Nucleotide Sequence of pZM3, a 1970 FIA:FIB:FII Plasmid Carrying Antibiotic Resistance and Virulence Determinants.

The multiresistance plasmid, pZM3, from a 1970 serovar Wien isolate from Algeria represents the multiresistance FI-type plasmids conferring resistance to ampicillin, chloramphenicol, kanamycin, neomycin, sulfonamides, streptomycin, spectinomycin, tetracycline, and mercuric ions circulating in the Middle East in the 1970s. pZM3 was sequenced to determine the relationship between IS, the IS-like insertion sequence it carries, and IS. IS is identical to IS.

An analysis of the IS/IS family of insertion sequences: is it a single family?

The relationships within a curated set of 112 insertion sequences (ISs) currently assigned to the IS family, here re-named the IS/IS family, in the ISFinder database were examined. The encoded DDE transposases include a helix-helix-turn-helix (H-HTH) potential DNA binding domain N-terminal to the catalytic (DDE) domain, but 10 from include one or two additional N-terminal domains. The transposase phylogeny clearly separated 75 derived from bacteria from 37 from archaea.

An IS26 variant with enhanced activity.

The insertion sequence IS26 plays a major role in the mobilization, expression and dissemination of antibiotic resistance genes in Gram-negative bacteria. Though IS26 is abundant in sequenced genomes and in plasmids that harbour antibiotic resistance genes, only a few minor variations in the IS26 sequence have been recorded. The most common variant, IS26* (also known as IS15Δ1), encodes a Tnp26 transposase with a single amino acid substitution, G184N in the catalytic domain.

Evolution and typing of IncC plasmids contributing to antibiotic resistance in Gram-negative bacteria.

The large, broad host range IncC plasmids are important contributors to the spread of key antibiotic resistance genes and over 200 complete sequences of IncC plasmids have been reported. To track the spread of these plasmids accurate typing to identify the closest relatives is needed. However, typing can be complicated by the high variability in resistance gene content and various typing methods that rely on features of the conserved backbone have been developed.