Extensively resistant Acinetobacter baumannii isolate RCH52 carries several resistance genes derived from an IncC plasmid.

OBJECTIVES: To identify the origins of resistance in a sporadic extensively resistant Acinetobacter baumannii isolate. METHODS: The complete genome of RCH52 was determined by combining available Illumina short reads with MinION (Oxford Nanopore) long reads using Unicycler. Bioinformatic searches were used to identify features of interest. RESULTS: The complete genome of RCH52 revealed an unusual chromosomal region containing all of the antibiotic resistance genes, except tet39, which is in a plasmid. A 129 585 bp segment was bounded by inversely oriented copies of ISAba1 and included two groups of resistance genes separated by the large segment of the backbone of type 1 IncC plasmids that lies between the ARI-A and ARI-B resistance islands but does not include the replication region. The ISAba1-bounded segment was located in a novel integrative element that had integrated into the chromosomal thyA gene but provided a replacement thyA gene. Several resistance genes are derived from either the ARI-A or the ARI-B resistance islands found in IncC plasmids that have been brought together by an IS26-mediated deletion of the original plasmid. This non-replicating circular molecule (or translocatable unit) has been incorporated into a smaller ISAba1-bounded unit that includes oxa23 in Tn2008B via homologous recombination between sul2-CR2-floR segments found in both. CONCLUSIONS: The plasmids shared by most Gram-negative pathogens, including the broad host range IncC plasmids, have not been detected in Acinetobacter species. However, it seems likely that they can conjugate into members of this genus and contribute pre-existing clusters of antibiotic resistance genes.

Cl415, a carbapenem-resistant Acinetobacter baumannii isolate containing four AbaR4 and a new variant of AbGRI2, represents a novel global clone 2 strain.

OBJECTIVES: To determine the genetic context of genes conferring antibiotic resistance on the carbapenem-resistant Acinetobacter baumannii Cl415, recovered in 2017 at El Youssef Hospital Centre in Akkar Governorate, North Lebanon. METHODS: Antibiotic resistance phenotype for 22 antibiotics was determined using disc diffusion or MIC determination. The whole-genome sequence of Cl415 was determined using a combination of the Illumina MiSeq and Oxford Nanopore (MinION) platforms. Complete genome was assembled using Unicycler and antibiotic resistance determinants and ISs were identified using ResFinder and ISFinder, respectively. RESULTS: Cl415 is a global clone 2 (GC2) strain and belongs to the most common STs of this clone, ST2IP and ST218OX. Cl415 is resistant to several antibiotics, including aminoglycosides and carbapenems to a high level. Genomic analysis of Cl415 revealed that it carries four chromosomal AbaR4 copies. One copy was found in the comM gene replacing the AbGRI1 island. Cl415 also contains a novel variant of AbGRI2, herein called AbGRI2-15, carrying only the blaTEM and aphA1 resistance genes. Cl415 belongs to a subclade of GC2 strains that appear to have diverged recently with a wide geographical distribution. CONCLUSIONS: The resistance gene complement of Cl415 was found in the chromosome with four oxa23 located in AbaR4 copies and the remaining genes in a novel variant of the AbGRI2 resistance island. Cl415 was isolated in Lebanon, but phylogenetic analysis suggests that Cl415 represents a new lineage with global distribution within GC2.

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. Plasmid sequences were compared to identify factors driving their evolution and divergence. IS26-mediated events, recombination between pdif sites and homologous recombination have all contributed. A translocatable unit that includes oxa58, generated by an IS26-mediated adjacent deletion, had been re-inserted by IS26 adjacent to an IS26 in pACICU1b to create the oxa58 gene duplication previously found in pACICU1. The oxa58 duplication has been lost from pACICU1b and the Tn6020 variant carrying the aphA1 (kanamycin, neomycin resistance) gene in pA388 has been lost from pACICU1/1b via recombination between directly-oriented IS26 copies. Two dif modules located between directly-oriented pdif sites in pA388 have been lost from pACICU1/1b and the product of this and other deletion events as well as inversion of dif modules located between inversely-oriented pdif sites were detected experimentally in pA388 DNA by PCR. Also, the new junctions were detected in a minority of reads in pA388 long-read sequence data. Inversion and deletion were only detected when the spacers in the pdif sites were identical and equivalent events involving mismatched spacers were not detected.

Evidence for a vestigial nematic state in the cuprate pseudogap phase.

The CuO2 antiferromagnetic insulator is transformed by hole-doping into an exotic quantum fluid usually referred to as the pseudogap (PG) phase. Its defining characteristic is a strong suppression of the electronic density-of-states D(E) for energies |E| < [Formula: see text], where [Formula: see text] is the PG energy. Unanticipated broken-symmetry phases have been detected by a wide variety of techniques in the PG regime, most significantly a finite-Q density-wave (DW) state and a Q = 0 nematic (NE) state. Sublattice-phase-resolved imaging of electronic structure allows the doping and energy dependence of these distinct broken-symmetry states to be visualized simultaneously. Using this approach, we show that even though their reported ordering temperatures T DW and T NE are unrelated to each other, both the DW and NE states always exhibit their maximum spectral intensity at the same energy, and using independent measurements that this is the PG energy [Formula: see text] Moreover, no new energy-gap opening coincides with the appearance of the DW state (which should theoretically open an energy gap on the Fermi surface), while the observed PG opening coincides with the appearance of the NE state (which should theoretically be incapable of opening a Fermi-surface gap). We demonstrate how this perplexing phenomenology of thermal transitions and energy-gap opening at the breaking of two highly distinct symmetries may be understood as the natural consequence of a vestigial nematic state within the pseudogap phase of Bi2Sr2CaCu2O8.

An outbreak of multiply antibiotic-resistant ST49:ST128:KL11:OCL8 Acinetobacter baumannii isolates at a Sydney hospital.

OBJECTIVES: To understand the acquisition of resistance genes by a non-GC1, non-GC2 Acinetobacter baumannii strain responsible for a 4 year outbreak at a Sydney hospital. METHODS: Representative isolates were screened for resistance to antibiotics. Three were subjected to WGS using Illumina HiSeq. One genome was completed with MinION long reads. Resistance regions were compared with known sequences using bioinformatics. RESULTS: Isolates were resistant to third-generation cephalosporins, gentamicin and tobramycin, sulfamethoxazole and erythromycin. Sequenced isolates were ST49 (Institut Pasteur scheme) and ST128 (Oxford scheme) and carried KL11 at the capsule locus and OCL8 at the lipooligosaccharide outer core locus. The complete genome of isolate J9 revealed that the resistance genes were all in plasmids; pRAY* contained aadB, and a large plasmid, pJ9-3, contained sul2 and floR genes and a dif module containing the mph(E)-msr(E) macrolide resistance genes. Transposon Tn6168, consisting of a second copy of the chromosomal ampC gene region flanked by ISAba1s, confers resistance to third-generation cephalosporins. Tn6168 is located inside the mph(E)-msr(E) dif module. pJ9-3 includes a set of four dif modules and the orientation of the pdif sites, XerC-XerD or XerD-XerC, alternates. A large transposon, Tn6175, containing tniCABDE transposition genes and genes annotated as being involved in heavy metal metabolism, uptake or export was found in the comM gene. Other ST49:ST128:KL11:OCL8 genomes found in the GenBank WGS database carried Tn6175 but neither of the plasmids carrying the resistance genes. CONCLUSIONS: An early carbapenem-susceptible A. baumannii outbreak recorded in Australia was caused by an unusual clone that had acquired plasmids carrying antibiotic resistance genes.

Emergence and rapid global dissemination of CTX-M-15-associated Klebsiella pneumoniae strain ST307.

OBJECTIVES: Recent reports indicate the emergence of a new carbapenemase-producing Klebsiella pneumoniae clone, ST307. We sought to better understand the global epidemiology and evolution of this clone and evaluate its association with antimicrobial resistance (AMR) genes. METHODS: We collated information from the literature and public databases and performed a comparative analysis of 95 ST307 genomes (including 37 that were newly sequenced). RESULTS: We show that ST307 emerged in the mid-1990s (nearly 20 years prior to its first report), is already globally distributed and is intimately associated with a conserved plasmid harbouring the blaCTX-M-15 ESBL gene and several other AMR determinants. CONCLUSIONS: Our findings support the need for enhanced surveillance of this widespread ESBL clone in which carbapenem resistance has occasionally emerged.

The AbaR antibiotic resistance islands found in Acinetobacter baumannii global clone 1 - Structure, origin and evolution.

In multiply resistant Acinetobacter baumannii, complex transposons located in the chromosomal comM gene carry antibiotic and heavy metal resistance determinants. For one type, known collectively as AbaR, the ancestral form, AbaR0, entered a member of global clone 1 (GC1) in the mid 1970s and continued to evolve in situ forming many variants. In AbaR0, antibiotic and mercuric ion resistance genes are located between copies of a cadmium-zinc resistance transposon, Tn6018, and this composite transposon is in a class III transposon, Tn6019, carrying arsenate/arsenite resistance genes and five tni transposition genes. The antibiotic resistance genes in the AbaR0 and derived AbaR3 configurations are aphA1b, blaTEM, catA1, sul1, tetA(A), and cassette-associated aacC1 and aadA1 genes. These genes are in a specific arrangement of fragments from well-known transposons, e.g. Tn1, Tn1721, Tn1696 and Tn2670, that arose in an IncM1 plasmid. All known GC1 lineage 1 isolates carry AbaR0 or AbaR3, which arose around 1990, or a variant derived from one of them. Variants arose via deletions caused by one of three internal IS26s, by recombination between duplicate copies of sul1 or Tn6018, or by gene cassette addition or replacement. A few GC2 isolates also carry an AbaR island with different cassette-associated genes, aacA4 and oxa20.

Commensurate 4a0-period charge density modulations throughout the Bi2Sr2CaCu2O8+x pseudogap regime.

Theories based upon strong real space (r-space) electron-electron interactions have long predicted that unidirectional charge density modulations (CDMs) with four-unit-cell (4a0) periodicity should occur in the hole-doped cuprate Mott insulator (MI). Experimentally, however, increasing the hole density p is reported to cause the conventionally defined wavevector QA of the CDM to evolve continuously as if driven primarily by momentum-space (k-space) effects. Here we introduce phase-resolved electronic structure visualization for determination of the cuprate CDM wavevector. Remarkably, this technique reveals a virtually doping-independent locking of the local CDM wavevector at [Formula: see text] throughout the underdoped phase diagram of the canonical cuprate Bi2Sr2CaCu2O8 These observations have significant fundamental consequences because they are orthogonal to a k-space (Fermi-surface)-based picture of the cuprate CDMs but are consistent with strong-coupling r-space-based theories. Our findings imply that it is the latter that provides the intrinsic organizational principle for the cuprate CDM state.

RCH51, a multiply antibiotic-resistant Acinetobacter baumannii ST103IP isolate, carries resistance genes in three plasmids, including a novel potentially conjugative plasmid carrying oxa235 in transposon Tn6252.

Objectives: To determine the identity and context of genes conferring antibiotic resistance in a sporadic multiply antibiotic-resistant Acinetobacter baumannii recovered at Royal Children's Hospital, Brisbane. Methods: The antibiotic resistance phenotype for 23 antibiotics was determined using disc diffusion or MIC determination. The whole-genome sequence of RCH51 was determined using the Illumina HiSeq platform. Antibiotic resistance determinants were identified using ResFinder. Plasmids were recovered by transformation. Results: Isolate RCH51 belongs to the uncommon STs ST103 IP (7-3-2-1-7-1-4) and ST514 OX (1-52-29-28-18-114-7). It was found to be resistant to sulfamethoxazole, tetracycline, gentamicin, tobramycin and kanamycin and also exhibited reduced susceptibility to imipenem (MIC 2 mg/L) and meropenem (MIC 6 mg/L). RCH51 carries the oxa235 , sul2 , floR , aadB and tet39 resistance genes, all located on plasmids. The largest of the three plasmids, pRCH51-3, is 52 789 bp and carries oxa235 in the ISAba1-bounded transposon Tn 6252 , as well as sul2 and floR . pRCH51-3 represents a new A. baumannii plasmid family that is potentially conjugative as it contains several genes predicted to encode transfer functions. However, conjugation of pRCH51-3 was not detected. The aadB and tet39 resistance genes were each found in small plasmids identical to the known plasmids pRAY*-v1 and pRCH52-1, respectively. Conclusions: The resistance gene complement of RCH51 was found in three plasmids. pRCH51-3, which carries the oxa235 , sul2 and floR resistance genes, represents a new, potentially conjugative A. baumannii plasmid type.

pIP40a, a type 1 IncC plasmid from 1969 carries the integrative element GIsul2 and a novel class II mercury resistance transposon.

The 167.5kb sequence of the conjugative IncC plasmid pIP40a, isolated from a Pseudomonas aeruginosa in 1969, was analysed. pIP40a confers resistance to kanamycin, neomycin, ampicillin, sulphonamides and mercuric ions, and several insertions in a type 1 IncC backbone were found, including copies of IS3, Tn1000 and a novel mercury resistance transposon, Tn6182. The antibiotic resistance genes were in two locations. Tn6023, containing the aphA1 kanamycin and neomycin resistance gene, is in a partial copy of Tn1/Tn2/Tn3 (blaTEM, ampicillin resistance) in the kfrA gene, and the sul2 sulphonamide resistance gene is in the integrative element GIsul2 in the position of ARI-B islands. The 11.5kb class II transposon Tn6182 is only distantly related to other class II transposons, with at most 33% identity between the TnpA of Tn6182 and TnpA of other group members. In addition, the inverted repeats are 37bp rather than 38bp, and the likely resolution enzyme is a tyrosine recombinase (TnpI). Re-annotation of GIsul2 revealed genes predicted to confer resistance to arsenate and arsenite, but resistance was not detected. The location of GIsul2 confirms it as the progenitor of the ARI-B configurations seen in many IncC plasmids isolated more recently. However, GIsul2 has integrated at the same site in type 1 and type 2 IncC plasmids, indicating that it targets this site. Analysis of the distribution of GIsul2 revealed that it in addition to its chromosomal integration site at the 3'-end of the guaA gene, it has also integrated into other plasmids, increasing its mobility.

Direct phase-sensitive identification of a d-form factor density wave in underdoped cuprates.

The identity of the fundamental broken symmetry (if any) in the underdoped cuprates is unresolved. However, evidence has been accumulating that this state may be an unconventional density wave. Here we carry out site-specific measurements within each CuO2 unit cell, segregating the results into three separate electronic structure images containing only the Cu sites [Cu(r)] and only the x/y axis O sites [Ox(r) and O(y)(r)]. Phase-resolved Fourier analysis reveals directly that the modulations in the O(x)(r) and O(y)(r) sublattice images consistently exhibit a relative phase of π. We confirm this discovery on two highly distinct cuprate compounds, ruling out tunnel matrix-element and materials-specific systematics. These observations demonstrate by direct sublattice phase-resolved visualization that the density wave found in underdoped cuprates consists of modulations of the intraunit-cell states that exhibit a predominantly d-symmetry form factor.

A large conjugative Acinetobacter baumannii plasmid carrying the sul2 sulphonamide and strAB streptomycin resistance genes.

Acinetobacter baumannii is an important nosocomial pathogen that often complicates treatment because of its high level of resistance to antibiotics. Though plasmids can potentially introduce various genes into bacterial strains, compared to other Gram-negative bacteria, information about the unique A. baumannii plasmid repertoire is limited. Here, whole genome sequence data was used to determine the plasmid content of strain A297 (RUH875), the reference strain for the globally disseminated multiply resistant A. baumannii clone, global clone 1(GC1). A297 contains three plasmids. Two known plasmids were present; one, pA297-1 (pRAY*), carries the aadB gentamicin, kanamycin and tobramycin resistance gene and another is an 8.7kb cryptic plasmid often found in GC1 isolates. The third plasmid, pA297-3, is 200kb and carries the sul2 sulphonamide resistance gene and strAB streptomycin resistance gene within Tn6172 and a mer mercuric ion resistance module elsewhere. pA297-3 transferred sulphonamide, streptomycin and mercuric ion resistance at high frequency to a susceptible A. baumannii recipient, and contains several genes potentially involved in conjugative transfer. However, a relaxase gene was not found. It also includes several genes encoding proteins involved in DNA metabolism such as partitioning. However, a gene encoding a replication initiation protein could not be found. pA297-3 includes two copies of a Miniature Inverted-Repeat Transposable Element (MITE), named MITE-297, bracketing a 77.5kb fragment, which contains several IS and the mer module. Several plasmids related to but smaller than pA297-3 were found in the GenBank nucleotide database. They were found in different A. baumannii clones and are wide spread. They all contain either Tn6172 or a variant in the same position in the backbone as Tn6172 in pA297-3. Some related plasmids have lost the segment between the MITE-297 copies and retain only one MITE-297. Others have segments of various lengths between two MITE-297 copies, and these can be derived from the region in pA297-3 via a deletion adjacent to IS related to IS26 such as IS1007 or IS1007-like. pA297-3 and its relatives represent a third type of conjugative Acinetobacter plasmid that contributes to the dissemination of antibiotic resistance in this species.

Destabilization of IncA and IncC plasmids by SGI1 and SGI2 type Salmonella genomic islands.

Both the Salmonella genomic islands (SGI) and the conjugative IncC plasmids are known to contribute substantially to the acquisition of resistance to multiple antibiotics, and plasmids in the A/C group are known to mobilize the Salmonella genomic island SGI1, which also carries multiple antibiotic resistance genes. Plasmid pRMH760 (IncC; A/C2) was shown to mobilize SGI1 variants SGI1-I, SGI1-F, SGI1-K and SGI2 from Salmonella enterica to Escherichia coli where it was integrated at the preferred location, at the end of the trmE (thdF) gene. The plasmid was transferred at a similar frequency. However, we observed that co-transfer of the SGI and the plasmid was rarer. In E. coli to E. coli transfer, the frequency of transfer of the IncC plasmid pRMH760 was at least 1000-fold lower when the donor carried SGI1-I or SGI1-K, indicating that the SGI suppresses transfer of the plasmid. In addition, pRMH760 was rapidly lost from both E. coli and S. enterica strains that also carried SGI1-I, SGI1-F or SGI2. However, plasmid loss was not seen when the SGI1 variant was SGI1-K, which lacks two segments of the SGI1 backbone. The complete sequence of the SGI1-I and SGI1-F were determined and SGI1-K also carries two single base substitutions relative to SGI1-I. The IncA (A/C1) plasmid RA1 was also shown to mobilize SGI2-A and though there are significant differences between the backbones of IncA and IncC plasmids, RA1 was also rapidly lost when SGI2-A was present in the same cell. We conclude that there are multiple interactions, both cooperative and antagonistic, between an IncA or IncC plasmid and the SGI1 and SGI2 family genomic islands.

ISMapper: identifying transposase insertion sites in bacterial genomes from short read sequence data.

BACKGROUND: Insertion sequences (IS) are small transposable elements, commonly found in bacterial genomes. Identifying the location of IS in bacterial genomes can be useful for a variety of purposes including epidemiological tracking and predicting antibiotic resistance. However IS are commonly present in multiple copies in a single genome, which complicates genome assembly and the identification of IS insertion sites. Here we present ISMapper, a mapping-based tool for identification of the site and orientation of IS insertions in bacterial genomes, directly from paired-end short read data. RESULTS: ISMapper was validated using three types of short read data: (i) simulated reads from a variety of species, (ii) Illumina reads from 5 isolates for which finished genome sequences were available for comparison, and (iii) Illumina reads from 7 Acinetobacter baumannii isolates for which predicted IS locations were tested using PCR. A total of 20 genomes, including 13 species and 32 distinct IS, were used for validation. ISMapper correctly identified 97 % of known IS insertions in the analysis of simulated reads, and 98 % in real Illumina reads. Subsampling of real Illumina reads to lower depths indicated ISMapper was able to correctly detect insertions for average genome-wide read depths >20x, although read depths >50x were required to obtain confident calls that were highly-supported by evidence from reads. All ISAba1 insertions identified by ISMapper in the A. baumannii genomes were confirmed by PCR. In each A. baumannii genome, ISMapper successfully identified an IS insertion upstream of the ampC beta-lactamase that could explain phenotypic resistance to third-generation cephalosporins. The utility of ISMapper was further demonstrated by profiling genome-wide IS6110 insertions in 138 publicly available Mycobacterium tuberculosis genomes, revealing lineage-specific insertions and multiple insertion hotspots. CONCLUSIONS: ISMapper provides a rapid and robust method for identifying IS insertion sites directly from short read data, with a high degree of accuracy demonstrated across a wide range of bacteria.

Genomic resistance island AGI1 carrying a complex class 1 integron in a multiply antibiotic-resistant ST25 Acinetobacter baumannii isolate.

OBJECTIVES: The objective of this study was to locate the antibiotic resistance determinants in the multiply antibiotic-resistant Acinetobacter baumannii isolate D4. METHODS: The genome was sequenced using Illumina HiSeq and assembled de novo using Velvet. PCR was used to link the relevant contigs and fill the gaps. Sequences were compared with ones in GenBank and annotated. RESULTS: A sporadic A. baumannii isolate D4, recovered in Sydney in 2006 from a wound, was multiply antibiotic resistant. D4 is ST25 (Institut Pasteur scheme) and exhibited resistance to third-generation cephalosporins and reduced susceptibility to ciprofloxacin, as well as resistance to aminoglycosides (gentamicin, kanamycin, neomycin and tobramycin) and further older antibiotics, nalidixic acid, sulfamethoxazole, streptomycin, spectinomycin and trimethoprim. The gyrA gene has a mutation consistent with nalidixic acid resistance. The bla PER conferring cephalosporin resistance, together with the aadB, aadA13/2, aadA2, strAB and sul1 resistance genes, are located within a 29 173 bp complex class 1 integron that includes three copies of intI1, three cassette arrays and two copies of the 3'-conserved segment. This integron is adjacent to the resG gene of an integrative genomic resistance island, AGI1 (Acinetobacter genomic island 1), with a backbone related to that of islands in the SGI1, SGI2 and PGI1 families. AGI1 is located at the 3'-end of the chromosomal trmE (formerly thdF) gene. CONCLUSIONS: AGI1 represents a new lineage of genomic resistance islands that belongs in the same broad group as members of the SGI1, SGI2 and PGI1 families. Genes conferring resistance to cephalosporins, aminoglycosides and sulphonamides are located in a complex class 1 integron within AGI1.

Tn6168, a transposon carrying an ISAba1-activated ampC gene and conferring cephalosporin resistance in Acinetobacter baumannii.

OBJECTIVES: To explore the cause of third-generation cephalosporin resistance in Australian Acinetobacter baumannii isolates belonging to global clone 1 (GC1). METHODS: GC1 isolates from Australia were tested for resistance to ceftazidime and cefotaxime using disc diffusion and MICs. PCR was used to determine the context of ISAba1-ampC configurations and amplicons were sequenced. The level of transcripts was measured using quantitative real-time PCR. Multilocus sequence typing was performed. RESULTS: All ceftazidime- and cefotaxime-resistant isolates carried an appropriately oriented ISAba1 adjacent to the ampC gene and ISAba1 increased ampC transcripts 8-12-fold. In three isolates, the ampC gene next to ISAba1 was not in the normal chromosomal position. Instead, ISAba1 was 7 bp upstream of an additional copy of ampC located in a 3155 bp duplicated segment of the chromosome that differs from the resident GC1 segment by 2.3% but is almost identical to the corresponding region in several non-GC1 draft genomes. The duplicated segment is bounded by directly oriented copies of ISAba1 and flanked by a 9 bp direct duplication. This 5.5 kb transposon, named Tn6168, is in the same position in the chromosome of the three Australian isolates and the GC1 isolate AB0057. Tn6168 was also detected in an unrelated A. baumannii strain, where it was in a different location. The central part of Tn6168 was probably acquired from a sequence type ST32 (Institut Pasteur scheme) A. baumannii strain. CONCLUSIONS: The ISAba1-ampC configuration, which increases ampC expression, can be part of a composite transposon Tn6168.

A GC1 Acinetobacter baumannii isolate carrying AbaR3 and the aminoglycoside resistance transposon TnaphA6 in a conjugative plasmid.

OBJECTIVES: To locate the acquired antibiotic resistance genes, including the amikacin resistance transposon TnaphA6, in the genome of an Australian isolate belonging to Acinetobacter baumannii global clone 1 (GC1). METHODS: A multiply antibiotic-resistant GC1 isolate harbouring TnaphA6 was sequenced using Illumina HiSeq, and reads were used to generate a de novo assembly and determine multilocus sequence types (STs). PCR was used to assemble the AbaR chromosomal resistance island and a large plasmid carrying TnaphA6. Plasmid DNA sequences were compared with ones available in GenBank. Conjugation experiments were conducted. RESULTS: The A. baumannii GC1 isolate G7 was shown to include the AbaR3 antibiotic resistance island. It also contains an 8.7 kb cryptic plasmid, pAb-G7-1, and a 70,100 bp plasmid, pAb-G7-2, carrying TnaphA6. pAb-G7-2 belongs to the Aci6 Acinetobacter plasmid family. It encodes transfer functions and was shown to conjugate. Plasmids related to pAb-G7-2 were detected in further amikacin-resistant GC1 isolates using PCR. From the genome sequence, isolate G7 was ST1 (Institut Pasteur scheme) and ST231 (Oxford scheme). Using Oxford scheme PCR-based methods, the isolate was ST109 and this difference was traced to a single base difference resulting from the inclusion of the original primers in the gpi segment analysed. CONCLUSIONS: The multiply antibiotic-resistant GC1 isolate G7 carries most of its resistance genes in AbaR3 located in the chromosome. However, TnaphA6 is on a conjugative plasmid, pAb-G7-2. Primers developed to locate TnaphA6 in pAb-G7-2 will simplify the detection of plasmids related to pAb-G7-2 in A. baumannii isolates.

A conjugative plasmid carrying the carbapenem resistance gene blaOXA-23 in AbaR4 in an extensively resistant GC1 Acinetobacter baumannii isolate.

OBJECTIVES: To locate the acquired bla(OXA-23) carbapenem resistance gene in an Australian A. baumannii global clone 1 (GC1) isolate. METHODS: The genome of the extensively antibiotic-resistant GC1 isolate A85 harbouring bla(OXA-23) in Tn2006 was sequenced using Illumina HiSeq, and the reads were used to generate a de novo assembly. PCR was used to assemble relevant contigs. Sequences were compared with ones in GenBank. Conjugation experiments were conducted. RESULTS: The sporadic GC1 isolate A85, recovered in 2003, was extensively resistant, exhibiting resistance to imipenem, meropenem and ticarcillin/clavulanate, to cephalosporins and fluoroquinolones and to the older antibiotics gentamicin, kanamycin and neomycin, sulfamethoxazole, trimethoprim and tetracycline. Genes for resistance to older antibiotics are in the chromosome, in an AbaR3 resistance island. A second copy of the ampC gene in Tn6168 confers cephalosporin resistance and the gyrA and parC genes have mutations leading to fluoroquinolone resistance. An 86 335 bp repAci6 plasmid, pA85-3, carrying bla(OXA-23) in Tn2006 in AbaR4, was shown to transfer imipenem, meropenem and ticarcillin/clavulanate resistance into a susceptible recipient. A85 also contains two small cryptic plasmids of 2.7 and 8.7 kb. A85 is sequence type ST126 (Oxford scheme) and carries a novel KL15 capsule locus and the OCL3 outer core locus. CONCLUSIONS: A85 represents a new GC1 lineage identified by the novel capsule locus but retains AbaR3 carrying genes for resistance to older antibiotics. Resistance to imipenem, meropenem and ticarcillin/clavulanate has been introduced into A85 by pA85-3, a repAci6 conjugative plasmid carrying Tn2006 in AbaR4.

How Kondo-holes create intense nanoscale heavy-fermion hybridization disorder.

Replacing a magnetic atom by a spinless atom in a heavy-fermion compound generates a quantum state often referred to as a "Kondo-hole". No experimental imaging has been achieved of the atomic-scale electronic structure of a Kondo-hole, or of their destructive impact [Lawrence JM, et al. (1996) Phys Rev B 53:12559-12562] [Bauer ED, et al. (2011) Proc Natl Acad Sci. 108:6857-6861] on the hybridization process between conduction and localized electrons which generates the heavy-fermion state. Here we report visualization of the electronic structure at Kondo-holes created by substituting spinless thorium atoms for magnetic uranium atoms in the heavy-fermion system URu(2)Si(2). At each thorium atom, an electronic bound state is observed. Moreover, surrounding each thorium atom we find the unusual modulations of hybridization strength recently predicted to occur at Kondo-holes [Figgins J, Morr DK (2011) Phys Rev Lett 107:066401]. Then, by introducing the "hybridization gapmap" technique to heavy-fermion studies, we discover intense nanoscale heterogeneity of hybridization due to a combination of the randomness of Kondo-hole sites and the long-range nature of the hybridization oscillations. These observations provide direct insight into both the microscopic processes of heavy-fermion forming hybridization and the macroscopic effects of Kondo-hole doping.

Evaluating phytoremediation potential and nutrients status of Bassia indica (Wight) A. J. Scott (Indian Bassia) in a cadmium-contaminated saline soil.

Bassia indica (Wight) A. J. Scott is a fast-growing halophyte suitable for the remediation of saline lands on a large scale. However, no information is available regarding its phytoremediation potential for cadmium (Cd) alone or in combination with salinity. Besides evaluating phytoremediation, assessing micronutrient hemostasis as a crucial physiological insight into the mechanism involved in the tolerance of B. indica under saline soil contaminated with Cd was subjected. Under salinity stress, a considerable amount of sodium accumulates in the plant. Moreover, the accumulation of sodium increased by Cd stress levels. The increase in the exchangeable form of Cd in the rhizosphere in the presence of NaCl ions further elevated the Cd content in the plant tissues. For instance, compared to non-saline conditions, applying 2.5 and 5 g NaCl kg-1 to soil treated with 60 mg Cd kg-1 increased exchangeable Cd by 28.4 and 49.5% in rhizosphere soil, which led to increased cadmium content by 16.1 and 29.6% in the root (as a main part of Cd accumulation), respectively. Under most stress conditions, potassium homeostasis in the shoot remained undisturbed. It was observed that this plant could transfer an optimal level of potassium from the roots to the shoots at a moderate salinity level. Changes and the distribution of Cu and Zn levels followed a similar pattern in the plant, indicating a common regulation mechanism for these nutrients. Generally, the plant could maintain an appropriate level of Fe, Zn, and Cu ions under most stressed conditions. However, the level of Mn decreased significantly under severe stress levels. Growth parameters, tolerance index, and the values of translocation factor < 1 and shoot bioconcentration factor > 1 under 5 mg Cd kg-1 soil treatment at different salinity levels indicated that B. indica could mitigate the detrimental effect of Cd toxicity and tolerate the NaCl stress via a phytostabilizer mechanism. However, the shoot bioconcentration factor values were very close to one at other Cd levels. Therefore, considering the obtained evidence and the innate ability of B. indica to remediation salinity, this plant is still recommended, even for higher Cd levels (even until 30 mg kg-1), in the presence of salinity.