Zhiyong Zong

Dominance of blaCTX-M within an Australian Extended-Spectrum β-Lactamase Gene Pool

ABSTRACT bla CTX-M genes, particularly blaCTX-M-15, are the dominant extended-spectrum β-lactamase (ESBL) genes among clinical isolates of Escherichia coli and Klebsiella pneumoniae in Sydney, Australia, where we also found one example of blaCTX-M-62, encoding a novel enzyme conferring ceftazidime resistance. ESBL genes were present in diverse community isolates and in a variety of associated conjugative plasmids.

Recombination in IS26 and Tn2 in the Evolution of Multiresistance Regions Carrying blaCTX-M-15 on Conjugative IncF Plasmids from Escherichia coli

ABSTRACT CTX-M-15 now appears to be the dominant extended-spectrum β-lactamase worldwide, and a number of different factors may contribute to this success. These include associations between blaCTX-M-15 and particular plasmids (IncF) and/or strains, such as Escherichia coli ST131, as well as the genetic contexts in which this gene is found. We previously identified blaCTX-M-15 as the dominant ESBL gene in the western Sydney area, Australia, and found that it was carried mainly on IncF or IncI1 plasmids. Here, we have mapped the multiresistance regions of the 11 conjugative plasmids with one or more IncF replicons obtained from that survey and conducted a limited comparison of plasmid backbones. Two plasmids with only an IncFII replicon appear to be very similar to the published plasmids pC15-1a and pEK516. The remaining nine plasmids, with multiple IncF replicons, have multiresistance regions related to those of pC15-1a and pEK516, but eight contain additional modules previously found in resistance plasmids from different geographic locations that carry a variety of different resistance genes. Differences between the multiresistance regions are largely due to IS26-mediated deletions, insertions, and/or rearrangements, which can explain the observed variable associations between blaCTX-M-15 and certain other resistance genes. We found no evidence of independent movement of blaCTX-M-15 or of a large multiresistance region between different plasmid backbones. Instead, homologous recombination between common components, such as IS26 and Tn2, appeared to be more important in creating new multiresistance regions, and this may be coupled with recombination in plasmid backbones to reassort multiple IncF replicons as well as components of multiresistance regions.

Complete Sequence of pJIE143, a pir-Type Plasmid Carrying ISEcp1-blaCTX-M-15 from an Escherichia coli ST131 Isolate

ABSTRACT pJIE143 (34 kb), from an Escherichia coli ST131 isolate, carries blaCTX-M-15 but could not be typed using the standard PCR-based replicon-typing primer set. Complete sequencing revealed a backbone with similarity to IncX plasmids, including a pir-like gene encoding a π-like replication protein and iterons related to those of other IncX plasmids. The 2.971-kb ISEcp1-blaCTX-M-15-orf477Δ transposition unit often found within Tn2 is inserted just beyond the end of pir, flanked by 5-bp direct repeats.

Limited diversity in the gene pool allows prediction of third-generation cephalosporin and aminoglycoside resistance in Escherichia coli and Klebsiella pneumoniae

Early appropriate antibiotic treatment reduces mortality in severe sepsis, but current methods to identify antibiotic resistance still generally rely on bacterial culture. Modern diagnostics promise rapid gene detection, but the apparent diversity of relevant resistance genes in Enterobacteriaceae is a problem. Local surveys and analysis of publicly available data sets suggested that the resistance gene pool is dominated by a relatively small subset of genes, with a very high positive predictive value for phenotype. In this study, 152 Escherichia coli and 115 Klebsiella pneumoniae consecutive isolates with a cefotaxime, ceftriaxone and/or ceftazidime minimum inhibitory concentration (MIC) of ≥ 2 μg/mL were collected from seven major hospitals in Sydney (Australia) in 2008-2009. Nearly all of those with a MIC in excess of European Committee on Antimicrobial Susceptibility Testing (EUCAST) resistance breakpoints contained one or more representatives of only seven gene types capable of explaining this phenotype, and this included 96% of those with a MIC ≥ 2 μg/mL to any one of these drugs. Similarly, 97% of associated gentamicin-non-susceptibility (MIC ≥ 8 μg/mL) could be explained by three gene types. In a country like Australia, with a background prevalence of resistance to third-generation cephalosporins of 5-10%, this equates to a negative predictive value of >99.5% for non-susceptibility and is therefore suitable for diagnostic application. This is an important proof-of-principle that should be tested in other geographic locations.

IncA/C Plasmid Carrying blaNDM-1, blaCMY-16, and fosA3 in a Salmonella enterica Serovar Corvallis Strain Isolated from a Migratory Wild Bird in Germany

ABSTRACT A Salmonella enterica serovar Corvallis strain was isolated from a wild bird in Germany. This strain carried the IncA/C2 pRH-1238 plasmid. Complete sequencing of the plasmid was performed, identifying the blaNDM-1, blaCMY-16, fosA3, sul1, sul2, strA, strB, aac(6′)-Ib, aadA5, aphA6, tetA(A), mphA, floR, dfrA7, and merA genes, which confer clinically relevant resistance to most of the antimicrobial classes, including β-lactams with carbapenems, fosfomycin, aminoglycosides, co-trimoxazole, tetracyclines, and macrolides. The strain likely originated from the Asiatic region and was transferred to Germany through the Milvus migrans migratory route.

ISEcp1-Mediated Transposition and Homologous Recombination Can Explain the Context of blaCTX-M-62 Linked to qnrB2

ABSTRACT bla CTX-M-62, a C508T variant of blaCTX-M-3b, was transferred from Klebsiella pneumoniae JIE137 on a conjugative plasmid together with a class 1 integron containing the dfrA12-gcuF-aadA2 cassette array, ISCR1, and qnrB2. blaCTX-M-62 lies between intact and rearranged copies of ISEcp1 in a configuration that can be explained by a combination of transposition and homologous recombination and which also illustrates the ability of ISEcp1 to mobilize an adjacent gene as part of transposition units of different sizes.

A blaVEB-1 Variant, blaVEB-6, Associated with Repeated Elements in a Complex Genetic Structure

ABSTRACT bla VEB-6 was found on the Proteus mirabilis chromosome in a context similar to those of blaVEB-1a and blaVEB-1b, in a truncated gene cassette flanked by 135-bp elements and duplications of the 3′-conserved segment of class 1 integrons. A linked aacA4-aadB-dfrA1-orfC cassette array includes components of Tn1331, illustrating the complex mosaicism of multiresistance regions.

RmtC 16S rRNA Methyltransferase in Australia

Proteus mirabilis isolate JIE273 was recovered in March 2007 from the urine of an inpatient at Blacktown Hospital, Sydney. In vitro susceptibility tests (Phoenix NMIC/ID-101 panel; BD, Franklin Lakes, NJ) indicated that JIE273 was susceptible to imipenem (MIC, ≤2 μg/ml), cefoxitin (MIC, ≤4 μg/

Prediction of major antibiotic resistance in Escherichia coli and Klebsiella pneumoniae in Singapore, USA and China using a limited set of gene targets

Antibiotic resistance in Gram-negative bacteria, especially Enterobacteriaceae, can be conferred by a large number of different acquired resistance genes, although it appears that relatively few dominate. A previous survey of Escherichia coli and Klebsiella pneumoniae isolates from Sydney, Australia, revealed that a limited set of genes could reliably predict resistance to third-generation cephalosporins (3GCs) and aminoglycosides. Here we tested E. coli and K. pneumoniae isolates with a cefotaxime, ceftriaxone and/or ceftazidime minimum inhibitory concentration of ≥ 2 μg/mL from China and Singapore, with significantly higher resistance rates than Australia, as well as the USA. Few targets were needed to predict non-susceptibility to 3GCs (95/95; 100%) and gentamicin (47/51; 92%). The gene types detected here are consistent with previous surveys in similar countries with similar resistance rates, where the majority of 3GC resistance can be explained by blaCTX-M genes. This study identified a limited set of genes capable of predicting resistance to 3GC and aminoglycoside antibiotics and implies a restriction in the global resistance gene pool that can be exploited for diagnostic purposes.

Different IncI1 plasmids from Escherichia coli carry ISEcp1-blaCTX-M-15 associated with different Tn2-derived elements

The bla(CTX-M-15) gene, encoding the globally dominant CTX-M-15 extended-spectrum β-lactamase, has generally been found in a 2.971-kb ISEcp1-bla(CTX-M-15)-orf477Δ transposition unit, with ISEcp1 providing a promoter. In available IncF plasmid sequences from Escherichia coli, this transposition unit interrupts a truncated copy of transposon Tn2 that lies within larger multiresistance regions. In E. coli, bla(CTX-M-15) is also commonly associated with IncI1 plasmids and here three such plasmids from E. coli clinical isolates from western Sydney 2006-2007 have been sequenced. The plasmid backbones are organised similarly to those of other IncI1 plasmids, but have insertions and/or deletions and sequence differences. Each plasmid also has a different insertion carrying bla(CTX-M-15). pJIE113 (IncI1 sequence type ST31) is almost identical to plasmids isolated from the 2011 E. coli O104:H4 outbreak in Europe, where the typical bla(CTX-M-15) transposition unit interrupts a complete Tn2 inserted directly in the plasmid backbone. In the novel plasmid pJIE139 (ST88), ISEcp1-blaC(TX-M-15)-orf477Δ lies within a Tn2/3 hybrid transposon. Homologous recombination could explain movement of ISEcp1-bla(CTX-M-15)-orf477Δ between copies of Tn2 on IncF and IncI1 plasmids and generation of the Tn2/3 hybrid. pJIE174 (ST37) is almost identical to pESBL-12 from the Netherlands and in these plasmids bla(CTX-M-15) is flanked by two copies of IS26 that truncate the transposition unit within a larger region bounded by the ends of Tn2. bla(CTX-M-15) and the associated ISEcp1-derived promoter may be able to move from this structure by the actions of IS26, independently of both ISEcp1 and Tn2.