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In previous studies, cefiderocol resistance in I E. coli i has been mainly attributed to the increased copy number and expression of the I bla i SB NDM sb -like gene.[2] Thus, we first investigated the gene copy number of I bla i SB NDM sb using the average coverage of reads generated from WGS data normalized to the average coverage of several intrinsic chromosomal genes used as reference ( I bla i SB EC sb -like, I gyrB i , I tonB i , I rpoB i ). Cefiderocol is a novel siderophore-conjugated cephalosporin that is very promising for the treatment of infections caused by MDR bacteria. [Extracted from the article]

Easy and robust antimicrobial susceptibility testing (AST) methods are essential in clinical bacteriology laboratories (CBL) in low-resource settings (LRS). We evaluated the Beckman Coulter MicroScan lyophilized broth microdilution panel designed to support Médecins Sans Frontières (MSF) CBL activity in difficult settings, in particular with the Mini-Lab. We evaluated the custom-designed MSF MicroScan Gram-pos microplate (MICPOS1) for Staphylococcus and Enterococcus species, MSF MicroScan Gram-neg microplate (MICNEG1) for Gram-negative bacilli, and MSF MicroScan Fastidious microplate (MICFAST1) for Streptococci and Haemophilus species using 387 isolates from routine CBLs from LRS against the reference methods. Results showed that, for all selected antibiotics on the three panels, the proportion of the category agreement was above 90% and the proportion of major and very major errors was below 3%, as per ISO standards. The use of the Prompt inoculation system was found to increase the MIC and the major error rate for some antibiotics when testing Staphylococci. The readability of the manufacturer's user manual was considered challenging for low-skilled staff. The inoculations and readings of the panels were estimated as easy to use. In conclusion, the three MSF MicroScan MIC panels performed well against clinical isolates from LRS and provided a convenient, robust, and standardized AST method for use in CBL in LRS. [ABSTRACT FROM AUTHOR]

Enterobacteriaceae are inhabitants of the intestinal flora J-J and are among the most common human pathogens, causing infections such as cystitis and pyelonephritis with fever, septicemia, pneumonia, peritonitis, meningitis, and [...]

Resistance of Klebsiella pneumoniae to carbapenems is mainly associated with acquired carbapenem-hydrolyzing β-lactamases (1). These β-lactamases can be metallo β-lactamases (IMP, VIM), expanded-spectrum oxacillinases (OXA-48), or Ambler class A enzymes [...]

Acinetobacter baumannii is a gram-negative organism that is increasingly recognized as a major pathogen causing nosocomial infections, including bacteremia and ventilator-associated pneumonia, particularly in patients admitted to intensive care units [...]

Ralstonia pickettii, an environmental bacterium that may also be responsible for human infections, produces two unrelated, inducible and chromosomally encoded oxacillinases, OXA-22 and OXA-60. In order to study the molecular basis of the induction process of these oxacillinase genes, the induction kinetics, the promoter/operator regions necessary for expression and induction, and the role of several ORFs located upstream and downstream of the [bla.su.b.OXA] genes were investigated. The [beta]-lactamase production reached a maximal level after 1 h induction, returned to its basal level within the following 3 h and was then again inducible. Using 5'RACE experiments, the promoter sequences of both oxacillinases were determined. These sequences showed weak promoter activities, which could, however, be increased approximately 200-fold by mutating the -35 promoter sequence. Deletion of the sequences located upstream of the promoter regions did not modify the basal [beta]-lactamase expression in R. pickettii, but resulted in the lack of induction. A minimum of 240 and 270 bp upstream of the transcription initiation sites was required for inducible expression of the [bla.sub.OXA-22] and [bla.sub.OXA-60] genes, respectively. Analysis of the genetic environment of both [bla.sub.OXA] genes revealed several ORFs that were inactivated by homologous recombination. Disruption of ORF-RP3, located 190 bp upstream of [bla.sub.OXA-60] and divergently transcribed, abolished induction of both [beta]-lactamases. ORF-RP3, which encoded a polypeptide of 532 aa with an estimated molecular mass of 58.7 kDa, displayed no obvious sequence homology with known regulatory proteins. Trans-complementation of ORF-RP3 restored the basal and inducible expression of both oxacillinase genes, indicating that the induction of both enzymes was related to the presence of ORF-RP3. In addition to the loss of induction, inactivation of the ORFRP3 in R. pickettii resulted in a complex pleiotropic phenotype, with increased lag phase and reduced survival after heat exposure, suggesting that ORF-RP3 might be a global regulator involved in unrelated regulatory pathways.

IS1999 and a point mutant derivative, IS1999.2, have been described inserted upstream of emerging antibiotic resistance genes [bla.sub.VEB-1] and [bla.sub.OXA-48]. 5' Rapid amplification of cDNA ends experiments revealed that expression of these [beta]-lactamase genes was driven by the outward-directed promoter, [P.sub.out], located in the IS1999 elements. These findings led us to study IS1999-mediated gene mobilization. Thus, the transposition properties of IS1999 and of IS1999-based composite transposons, made of two copies of IS1999 in different orientations, were investigated. IS1999 or IS1999-based composite transposons were capable of transposing onto the conjugative plasmid pOX38-Gen. Sequence analysis of the insertion sites revealed that IS1999 inserted preferentially into DNA targets containing the consensus sequence NGCNNNGCN. Transposition was more efficient when at least one left inverted repeat end was located at an outside end of the transposon. The transposition frequency of IS1999.2 was 10-fold lower than that of IS1999, and transposition frequencies of the putative natural transposon, Tn1999, were below detection limits of our transposition assay. This reduced transposition frequency of IS1999.2-based elements may result from a lower transcription of the transposase gene, as revealed by reverse transcription-PCR analyses.

VEB-1 extended-spectrum [beta]-lactamase-producing Acinetobacter baumannii was responsible for an outbreak in hospitals in France. A national alert was triggered in September 2003 when 4 hospitals reported clusters of A. baumannfi [...]

The integron-borne bl[a.sub.VEB-1] gene encodes an extended-spectrum [beta]-lactamase. This gene was associated mostly with IS1999 and rarely with an additional IS2000 element in Pseudomonas aeruginosa isolates from Thailand, whereas IS1999 was only very rarely associated with bl[a.sub.VEB.-1] in Enterobacteriaceae. Expression experiments and promoter study identified promoter sequences in IS1999 that increased the expression of VEB-1 in P.aeruginosa.

Summary: Early detection of expanded‐spectrum cephalosporin (ESC) resistance is essential not only for an effective therapy but also for the prompt implementation of infection control measures to prevent dissemination in the hospital. We have developed and validated a lateral flow immunoassay (LFIA), called LFIA‐CTX test, for the detection of ESC (cefotaxime) hydrolytic activity based on structural discrimination between the intact antibiotic and its hydrolysed product. A single bacterial colony was suspended in an extraction buffer containing cefotaxime. After a 30‐min incubation, the solution is loaded on the LFIA for reading within 10 min. A total of 348 well‐characterized Gram‐negative isolates were tested. Among them, the 38 isolates that did not express any cefotaxime‐hydrolysing β‐lactamase gave negative results. Of the 310 isolates expressing at least one cefotaxime‐hydrolysing β‐lactamase, all were tested positive, except three OXA‐48‐like producers, which were repeatedly detected negative. Therefore, the sensitivity was 99.1% and the specificity was 100%. The LFIA‐CTX test is efficient, fast, low‐cost and easy to implement in the workflow of a routine microbiology laboratory. This paper details the development and validation of a lateral flow immunochromatographic assay for the detection of hydrolysis activity of extended spectrum cephalosporins. This test, based on the use of monoclonal antibodies specific to intact antibiotics, takes advantage of the LFIA and the universality of biochemical tests (detection of enzymatic activity whatever the enzyme or its variants). Its validation on 348 bacterial strains indicates a sensitivity of 99.1% and a specificity of 100%, making it an efficient, rapid, low‐cost and easy to implement test in the workflow of a routine microbiology laboratory, without additional equipment or trained personnel. [ABSTRACT FROM AUTHOR]

Research reveals that the plasmid-located class 1 integron In53 is a truncated integron and carries eight resistance gene cassette and is associated with the beta-lactamase gene environment in Escherichia coli. The integron is localized to the Tn2000 composite transposon and is bounded by two insertion sequence elements. In53 .