Characterization of Pseudomonas aeruginosa isolates: Occurrence rates, antimicrobial susceptibility patterns, and molecular typing in the global SENTRY Antimicrobial Surveillance Program, 1997-1999

During 1997–1999, a total of 70,067 isolates (6631 Pseudomonas aeruginosa isolates) were analyzed in the SENTRY program by geographic region and body site of infection. The respiratory tract was the most common source of P. aeruginosa. P. aeruginosa isolation rates increased during the study interval. Europe was the only region to show a significant decline in β-lactam and aminoglycoside susceptibility rates. There was a reduction in the rates of susceptibility of Canadian isolates to imipenem and of Latin American isolates to meropenem. A total of 218 multidrug-resistant P. aeruginosa isolates (MDR-PSA; resistant to piperacillin, ceftazidime, imipenem, and gentamicin) were observed; MDR-PSA occurrence rates (percentages of all isolates) ranged from 8.2% (Latin America) to 0.9% (Canada). No antimicrobial inhibited >50% of MDR-PSA strains. Molecular characterization of selected, generally resistant strains was performed. Isolates showing unique ribogroups were found in Europe, Latin America, and the United States, but clonal spread was documented in several medical centers.A. C. Gales, R. N. Jones, J. Turnidge, R. Rennie, and R. Rampha

Laboratory variants GESG170L, GESG170K, and GESG170H increase carbapenem hydrolysis and confer resistance to clavulanic acid

The Guiana extended-spectrum (GES) b-lactamase GESG170H, GESG170L, and GESG170K mutants showed kcat, Km, and kcat/Km values very dissimilar to those of GES-1 and GES-5. The enhancement of the hydrolytic activity against carbapenems is potentially due to a shift of the substrate in the active site that provides better positioning of the deacylating water molecule caused by the presence of the imidazole ring of H170 and of the long side chain of K170 and L170

A Two Amino Acid Duplication, L167E168, in the X-Loop Drastically Decreases Carbapenemase Activity of KPC-53, a Natural Class A β-Lactamase

KPC-53 enzyme is a natural KPC variant which showed a duplication of L167E168 residues in the X-loop structure. The blaKPC-53 gene was cloned both into pBC-SK and pET-24a vectors, and the recombinant plasmids were transferred by transformation in Escherichia coli competent cells to evaluate the antimicrobial susceptibility and to produce the enzyme. Compared to KPC-3, the KPC-53 was less stable and showed a dramatic reduction of kcat and kcat/Km versus several β-lactams, in particular carbapenems. Indeed, a 2,000-fold reduction was observed in the kcat values of KPC-53 for imipenem and meropenem. Concerning inhibitors, KPC-53 was susceptible to tazobactam and clavulanic acid but maintained resistance to avibactam. The molecular modeling indicates that the L167E168 duplication in KPC-53 modifies the interactions between residues involved in the catalytic pocket, changing the flexibility of the X-loop, which is directly coupled with the catalytic properties of the KPC enzymes

Culture-dependent and sequencing methods revealed the absence of a bacterial community residing in the urine of healthy cats

A growing number of studies suggest that the lower urinary tract of humans and dogs can harbor a urinary microbiota. Nevertheless, a certain concern has developed that the microbiota reported could be due to unaccounted contamination, especially in low-biomass samples. The aim of this study was to investigate the bacterial community which populates the urine of healthy cats using two approaches: a culture-dependent approach which consisted of the expanded quantitative urine culture (EQUC) techniques capable of identifying live bacteria not growing in standard urine cultures, and a culture-independent approach which consisted of 16S ribosomal RNA next generation sequencing (16S rRNA NGS) capable of identifying bacterial DNA and exploring microbial diversity with high resolution. To avoid confounding factors of possible bacterial contamination, the urine was sampled using ultrasound-guided cystocentesis, and several sample controls and negative controls were analyzed. The urine sampled from the 10 cats included in the study showed no bacterial growth in the EQUC procedure. Although several reads were successfully originated using 16S rRNA NGS, a comparable pattern was observed between urine samples and the negative control, and no taxa were statistically accepted as non-contaminant. Taken together, the results obtained allowed stating that no viable bacteria were present in the urine of healthy cats without lower urinary tract disease and urinary tract infections, and that the bacterial DNA detected was of contaminant origin

Potent inhibitory activity of taniborbactam towards NDM-1 and NDM-1Q119X mutants, and in vitro activity of cefepime/taniborbactam against MBLs producing Enterobacterales

Objective: This study aimed to investigate the in vitro activity of taniborbactam (VNRX-5133), a novel broad-spectrum bicyclic boronate, against NDM-1 and Q119E, Q119K, Q119C, Q119F, Q119V, and Q119Y NDM-1 variants, which showed an increased activity towards some β-lactams, including cefepime. Methods: Inhibition kinetic assays were spectrophotometrically performed using cefepime (50 μM) as the reporter substrate and 80 nM of each enzyme. Taniborbactam behaves as a competitive inhibitor towards NDM-1 and NDM-1 Q119 variants with lower Ki values (range 3–16 nM). The phenotypic profile was assessed in both Enterobacterales clinical isolates and engineered Escherichia coli BL21(DE3) strains by conventional broth microdilution procedures according to the Clinical and Laboratory Standards Institute (CLSI). Results: Taniborbactam at a fixed concentration of 4 mg/L was able to restore activity of cefepime in 24 of 26 Enterobacterales clinical isolates harbouring metallo-β-lactamases with MIC50/MIC90 values of 14 mg/L. Cefepime MICs were drastically reduced in all clinical isolates and in NDM-1 and Q119X producing Escherichia coli BL21(DE3). Taniborbactam was unable to restore susceptibility to cefepime in two IMP variants producing clinical isolates. Conclusion: The inhibition level of NDM enzymes provided by taniborbactam protects the antibacterial activity of cefepime from this important metallo-β-lactamase

<i>In Vitro</i>Activity of Cefodizime (HR-221) in Combination with β-Lactamase Inhibitors

Cefodizime (formerly HR221) was tested either for in vitro microbiological activity or for its stability to β-lactamases in the presence of two β- lactamase inhibitors (clavulanic acid, tazobactam). Cefodizime was a poor substrate of class C enzymes but hyperproducer strains were generally resistant with or without a β-lactamase inhibitor used in combination. On the contrary, class A enzymes were able to hydrolyze cefodizime. However, strains expressing class A β-lactamase were susceptible to cefodizime in combination with clavulanic acid