Of the 181 unduplicated Escherichia coli strains isolated in nine different hospitals in three Portuguese regions, 119 were extended-spectrum ␤-lactamase (ESBL)-CTX-M producers and were selected for phenotype and genotype characterization. CTX-M producer strains were prevalent among community-acquired infections (56%), urinary tract infections (76%), and patients >60 years old (76%). In MIC tests, all strains were resistant to cefotaxime, 92% were resistant to ceftazidime, 93% were resistant to quinolones, 89% were resistant to aminoglycoside, and 26% were resistant to trimethoprim-sulfamethoxazole; all strains were sensitive to carbapenems, and 92% of the strains had a multidrug resistance phenotype. Molecular methods identified 109 isolates harboring a bla CTX-M-15 gene, 1 harboring the bla CTX-M-32 gene (first identification in the country), and 9 harboring the bla CTX-M-14 gene. All isolates presented the ISEcp1 element upstream from the bla CTX-M genes; one presented the IS903 element (downstream of bla CTX-M-14 gene), and none had the IS26 element; 85% carried bla TEM-1B , and 84% also carried a bla OXA-30 . Genetic relatedness analysis based on pulsed-field gel electrophoresis defined five clusters and indicated that 76% of all isolates (from cluster IV) corresponded to a single epidemic strain. Of the 47 strains from one hospital, 41 belonged to cluster IV and were disseminated in three main wards. CTX-M-producing E. coli strains are currently a problem in Portugal, with CTX-M-15 particularly common. This study suggests that the horizontal transfer of bla CTX-M genes, mediated by plasmids and/or mobile elements, contributes to the dissemination of CTX-M enzymes to community and hospital environments. The use of extended-spectrum cephalosporins, quinolones, and aminoglycosides is compromised, leaving carbapenems as the therapeutic option for severe infections caused by ESBL producers.

With an increasing prevalence during the past decades, atopic dermatitis has become a global health issue. A literature search following a targeted approach was undertaken to perform this non-systematic review, which intends to provide an overview of the epidemiology, pathophysiology, clinical features, comorbidities, and current therapies for the treatment of atopic dermatitis. In sum, this is a heterogeneous skin disorder associated with variable morphology, distribution, and disease course. Although not completely understood, its pathogenesis is complex and seems to result from a combination of genetic and environmental factors that induce skin barrier dysfunction, cutaneous and systemic immune dysregulation, skin microbiota dysbiosis, and a strong genetic influence. Diagnosis is based on specific criteria that consider patient and family history and clinical manifestations. Overall disease severity must be determined by evaluating both objective signs and subjective symptoms. Therapeutic goals require a multistep approach, focusing on reducing pruritus and establishing disease control. Patients should be advised on basic skin care and avoidance of triggers. Topical anti-inflammatory agents should be considered in disease flares or chronic/recurrent lesions. In case of inadequate response, phototherapy, systemic immunosuppressants and, more recently, dupilumab, should be added. Nevertheless, the treatment of moderate-to-severe atopic dermatitis remains challenging and novel, efficacious, safe and targeted treatments are urgently needed. In conclusion, although the last few years have seen important improvement in the understanding of the disease, future research in atopic dermatitis will continue exploring gene-environment interactions and how it affects pathophysiology, disease severity, and treatment outcomes.

cAmong the 2,105 Enterobacteriaceae tested in a survey done in Portugal, 165 were nonsusceptible to carbapenems, from which 35 (26 Klebsiella pneumoniae, 3 Escherichia coli, 2 Enterobacter aerogenes, and 3 Enterobacter cloacae isolates and 1 Klebsiella oxytoca isolate) were confirmed to be carbapenemase producers by the presence of 30 Tn4401d-bla KPC-3 , 4 intI3-bla GES-5 , and one intI1-bla VIM-2 gene, alone or in combination with other bla genes. The dissemination of bla KPC-3 gene carried by an IncF plasmid suggests lateral gene transfer as a major mechanism of dissemination.

The spread of antibiotic resistant bacteria throughout the food chain constitutes a public health concern. To understand the contribution of fresh produce in shaping antibiotic resistance bacteria and integron prevalence in the food chain, 333 antibiotic resistance Gram negative isolates were collected from organic and conventionally produced fruits (pears, apples, and strawberries) and vegetables (lettuces, tomatoes, and carrots). Although low levels of resistance have been detected, the bacterial genera identified in the assessed fresh produce are often described not only as environmental, but mostly as commensals and opportunistic pathogens. The genomic characterization of integron-harboring isolates revealed a high number of mobile genetic elements and clinically relevant antibiotic resistance genes, of which we highlight the presence of as mcr-1, qnrA1, blaGES−11, mphA, and oqxAB. The study of class 1 (n = 8), class 2 (n = 3) and class 3 (n = 1) integrons, harbored by species such as Morganella morganii, Escherichia coli, Klebsiella pneumoniae, led to the identification of different integron promoters (PcW, PcH1, PcS, and PcWTNG−10) and cassette arrays (containing drfA, aadA, cmlA, estX, sat, and blaGES). In fact, the diverse integron backbones were associated with transposable elements (e.g., Tn402, Tn7, ISCR1, Tn2*, IS26, IS1326, and IS3) that conferred greater mobility. This is also the first appearance of In1258, In1259, and In3-13, which should be monitored to prevent their establishment as successfully dispersed mobile resistance integrons. These results underscore the growing concern about the dissemination of acquired resistance genes by mobile elements in the food chain.

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The new ␤-lactamase SHV-72 was isolated from clinical Klebsiella pneumoniae INSRA1229, which exhibited the unusual association of resistance to the amoxicillin-clavulanic acid combination (MIC, 64 g/ml) and susceptibility to cephalosporins, aztreonam, and imipenem. SHV-72 (pI 7.6) harbored the three amino acid substitutions Ile8Phe, Ala146Val, and Lys234Arg. SHV-72 had high catalytic efficiency against penicillins (k cat /K m , 35 to 287 M ؊1 ⅐ s ؊1 ) and no activity against oxyimino ␤-lactams. The concentration of clavulanic acid necessary to inhibit the enzyme activity by 50% was 10-fold higher for SHV-72 than for SHV-1. Moleculardynamics simulation suggested that the Lys234Arg substitution in SHV-72 stabilized an atypical conformation of the Ser130 side chain, which moved the O␥ atom of Ser130 around 3.5 Å away from the key O␥ atom of the reactive serine (Ser70). This movement may therefore decrease the susceptibility to clavulanic acid by preventing cross-linking between Ser130 and Ser70.The most common resistance mechanism in bacteria against ␤-lactam antibiotics is the production of ␤-lactamases (EC 3.5.2.6), which hydrolyze and inactivate ␤-lactams. ␤-Lactamases are divided into four major classes (A to D) on the basis of their primary sequence (1). While class B is composed of metalloenzymes that necessitate the presence of zinc cations for activity, classes A, C, and D are serine hydrolases (1, 7). Class A enzymes comprise several enzyme families, including the clinically relevant enzymes TEM and SHV (26).TEM and SHV enzymes initially had preferential activity against penicillins, as in the case of enzymes SHV-1 and TEM-1. Oxyimino ␤-lactams that are resistant to their hydrolytic activity and ␤-lactam inhibitors, such as clavulanic acid and tazobactam, have been developed to get around the activities of these enzymes (6,9,26). Nevertheless, the presence of point mutations in TEM and SHV enzymes has expanded the substrate spectrum to include oxyimino ␤-lactams and/or has conferred resistance to the inhibitors (3, 6, 9, 26).More than 28 inhibitor-resistant TEM enzymes have been detected. They harbor amino acid substitutions at positions 69, 130, 165, 182, 244, 275, and/or 276 that confer resistance to inhibitors (9, 15). Only three natural inhibitor-resistant SHVs (IRS) have been reported (http://www.lahey.org/studies). It has been proposed that substitutions at positions 69, 130, and 187 are involved in their resistance to inhibitors (10,14,31). IRS enzymes have also been constructed in vitro by site saturation mutagenesis at position 244 (37).In this study, we performed a phenotypic, molecular, and biochemical characterization of the new IRS-type ␤-lactamase SHV-72 from a clinical K. pneumoniae strain and investigated by molecular-dynamics simulations (MDSs) the role of the Lys234Arg substitution in its resistance to clavulanic acid. MATERIALS AND METHODSBacterial strains and plasmid. Klebsiella pneumoniae INSRA1229 was isolated from sputum of an 80-year-old male in an internal medicine service of a ge...

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