Coping with antibiotic resistance: contributions from genomics

Antibiotic resistance is a public health issue of global dimensions with a significant impact on morbidity, mortality and healthcare-associated costs. The problem has recently been worsened by the steady increase in multiresistant strains and by the restriction of antibiotic discovery and development programs. Recent advances in the field of bacterial genomics will further current knowledge on antibiotic resistance and help to tackle the problem. Bacterial genomics and transcriptomics can inform our understanding of resistance mechanisms, and comparative genomic analysis can provide relevant information on the evolution of resistant strains and on resistance genes and cognate genetic elements. Moreover, bacterial genomics, including functional and structural genomics, is also proving to be instrumental in the identification of new targets, which is a crucial step in new antibiotic discovery programs

Characterization of a Multiresistance Plasmid Carrying the optrA and cfr Resistance Genes From an Enterococcus faecium Clinical Isolate

open13noEnterococcus faecium E35048, a bloodstream isolate from Italy, was the first strain where the oxazolidinone resistance gene optrA was detected outside China. The strain was also positive for the oxazolidinone resistance gene cfr. WGS analysis revealed that the two genes were linked (23.1 kb apart), being co-carried by a 41,816-bp plasmid that was named pE35048-oc. This plasmid also carried the macrolide resistance gene erm(B) and a backbone related to that of the well-known Enterococcus faecalis plasmid pRE25 (identity 96%, coverage 65%). The optrA gene context was original, optrA being part of a composite transposon, named Tn6628, which was integrated into the gene encoding for the ζ toxin protein (orf19 of pRE25). The cfr gene was flanked by two ISEnfa5 insertion sequences and the element was inserted into an lnu(E) gene. Both optrA and cfr contexts were excisable. pE35048-oc could not be transferred to enterococcal recipients by conjugation or transformation. A plasmid-cured derivative of E. faecium E35048 was obtained following growth at 42°C, and the complete loss of pE35048-oc was confirmed by WGS. pE35048-oc exhibited some similarity but also notable differences from pEF12-0805, a recently described enterococcal plasmid from human E. faecium also co-carrying optrA and cfr; conversely it was completely unrelated to other optrA- and cfr-carrying plasmids from Staphylococcus sciuri. The optrA-cfr linkage is a matter of concern since it could herald the possibility of a co-spread of the two genes, both involved in resistance to last resort agents such as the oxazolidinones.openMorroni, Gianluca; Brenciani, Andrea; Antonelli, Alberto; Maria D’Andrea, Marco; Di Pilato, Vincenzo; Fioriti, Simona; Mingoia, Marina; Vignaroli, Carla; Cirioni, Oscar; Biavasco, Francesca; Varaldo, Pietro E.; Rossolini, Gian Maria; Giovanetti, EleonoraMorroni, Gianluca; Brenciani, Andrea; Antonelli, Alberto; Maria D’Andrea, Marco; Di Pilato, Vincenzo; Fioriti, Simona; Mingoia, Marina; Vignaroli, Carla; Cirioni, Oscar; Biavasco, Francesca; Varaldo, Pietro E.; Rossolini, Gian Maria; Giovanetti, Eleonor

Structure of the capsular polysaccharide of the KPC-2-producing Klebsiella pneumoniae strain KK207-2 and assignment of the glycosyltransferases functions

Klebsiella pneumoniae strain KK207-2 was isolated in 2010 froma bloodstreaminfection of an inpatient at an Italian hospital. It was previously found to produce the KPC-2 carbapenemase and to belong to clade 1 of sequence type 258. Genotyping of the conserved wzi and wzc genes from strain KK207-2 yielded contrasting results: the wzc-based method assigned the cps207\u20132 to a new K-type, while the wzi-based method assigned it to the known K41 K-type. In order to resolve this contradiction, the capsular polysaccharide of K. pneumoniae KK207-2 was purified and its structure determined by using GLC-MS of appropriate carbohydrate derivatives, ESI-MS of both partial hydrolysis and Smith degradation derived oligosaccharides, andNMR spectroscopy of oligosaccharides, and the lithium degraded, native and de-O-acetylated polysaccharide. All the collected data demonstrated the following repeating unit for the K. pneumoniae KK207-2 capsular polysaccharide: OAc 6 [3)-\u3b2-D-Gal-(1-4)-\u3b2-D-Glc-(1-]n 4 I 1 \u3b2-D-Glcp-(1-6)-\u3b1-D-Glcp-(1-4)-\u3b2-D-GlcpA-(1-6)-\u3b1-D-Glcp The polysaccharide contains about 0.60 acetyl groups per repeating unit on C6 of the Gal residue. The reactions catalysed by each glycosyltransferase in the cpsKK207-2 gene cluster were assigned on the basis of structural homology with other Klebsiella K antigens

Pseudomonas aeruginosa bloodstream infections: risk factors and treatment outcome related to expression of the PER-1 extended-spectrum beta-lactamase

BACKGROUND: Bloodstream infection (BSI) due to Pseudomonas aeruginosa (Pa) has relevant clinical impact especially in relation to drug resistance determinants. The PER-1 extended-spectrum beta-lactamase (ESBL) is a common enzyme conferring high-level resistance to anti-pseudomonal cephalosporins. Risk factors and treatment outcome of BSI episodes caused by PER-1-positive Pa (PER-1-Pa) strains were compared to those caused by ESBL-negative Pa isolates (ESBL-N-Pa). METHODS: Twenty-six BSI cases due to ceftazidime-resistant Pa strains have been investigated. MIC values of anti-pseudomonal drugs were determined by the Etest method (AB Biodisk, Solna, Sweden). The double-disk synergy test was used to detect ESBL production. PCR amplification and DNA sequencing were used to characterize ESBL types. Clinical records of BSI-patients were examined retrospectively. Demographic data, underlying diseases (McCabe-Jackson classification and Charlson weighted index), risk factors, antimicrobial therapy, and treatment outcome were evaluated in cases due to ESBL-positive and cases due to ESBL-N-Pa isolates. Unpaired Student's t-test, Mann-Whitney U-test, Fisher's exact test and the χ(2 )test were used for statistical analysis. RESULTS: Nine Pa isolates expressed the PER-1 ESBL; the remaining 17 isolates did not produce ESBLs. Severe sepsis (P = 0.03), bladder and intravascular catheters (both P = 0.01), immunosuppressive therapy (P = 0.04), and mechanical ventilation (P = 0.03) were significantly associated with BSI due to PER-1-Pa. Empirical treatment (P = 0.02) and treatment after ID/AST (P < 0.01) were rarely adequate in PER-1-Pa cases. With regard to treatment outcome, 77.8% BSI cases due to PER-1-Pa vs. 28.6% cases due to ESBL-N-Pa isolates failed to respond (P < 0.03). All cases due to PER-1-Pa that were treated with carbapenems (alone or in combination with amikacin) failed to respond. In contrast, 7/8 cases due to ESBL-N-Pa given carbapenems were responders. CONCLUSION: Therapeutic failure and increased hospital costs are associated with BSI episodes caused by PER-1-Pa strains. Thus, recognition and prompt reporting of ESBL-production appears a critical factor for the management of patients with serious P. aeruginosa infections

Plasmid-mediated quinolone resistance determinant qnrB19 in non-typhoidal Salmonella enterica strains isolated in Venezuela

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Tracking Acquired Antibiotic Resistance in Commensal Bacteria of Galápagos Land Iguanas: No Man, No Resistance

BACKGROUND: Antibiotic resistance, evolving and spreading among bacterial pathogens, poses a serious threat to human health. Antibiotic use for clinical, veterinary and agricultural practices provides the major selective pressure for emergence and persistence of acquired resistance determinants. However, resistance has also been found in the absence of antibiotic exposure, such as in bacteria from wildlife, raising a question about the mechanisms of emergence and persistence of resistant strains under similar conditions, and the implications for resistance control strategies. Since previous studies yielded some contrasting results, possibly due to differences in the ecological landscapes of the studied wildlife, we further investigated this issue in wildlife from a remote setting of the Galapagos archipelago. METHODOLOGY/PRINCIPAL FINDINGS: Screening for acquired antibiotic resistance was carried out in commensal enterobacteria from Conolophus pallidus, the terrestrial iguana of Isla Santa Fe, where: i) the abiotic conditions ensure to microbes good survival possibilities in the environment; ii) the animal density and their habits favour microbial circulation between individuals; and iii) there is no history of antibiotic exposure and the impact of humans and introduced animal species is minimal except for restricted areas. Results revealed that acquired antibiotic resistance traits were exceedingly rare among bacteria, occurring only as non-dominant strains from an area of minor human impact. CONCLUSIONS/SIGNIFICANCE: Where both the exposure to antibiotics and the anthropic pressure are minimal, acquired antibiotic resistance traits are not normally found in bacteria from wildlife, even if the ecological landscape is highly favourable to bacterial circulation among animals. Monitoring antibiotic resistance in wildlife from remote areas could also be a useful tool to evaluate the impact of anthropic pressure

Key considerations on the potential impacts of the COVID-19 pandemic on antimicrobial resistance research and surveillance

Antibiotic use in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patients during the COVID-19 pandemic has exceeded the incidence of bacterial coinfections and secondary infections, suggesting inappropriate and excessive prescribing. Even in settings with established antimicrobial stewardship (AMS) programmes, there were weaknesses exposed regarding appropriate antibiotic use in the context of the pandemic. Moreover, antimicrobial resistance (AMR) surveillance and AMS have been deprioritised with diversion of health system resources to the pandemic response. This experience highlights deficiencies in AMR containment and mitigation strategies that require urgent attention from clinical and scientific communities. These include the need to implement diagnostic stewardship to assess the global incidence of coinfections and secondary infections in COVID-19 patients, including those by multidrug-resistant pathogens, to identify patients most likely to benefit from antibiotic treatment and identify when antibiotics can be safely withheld, de-escalated or discontinued. Long-term global surveillance of clinical and societal antibiotic use and resistance trends is required to prepare for subsequent changes in AMR epidemiology, while ensuring uninterrupted supply chains and preventing drug shortages and stock outs. These interventions present implementation challenges in resource-constrained settings, making a case for implementation research on AMR. Knowledge and support for these practices will come from internationally coordinated, targeted research on AMR, supporting the preparation for future challenges from emerging AMR in the context of the current COVID-19 pandemic or future pandemics