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Preview: Journal of Antimicrobial Chemotherapy - current issue

Journal of Antimicrobial Chemotherapy Current Issue





Published: Mon, 19 Mar 2018 00:00:00 GMT

Last Build Date: Fri, 20 Apr 2018 06:49:22 GMT

 



Pharmacodynamics of inhaled amikacin (BAY 41-6551) studied in an in vitro pharmacokinetic model of infection

Mon, 19 Mar 2018 00:00:00 GMT

Abstract
Background
The pharmacodynamics of inhaled antimicrobials are poorly studied. Amikacin is being developed for inhalational therapy as BAY 41-6551.
Objectives
We employed an in vitro pharmacokinetic model to study the pharmacokinetics/pharmacodynamics of amikacin.
Methods
A dose-ranging design was used to establish fAUC/MIC and fCmax/MIC targets for static, −1 log drop and −2 log drop effects for strains of Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. We then modelled epithelial lining fluid (ELF) concentration associated with inhaled amikacin (400 mg every 12 h), over 5 days using mean human concentrations.
Results
The 24 h static effect fAUC/MIC targets and −1 log drop targets were 51.0 ± 26.7 and 71.6 ± 27.6 for all species of aerobic Gram-negative bacilli. fAUC/MIC targets for static effect, −1 log drop or −2 log drop were smaller than the 24 h values at 12 h and larger at 48 h. Emergence of resistance occurred maximally with E. coli in the fAUC/MIC range 12–60; K. pneumoniae 0–60 (48 h) and P. aeruginosa 12–80. When human ELF concentrations were modelled for strains with MIC ≤8 mg/L, there was rapid clearance and no regrowth. For strains with MIC ≥32 mg/L, there was initial clearance followed by regrowth. If MIC values were related to bacterial clearance then at least a static effect or −1 log drop in count would be expected for bacterial strains with MICs of ≤180 mg/L (static effect) or ≤148 mg/L (−1 log drop effect).
Conclusions
An fAUC/MIC amikacin target of 50–80 is appropriate for aerobic Gram-negative bacilli and mean ELF concentrations of BAY 41-6551 would produce a static to −1 log clearance with strains up to 128 mg/L.



CTX-M ESBL-producing Enterobacteriaceae: estimated prevalence in adults in England in 2014

Mon, 05 Mar 2018 00:00:00 GMT

Abstract
Background
ESBL-producing Enterobacteriaceae (ESBLPE) are increasing in prevalence worldwide and are more difficult to treat than non-ESBLPE. Their prevalence in the UK general population is unknown, as the only previous UK ESBLPE faecal colonization study involved patients with diarrhoea.
Objectives
To estimate the prevalence of CTX-M ESBLPE faecal colonization in the general adult population of England in 2014, and investigate risk factors.
Methods
A stratified random sample of 58 337 registered patients from 16 general practices within four areas of England were invited to participate by returning faeces specimens and self-completed questionnaires. Specimens were tested for ESBLPE and carbapenemase-producing Enterobacteriaceae (CPE).
Results
2430 individuals participated (4% of those invited). The estimated prevalence of colonization with CTX-M ESBLPE in England was 7.3% (95% CI 5.6%–9.4%) (Shropshire 774 participants, 4.9% colonization; Southampton City 740 participants, 9.2%; Newham 612 participants, 12.7%; Heart of Birmingham 234 individuals, 16.0%) and was particularly high in: those born in Afghanistan (10 participants, 60.0% colonization, 95% CI 29.7%–84.2%); those born on the Indian subcontinent (India, Pakistan, Bangladesh or Sri Lanka) (259 participants, 25.0% colonization, 95% CI 18.5%–32.9%); travellers to South Asia (India, Pakistan, Bangladesh, Sri Lanka or Nepal) in the last year (140 participants, 38.5% colonization, 95% CI 27.8%–50.5%); and healthcare domestics (8 participants, unweighted 37.5% colonization, 95% CI 8.5%–75.5%). Risk factors identified included: being born in the Indian subcontinent (aOR 5.4, 95% CI 3.0–9.7); travel to South Asia (aOR 2.9, 95% CI 1.8–4.8) or to Africa, China, South or Central America, South East or Pacific Asia or Afghanistan (aOR 2.6, 95% CI 1.7–4.1) in the last year; and working as a healthcare domestic (aOR 6.2, 95% CI 1.3–31). None of the 48 participants who took co-amoxiclav in the last year was colonized with CTX-M ESBLPE. blaCTX-M-15 accounted for 66% of CTX-M ESBLPE positives. 0.1% (two participants) were colonized with CPE.
Conclusions
CTX-M ESBLPE are established in the general population in England and prevalence is particularly high in people from certain countries of birth or with recent travel. We recommend that these findings be taken into account in guidance on the empirical management of patients presenting with a likely Enterobacteriaceae infection.



Effects of primary care antimicrobial stewardship outreach on antibiotic use by general practice staff: pragmatic randomized controlled trial of the TARGET antibiotics workshop

Mon, 05 Mar 2018 00:00:00 GMT

Abstract
Objectives
To determine whether local trainer-led TARGET antibiotic interactive workshops improve antibiotic dispensing in general practice.
Methods
Using a McNulty–Zelen-design randomized controlled trial within three regions of England, 152 general practices were stratified by clinical commissioning group, antibiotic dispensing rate and practice patient list size, then randomly allocated to intervention (offered TARGET workshop that incorporated a presentation, reflection on antibiotic data, promotion of patient and general practice (GP) staff resources, clinical scenarios and action planning, 73 practices) or control (usual practice, 79 practices). The primary outcome measure was total oral antibiotic items dispensed/1000 patients for the year after the workshop (or pseudo-workshop date for controls), adjusted for the previous year’s dispensing.
Results
Thirty-six (51%) intervention practices (166 GPs, 51 nurses and 101 other staff) accepted a TARGET workshop invitation. In the ITT analysis total antibiotic dispensing was 2.7% lower in intervention practices (95% CI −5.5% to 1%, P =0.06) compared with controls. Dispensing in intervention practices was 4.4% lower for amoxicillin/ampicillin (95% CI 0.6%–8%, P =0.02); 5.6% lower for trimethoprim (95% CI 0.7%–10.2%, P =0.03); and a non-significant 7.1% higher for nitrofurantoin (95% CI −0.03 to 15%, P =0.06). The Complier Average Causal Effect (CACE) analysis, which estimates impact in those that comply with assigned intervention, indicated 6.1% (95% CI 0.2%–11.7%, P =0.04) lower total antibiotic dispensing in intervention practices and 11% (95% CI 1.6%–19.5%, P =0.02) lower trimethoprim dispensing.
Conclusions
This study within usual service provision found that TARGET antibiotic workshops can help improve antibiotic use, and therefore should be considered as part of any national antimicrobial stewardship initiatives. Additional local facilitation will be needed to encourage all general practices to participate.



Pharmacodynamics and cellular accumulation of amphotericin B and miltefosine in Leishmania donovani-infected primary macrophages

Wed, 28 Feb 2018 00:00:00 GMT

Abstract
Objectives
We examined the in vitro pharmacodynamics and cellular accumulation of the standard anti-leishmanial drugs amphotericin B and miltefosine in intracellular Leishmania donovani amastigote–macrophage drug assays.
Methods
Primary mouse macrophages were infected with L. donovani amastigotes. In time–kill assays infected macrophages were exposed to at least six different concentrations of serially diluted drugs and the percentage of infected macrophages was determined after 6, 12, 24, 48, 72 and 120 h of exposure. Cellular drug accumulation was measured following exposure to highly effective drug concentrations for 1, 6, 24, 48 and 72 h. Data were analysed through a mathematical model, relating drug concentration to the percentage of infected cells over time. Host cell membrane damage was evaluated through measurement of lactate dehydrogenase release. The effect of varying the serum and albumin concentrations in medium on the cellular accumulation levels of miltefosine was measured.
Results
Amphotericin B was more potent than miltefosine (EC50 values of 0.65 and 1.26 μM, respectively) and displayed a wider therapeutic window in vitro. The kinetics of the cellular accumulation of amphotericin B was concentration- and formulation-dependent. At an extracellular concentration of 10 μM miltefosine maximum cellular drug levels preceded maximum anti-leishmanial kill. Miltefosine induced membrane damage in a concentration-, time- and serum-dependent manner. Its cellular accumulation levels increased with decreasing amounts of protein in assay medium.
Conclusions
We have developed a novel approach to investigate the cellular pharmacology of anti-leishmanial drugs that serves as a model for the characterization of new drug candidates.



Evaluation of three broth microdilution systems to determine colistin susceptibility of Gram-negative bacilli

Thu, 22 Feb 2018 00:00:00 GMT

Abstract
Background
The broth microdilution (BMD) method is currently the recommended technique to determine susceptibility to colistin.
Objectives
We evaluated the accuracy of three commercialized BMD panels [Sensititre (ThermoFisher Diagnostics), UMIC (Biocentric) and MicroScan (Beckman Coulter)] to determine colistin susceptibility.
Methods
A collection of 185 isolates of Gram-negative bacilli (133 colistin resistant and 52 colistin susceptible) was tested. Manual BMD according to EUCAST guidelines was used as the reference method, and EUCAST 2017 breakpoints were used for susceptibility categorization.
Results
The UMIC system gave the highest rate of very major errors (11.3%) compared with the Sensititre and MicroScan systems (3% and 0.8%, respectively). A high rate of major errors (26.9%) was found with the MicroScan system due to an overestimation of the MICs for the non-fermenting Gram-negative bacilli, whereas no major errors were found with the Sensititre and UMIC systems.
Conclusions
The UMIC system was easy to use, but failed to detect >10% of colistin-resistant isolates. The MicroScan system showed excellent results for enterobacterial isolates, but non-susceptible results for non-fermenters should be confirmed by another method and the range of MICs tested was narrow. The Sensititre system was the most reliable marketed BMD panel with a categorical agreement of 97.8%.



Selection of hyperproduction of AmpC and SME-1 in a carbapenem-resistant Serratia marcescens isolate during antibiotic therapy

Mon, 19 Feb 2018 00:00:00 GMT

Abstract
Objectives
Antibiotic selective pressure may result in changes to antimicrobial susceptibility throughout the course of infection, especially for organisms that harbour chromosomally encoded AmpC β-lactamases, notably Enterobacter spp., in which hyperexpression of ampC may be induced following treatment with cephalosporins. In this study, we document a case of bacteraemia caused by a blaSME-1-harbouring Serratia marcescens that subsequently developed resistance to expanded-spectrum cephalosporins, piperacillin/tazobactam and fluoroquinolones, over the course of several months of treatment with piperacillin/tazobactam and ciprofloxacin.
Methods
Susceptibility testing and WGS were performed on three S. marcescens isolates from the patient. β-Lactamase activity in the presence or absence of induction by imipenem was measured by nitrocefin hydrolysis assays. Expression of ampC and blaSME-1 under the same conditions was determined by real-time PCR.
Results
WGS demonstrated accumulation of missense and nonsense mutations in ampD associated with stable derepression of AmpC. Gene expression and β-lactamase activity of both AmpC and SME-1 were inducible in the initial susceptible isolate, but were constitutively high in the resistant isolate, in which total β-lactamase activity was increased by 128-fold.
Conclusions
Although development of such in vitro resistance due to selective pressure imposed by antibiotics is reportedly low in S. marcescens, our findings highlight the need to evaluate isolates on a regular basis during long-term antibiotic therapy.



A multicentre stewardship initiative to decrease excessive duration of antibiotic therapy for the treatment of community-acquired pneumonia

Fri, 16 Feb 2018 00:00:00 GMT

Abstract
Background
The increased emphasis on pneumonia-related performance measures and patient outcomes has led hospitals to implement multifaceted approaches to quickly identify patients with community-acquired pneumonia (CAP), start timely therapy and reduce readmission. However, there has been minimal focus on duration of therapy (DOT) and patients often receive prolonged antibiotic courses. The IDSA and American Thoracic Society (IDSA/ATS) CAP guidelines recommend 5 days of therapy for clinically stable patients that quickly defervesce and stewardship teams are well positioned to influence prescribing practices.
Objectives
Determine the impact of a prospective stewardship intervention on total antibiotic DOT and associated clinical outcomes in hospitalized patients with CAP.
Methods
This multicentre, quasi-experimental study evaluated three concurrent interventions over a 6 month period to promote appropriate DOT. All centres updated institutional CAP guidelines to promote IDSA/ATS-concordant DOT, provided education and conducted daily audit and feedback with intervention to provide patient-specific DOT recommendations.
Results
A total of 600 patients with CAP were included (307 in the historical control group and 293 in the stewardship intervention group). The stewardship intervention increased compliance with DOT recommendations (42% versus 5.6%, P <0.001) and reduced the median DOT per patient (6 versus 9 days, P <0.001). Clinical outcomes, including mortality, readmission with pneumonia, presentation to the emergency centre/clinic with pneumonia and incidence of Clostridium difficile infection within 30 days of discharge, were not different between groups.
Conclusions
This multicentre evaluation of a stewardship intervention in hospitalized CAP patients reduced the total antibiotic DOT and increased guideline-concordant DOT without adversely affecting patient outcomes.



Education and management of antimicrobials amongst nurses in Africa—a situation analysis: an Infection Control Africa Network (ICAN)/BSAC online survey

Thu, 15 Feb 2018 00:00:00 GMT

Abstract
Objectives
To assess the current involvement of nurses in the use and management of antimicrobials and their training in antimicrobial stewardship (AMS) across Africa.
Methods
After a pilot study, an online questionnaire (SurveyMonkey) in both French and English was circulated via the Infection Control Africa Network (ICAN) mailing list to both members and non-members in Africa. The study was conducted from 26 May to 19 August 2016. Data were summarized in proportions and bar charts; proportions were compared using the χ2 test. A multivariate logistic regression model was built to identify independent factors associated with the practice of AMS.
Results
While 96% of the 173 respondents were aware of the term ‘AMS’, 88.5% (146/165) undertook AMS tasks as part of their job; 91.9% (158/172) wanted to be more involved in AMS but 44.9% (71/158) reported there were barriers in doing so. AMS training was delivered to 36.7% (62/169) and 53.6% (90/168), respectively, during their undergraduate and postgraduate education. AMS training for healthcare workers in their institutions was reported by 50.3% (86/171), including training aimed at doctors (56.9%), pharmacists (76.7%), microbiologists (31.4%) and nurses (95.3%). However, 95.4% (164/172) of respondents asked for further education on AMS and the majority preferred AMS training to be part of the infection prevention curriculum (IPC) education. Three-quarters of institutions had an AMS initiative, but only ∼41% reported having seen a national AMS guideline.
Conclusions
For Africa, we recommend AMS education at undergraduate level, AMS policies at institution and national levels and incorporating AMS training into the IPC for nurses.



Primary resistance to integrase strand transfer inhibitors in patients infected with diverse HIV-1 subtypes in sub-Saharan Africa

Wed, 14 Feb 2018 00:00:00 GMT

Abstract
Objectives
To investigate the prevalence and patterns of major and accessory resistance mutations associated with integrase strand transfer inhibitors (INSTIs), across diverse HIV-1 subtypes in sub-Saharan Africa.
Methods
pol gene sequences were obtained using Illumina next-generation sequencing from 425 INSTI-naive HIV-infected adults from Kenya (21.2%), Nigeria (7.3%), South Africa (22.8%), Uganda (25.2%) and Zambia (23.5%). Drug resistance interpretation was based on the IAS 2017 mutation list and accessory mutations from Stanford HIVdb with resistance penalty scores of ≥10 to at least 1 INSTI. Resistance was further classified based on sensitivity thresholds of ≥20% (Sanger sequencing) and 1%–20% for low-frequency variants (next-generation sequencing).
Results
Of 425 genotypes, 48.7% were subtype C, 28.5% A, 10.1% D, 2.8% G and 9.9% were recombinants. Major INSTI resistance mutations were detected only at <20% threshold, at a prevalence of 2.4% (2.5% in subtype A, 2.4% C, 0% D, 8.3% G and 2.4% in recombinants) and included T66A/I (0.7%), E92G (0.5%), Y143C/S (0.7%), S147G (0.2%) and Q148R (0.5%). Accessory mutations occurred at a prevalence of 15.1% at the ≥20% threshold (23.1% in subtype A, 8.7% C, 11.6% D, 25% G and 23.8% in recombinants), and included L74I/M (10.4%), Q95K (0.5%), T97A (4%), E157Q (0.7%) and G163R/K (0.7%).
Conclusions
Major INSTI resistance mutations were rare and only occurred at low-level resistance detection thresholds. INSTI-based regimens are expected to be effective across the different major HIV-1 subtypes in the region.



Overexpression of Stenotrophomonas maltophilia major facilitator superfamily protein MfsA increases resistance to fluoroquinolone antibiotics

Wed, 14 Feb 2018 00:00:00 GMT

Abstract
Background
Stenotrophomonas maltophilia is an opportunistic human pathogen causing nosocomial infections worldwide. S. maltophilia infection is of particular concern due to its inherent resistance to currently used antibiotics. Proton motive force-driven transporters of the major facilitator superfamily frequently contribute to the efflux of substances, including antibiotics, across cell membranes.
Methods
An mfsA expression plasmid (pMfsA) was constructed and transferred into bacterial strains by electroporation. The antibiotic susceptibility levels of S. maltophilia strains were determined using standard methods.
Results and conclusions
S. maltophilia MfsA is an efflux pump associated with paraquat resistance. We show here that plasmid-mediated overexpression of mfsA in WT S. maltophilia K279a increased resistance not only to paraquat but also to second-generation fluoroquinolone antibiotics, i.e. ciprofloxacin, norfloxacin, levofloxacin and ofloxacin. Ciprofloxacin was used as a representative drug. Addition of the proton motive force inhibitor carbonyl cyanide-m-chlorophenylhydrazone increases susceptibility to ciprofloxacin. Taken together these results suggest that MsfA is a novel fluoroquinolone efflux pump of S. maltophilia. Moreover, heterologous expression of mfsA in other Gram-negative pathogenic bacteria conferred resistance to paraquat as well as to fluoroquinolones. Thus, if this determinant was horizontally transferred, it could cause the spread of fluoroquinolone resistance among bacterial species.









HIV-1 genotypic resistance testing using the Vela automated next-generation sequencing platform

Mon, 12 Feb 2018 00:00:00 GMT

Abstract
Objectives
To evaluate the diagnostic performance of the Vela next-generation sequencing (NGS) system in conjunction with the Sentosa SQ HIV Genotyping Assay for genotyping HIV-1.
Methods
Plasma RNA was extracted and templates prepared with the Sentosa SX instrument before sequencing the HIV-1 polymerase on the Sentosa SQ301 Sequencer (PGM IonTorrent). The Vela NGS System was compared with direct sequencing and the 454 GS-FLX (Roche) and MiSeq (Illumina) systems for genotypic resistance testing on clinical samples.
Results
The Vela NGS system detected majority resistance mutations in subtype B and CRF02-AG samples at 500 copies/mL and minority variants with a sensitivity of 5% at 100 000 copies/mL. The Vela NGS system and direct sequencing identified resistance mutations with 97% concordance in 46 clinical samples. Vela identified 1/20 of the 1%–5% mutations identified by 454, 5/12 of the 5%–20% mutations and 60/61 of the >20% mutations. Vela identified 3/14 of the 1%–5% mutations identified by MiSeq, 0/2 of the 5%–20% mutations and 47/47 of the >20% mutations. The resistance mutation quantifications by Vela and 454 were concordant (bias: 2.31%), as were those by Vela and MiSeq (bias: 1.06%).
Conclusions
The Vela NGS system provides automated nucleic acid extraction, PCR reagent distribution, library preparation and bioinformatics analysis. The analytical performance was very good when compared with direct sequencing, but was less sensitive than two other NGS platforms for detecting minority variants.



In vitro activity of zoliflodacin (ETX0914) against macrolide-resistant, fluoroquinolone-resistant and antimicrobial-susceptible Mycoplasma genitalium strains

Mon, 12 Feb 2018 00:00:00 GMT

Abstract
Background
Mycoplasma genitalium is estimated to be the second most common cause of bacterial sexually transmitted infection in Europe. It is of increasing public health concern due to the rapid development of resistance to different antimicrobial classes, including the preferred first- and second-line treatments azithromycin and moxifloxacin. Thus, new antimicrobial agents are urgently needed, especially for the treatment of MDR strains.
Methods
The in vitro activity of the new spiropyrimidinetrione zoliflodacin against 47 M. genitalium strains was assessed by growing M. genitalium in Vero cell culture and measuring growth by quantitative PCR. The collection included 34 moxifloxacin-susceptible (MIC <1 mg/L) and 13 moxifloxacin-resistant (MIC ≥1 mg/L) strains. Twenty-three of the strains were azithromycin resistant (MIC ≥16 mg/L) and 12 of these strains were MDR.
Results
Only one (2.1%) strain with substantially increased MIC (4 mg/L) and potential resistance to zoliflodacin was found. Zoliflodacin was overall more potent than moxifloxacin (P =0.009) and no cross-resistance was observed between the two drug classes of topoisomerase II inhibitors. Differences in the MICs of zoliflodacin and azithromycin were not statistically significant; however, 23 (48.9%) compared with potentially 1 (2.1%) of the strains were resistant to azithromycin and zoliflodacin, respectively.
Conclusions
Zoliflodacin is a promising candidate for the treatment of M. genitalium and it is important to further develop and evaluate this drug.



Gentamicin-intercalated smectite as a new therapeutic option for Helicobacter pylori eradication

Mon, 12 Feb 2018 00:00:00 GMT

Abstract
Objectives
Novel antibacterial strategies against Helicobacter pylori are needed because H. pylori strains are acquiring resistance to antibiotics. We evaluated the efficacy of gentamicin-intercalated smectite hybrid (S-GEN)-based treatment regimens in a murine model of H. pylori infection.
Methods
Two groups of 10 rats were administered either smectite or S-GEN to measure coverage of the gastric mucosa. To evaluate anti-H. pylori efficacy, mice were divided into eight groups of 10 mice each given different treatments, and H. pylori eradication was assessed by a Campylobacter-like organism (CLO) test and H. pylori PCR of the gastric mucosa, and H. pylori antigen and H. pylori PCR analysis of mouse faeces. The levels of proinflammatory cytokines were examined.
Results
S-GEN was retained in the gastric mucosal layer with a >60% distribution ratio for up to 1 h, and the S-GEN-based triple regimen decreased bacterial burden in vivo compared with that of untreated mice or mice treated with other regimens. The cure rates in the CLO test and H. pylori PCR from gastric mucosa were 70%, 60%, 80%, 50%, 60% and 60% in Groups III–VIII, respectively. Those for H. pylori PCR in the faeces of mice were 90% and 100% in Group III with standard therapy and Group V with triple therapy including S-GEN, respectively. S-GEN triple therapy also reduced the levels of proinflammatory cytokines.
Conclusions
These results suggest that S-GEN is a promising and effective therapeutic agent for the treatment of H. pylori infection.



Emergence and control of linezolid-resistant Staphylococcus epidermidis in an ICU of a German hospital

Fri, 09 Feb 2018 00:00:00 GMT

Abstract
Objectives
To investigate an outbreak of linezolid-resistant Staphylococcus epidermidis (LRSE) in an interdisciplinary ICU, linezolid consumption and infection control measures taken.
Methods
Routine surveillance of nosocomial infections revealed colonization and infection with LRSE affecting 14 patients during a 15 month period. LRSE isolates were analysed with respect to their clonal relatedness, antimicrobial susceptibility, the presence of cfr and/or mutations in the 23S rRNA, rplC, rplD and rplV genes. cfr plasmids were characterized by Illumina sequencing. Medical records were reviewed and antibiotic consumption was determined.
Results
Molecular typing identified the presence of three different LRSE clusters: PFGE type I/ST168 (n =5), PFGE type II/ST5 (n =10) and PFGE type III/ST2 (n =1). Ten strains harboured the cfr gene; we also detected mutations in the respective ribosomal protein genes. WGS revealed an almost identical 39 kb cfr plasmid obtained from strains of different genetic background (ST2, ST5, ST168) that shows high similarity to the recently published LRSE plasmid p12-02300. Due to an increase in the number of patients treated for infections with MRSA, a significant increase in linezolid usage was noted from January to July 2014 (from 5.55 to 20.41 DDDs/100 patient-days).
Conclusions
Here, we report the molecular epidemiology of LRSE in an ICU. Our results suggest the selection of resistant mutants under linezolid treatment as well as the spread of cfr-carrying plasmids. The reduction of linezolid usage and the strengthening of contact precautions proved to be effective infection control measures.



Novel Tn916-like elements confer aminoglycoside/macrolide co-resistance in clinical isolates of Streptococcus gallolyticus ssp. gallolyticus

Fri, 09 Feb 2018 00:00:00 GMT

Abstract
Background
Streptococcus gallolyticus ssp. gallolyticus (Sgg) is a commensal bacterium and an opportunistic pathogen. In humans it has been clinically associated with the incidence of colorectal cancer (CRC) and epidemiologically recognized as an emerging cause of infective endocarditis (IE). The standard therapy of Sgg includes the administration of a penicillin in combination with an aminoglycoside. Even though penicillin-resistant isolates have still not been reported, epidemiological studies have shown that this microbe is a reservoir of multiple acquired genes, conferring resistance to tetracyclines, aminoglycosides, macrolides and glycopeptides. However, the underlying antibiotic resistance mobilome of Sgg remains poorly understood.
Objectives
To investigate the mobile genetic basis of antibiotic resistance in multiresistant clinical Sgg.
Methods
Isolate NTS31106099 was recovered from a patient with IE and CRC at Nantes University Hospital, France and studied by Illumina WGS and comparative genomics. Molecular epidemiology of the identified mobile element(s) was performed using antibiotic susceptibility testing (AST), PCR, PFGE and WGS. Mobility was investigated by PCR and filter mating.
Results
Two novel conjugative transposons, Tn6263 and Tn6331, confer aminoglycoside/macrolide co-resistance in clinical Sgg. They display classical family Tn916/Tn1545 modular architecture and harbour an aph(3′)-III→sat4ant(6)-Ia→erm(B) multiresistance gene cluster, related to pRE25 of Enterococcus faecium. These and/or closely related elements are highly prevalent among genetically heterogeneous clinical isolates of Sgg.
Conclusions
Previously unknown Tn916-like mobile genetic elements conferring aminoglycoside/macrolide co-resistance make Sgg, collectively with other gut Firmicutes such as enterococci and eubacteria, a potential laterally active reservoir of these antibiotic resistance determinants among the mammalian gastrointestinal microbiota.



Discrepancies in national time trends of outpatient antibiotic utilization using different measures: a population-based study in France

Fri, 09 Feb 2018 00:00:00 GMT

Abstract
Objectives
To assess time trends of outpatient antibiotic utilization using different measures and explore their discrepancies.
Methods
Based on French sales data from the IQVIA SDM database, 2009–16, we assessed time trends in outpatient antibiotic utilization using PrID, DID, PID and SID (defined as the number of prescriptions, DDDs, packages and standard units per 1000 inhabitants per day, respectively). We explored discrepancies between trends in PrID and DID by modelling the number of DDDs per prescription.
Results
Outpatient antibiotic utilization (n =538.2 million projected prescriptions) decreased in terms of PrID, PID and SID (–10%, –8% and –8%, respectively; negative regression slopes; P <0.01), but remained stable according to DID (+2%; slope 0.009; P =0.4). The number of DDDs per prescription increased over time (+14%; slope 0.019; P <0.001). The proportions of amoxicillin and amoxicillin/clavulanate were positively associated with the number of DDDs per prescription (adjusted coefficients 0.10 and 0.15, respectively; both P <0.05), as well as the proportion of adult and hospital prescriptions (adjusted coefficients 0.07 and 0.05, respectively; both P <0.05). The discrepancy between DID and PrID disappeared when the DDD of amoxicillin was increased to values higher than the current DDD.
Conclusions
Time trends in outpatient antibiotic utilization expressed as PrID, DID, PID and SID provided conflicting results. We caution against using DID alone when monitoring antibiotic utilization. Instead, we recommend monitoring both DID and PrID as they provide different types of relevant information, especially when studying trends at a national level.



Effectiveness of general practitioner online training and an information booklet for parents on antibiotic prescribing for children with respiratory tract infection in primary care: a cluster randomized controlled trial

Fri, 09 Feb 2018 00:00:00 GMT

Abstract
Objectives
Antibiotics are too often prescribed in childhood respiratory tract infection (RTI), despite limited effectiveness, potential side effects and bacterial resistance. We aimed to reduce antibiotic prescribing for children with RTI by online training for general practitioners (GPs) and information for parents.
Methods
A pragmatic cluster randomized controlled trial in primary care. The intervention consisted of online training for GPs and an information booklet for parents. The primary outcome was the antibiotic prescription rate for children presenting with RTI symptoms, as registered by GPs. Secondary outcomes were number of reconsultations within the same disease episode, consultations for new episodes, hospital referrals and pharmacy-dispensed antibiotic courses for children. This trial was registered at the Dutch Trial Register (NTR), registration number: NTR4240.
Results
After randomization, GPs from a total of 32 general practices registered 1009 consultations. An antibiotic was prescribed in 21% of consultations in the intervention group, compared with 33% in the usual care group, controlled for baseline prescribing (rate ratio 0.65, 95% CI 0.46–0.91). The probability of reconsulting during the same RTI episode did not differ significantly between the intervention and control groups, and nor did the numbers of consultations for new episodes and hospital referrals. In the intervention group antibiotic dispensing was 32 courses per 1000 children/year lower than the control group, adjusted for baseline prescribing (rate ratio 0.78, 95% CI 0.66–0.92). The numbers and proportion of second-choice antibiotics did not differ significantly.
Conclusions
Concise, feasible, online GP training, with an information booklet for parents, showed a relevant reduction in antibiotic prescribing for children with RTI.



Presence and molecular characteristics of oxazolidinone resistance in staphylococci from household animals in rural China

Wed, 07 Feb 2018 00:00:00 GMT

Abstract
Objectives
To investigate the presence and molecular characteristics of oxazolidinone resistance genes cfr and optrA in staphylococci from household animals in rural China.
Methods
Various samples were collected from household animals in 12 rural villages. Staphylococcal isolates showing florfenicol MICs ≥10 mg/L were identified and screened for the presence of cfr and/or optrA. PCR-positive isolates were characterized by antimicrobial susceptibility testing, S1 nuclease PFGE and Southern blotting. WGS data were analysed to identify the core-genome phylogenetic profile of each isolate as well as the genetic environment of cfr and/or optrA.
Results
Nine optrA-positive (seven Staphylococcus sciuri and two Staphylococcus simulans) and 10 cfr-positive staphylococci were identified from eight and five villages, respectively. The gene optrA was chromosomally encoded in all nine isolates, whereas cfr was located on a plasmid in one S. sciuri and three Staphylococcus saprophyticus and in the chromosomal DNA of single Staphylococcus cohnii and Staphylococcus lentus isolates and two S. sciuri isolates. The remaining two cfr-carrying Staphylococcus haemolyticus isolates were indistinguishable by PFGE. Most optrA- or cfr-carrying staphylococci also harboured phenicol, tetracycline and/or macrolide-lincosamide-streptogramin B resistance genes. Genetic environment analysis showed that, for the first time, optrA was associated with transposon Tn6261, while cfr was adjacent to both a tnp (transposase) gene and a Tn558 transposon.
Conclusions
The current study reveals for the first time the wide distribution of oxazolidinone resistance genes optrA and cfr in household animals in rural areas of China and is the first identification of optrA in S. simulans isolates.



Development of a dosing nomogram for continuous-infusion meropenem in critically ill patients based on a validated population pharmacokinetic model

Wed, 07 Feb 2018 00:00:00 GMT

Abstract
Background
Optimal antibiotic exposure is a vital but challenging prerequisite for achieving clinical success in ICU patients.
Objectives
To develop and externally validate a population pharmacokinetic model for continuous-infusion meropenem in critically ill patients and to establish a nomogram based on a routinely available marker of renal function.
Methods
A population pharmacokinetic model was developed in NONMEM® 7.3 based on steady-state meropenem concentrations (CSS) collected during therapeutic drug monitoring. Different serum creatinine-based markers of renal function were compared for their influence on meropenem clearance (the Cockcroft–Gault creatinine clearance CLCRCG, the CLCR bedside estimate according to Jelliffe, the Chronic Kidney Disease Epidemiology Collaboration equation and the four-variable Modification of Diet in Renal Disease equation). After validation of the pharmacokinetic model with independent data, a dosing nomogram was developed, relating renal function to the daily doses required to achieve selected target concentrations (4/8/16 mg/L) in 90% of the patients. Probability of target attainment was determined for efficacy (CSS ≥8 mg/L) and potentially increased likelihood of adverse drug reactions (CSS >32 mg/L).
Results
In total, 433 plasma concentrations (3.20–48.0 mg/L) from 195 patients (median/P0.05 – P0.95 at baseline: weight 77.0/55.0–114 kg, CLCRCG 63.0/19.6–168 mL/min) were used for model building. We found that CLCRCG best described meropenem clearance (CL = 7.71 L/h, CLCRCG = 80 mL/min). The developed model was successfully validated with external data (n =171, 73 patients). According to the nomogram, daily doses of 910/1480/2050/2800/3940 mg were required to reach a target CSS = 8 mg/L in 90% of patients with CLCRCG = 20/50/80/120/180 mL/min, respectively. A low probability of adverse drug reactions (<0.5%) was associated with these doses.
Conclusions
A dosing nomogram was developed for continuous-infusion meropenem based on renal function in a critically ill population.



Meropenem potentiation of aminoglycoside activity against Pseudomonas aeruginosa: involvement of the MexXY-OprM multidrug efflux system

Tue, 06 Feb 2018 00:00:00 GMT

Abstract
Objectives
To assess the ability of meropenem to potentiate aminoglycoside (AG) activity against laboratory and AG-resistant cystic fibrosis (CF) isolates of Pseudomonas aeruginosa and to elucidate its mechanism of action.
Methods
AG resistance gene deletions were engineered into P. aeruginosa laboratory and CF isolates using standard gene replacement technology. Susceptibility to AGs ± meropenem (at ½ MIC) was assessed using a serial 2-fold dilution assay. mexXY expression and MexXY-OprM efflux activity were quantified using quantitative PCR and an ethidium bromide accumulation assay, respectively.
Results
A screen for agents that rendered WT P. aeruginosa susceptible to a sub-MIC concentration of the AG paromomycin identified the carbapenem meropenem, which potentiated several additional AGs. Meropenem potentiation of AG activity was largely lost in a mutant lacking the MexXY-OprM multidrug efflux system, an indication that it was targeting this efflux system in enhancing P. aeruginosa susceptibility to AGs. Meropenem failed to block AG induction of mexXY expression or MexXY-OprM efflux activity, suggesting that it may be interfering with some MexXY-dependent process linked to AG susceptibility. Meropenem potentiated AG activity versus AG-resistant CF isolates, enhancing susceptibility to at least one AG in all isolates and susceptibility to all tested AGs in 50% of the isolates. Notably, meropenem potentiation of AG activity was linked to MexXY in some but not all CF isolates in which this was examined.
Conclusions
Meropenem potentiates AG activity against laboratory and CF strains of P. aeruginosa, both dependent on and independent of MexXY, highlighting the complexity of AG resistance in this organism.



Potent Plasmodium falciparum gametocytocidal compounds identified by exploring the kinase inhibitor chemical space for dual active antimalarials

Tue, 06 Feb 2018 00:00:00 GMT

Abstract
Objectives
Novel chemical tools to eliminate malaria should ideally target both the asexual parasites and transmissible gametocytes. Several imidazopyridazines (IMPs) and 2-aminopyridines (2-APs) have been described as potent antimalarial candidates targeting lipid kinases. However, these have not been extensively explored for stage-specific inhibition of gametocytes in Plasmodium falciparum parasites. Here we provide an in-depth evaluation of the gametocytocidal activity of compounds from these chemotypes and identify novel starting points for dual-acting antimalarials.
Methods
We evaluated compounds against P. falciparum gametocytes using several assay platforms for cross-validation and stringently identified hits that were further profiled for stage specificity, speed of action and ex vivo efficacy. Physicochemical feature extraction and chemogenomic fingerprinting were applied to explore the kinase inhibition susceptibility profile.
Results
We identified 34 compounds with submicromolar activity against late stage gametocytes, validated across several assay platforms. Of these, 12 were potent at <100 nM (8 were IMPs and 4 were 2-APs) and were also active against early stage gametocytes and asexual parasites, with >1000-fold selectivity towards the parasite over mammalian cells. Front-runner compounds targeted mature gametocytes within 48 h and blocked transmission to mosquitoes. The resultant chemogenomic fingerprint of parasites treated with the lead compounds revealed the importance of targeting kinases in asexual parasites and gametocytes.
Conclusions
This study encompasses an in-depth evaluation of the kinase inhibitor space for gametocytocidal activity. Potent lead compounds have enticing dual activities and highlight the importance of targeting the kinase superfamily in malaria elimination strategies.



Minority resistant variants are also present in HIV-2-infected antiretroviral-naive patients

Sat, 03 Feb 2018 00:00:00 GMT

Abstract
Objectives
To assess the prevalence of minority resistant variants (MRV) and X4-tropic minority variants in ART-naive HIV-2-infected patients.
Patients and methods
ART-naive HIV-2-infected patients with detectable plasma viral load (>100 copies/mL) included in the ANRS HIV-2 CO5 Cohort were assessed. We performed ultra-deep sequencing (UDS) of protease, RT, integrase and gp105 regions. Only mutations in the HIV-2 ANRS list >1% were considered. HIV-2 tropism was assessed by V3 loop region UDS, and each read was interpreted with determinants of CXCR4-coreceptor use.
Results
Among the 47 patients assessed, three displayed plasma viruses with a resistance-associated mutation (RAM) above the 20% detection threshold, all in RT, resulting in a prevalence of transmitted drug resistance for NRTI of 7.9% (95% CI 0.0%–16.5%). No RAM above the 20% detection threshold was found in protease or integrase. At the 1% detection threshold the transmitted drug resistance prevalence was 9.8% (95% CI 0.6%–19.0%), 13.2% (95% CI 3.5%–22.9%) and 4.5% (95% CI 0%–17.5%) for PI, NRTI and integrase inhibitors. The most prevalent MRV was the PI RAM I50V detected in three samples. Tropism analysis showed that 21% of patients (4 of 19) exhibited X4-tropic viruses: two in majority proportion and two in minority proportions (1.5% and 1.9%).
Conclusions
In this first study assessing the prevalence of MRV in HIV-2 infection among ART-naive patients, we observed a 2–3-fold higher prevalence of RAM when a 1% detection threshold of mutations was used compared with a 20% threshold. Similarly, the proportion of patients with X4-tropic viruses was twice as high when UDS was used.



Analysis of the presence of erroneous Qnr sequences in GenBank

Sat, 03 Feb 2018 00:00:00 GMT

Abstract
Background
Twenty years ago the first transferable mechanism of quinolone resistance (TMQR), QnrA, was described. Thereafter, innumerable TMQRs, either Qnr related or not, were described. Ten years ago the exponential description of Qnr genes/alleles led to the proposal of a common nomenclature.
Objectives
This analysis aims to determine the degree of correctness of the Qnr sequences currently present in GenBank.
Methods
The Qnr amino acid type sequence of the first allele (e.g. QnrA1) of each Qnr family present in http://www.lahey.org/qnrStudies/ was compared with what is present in GenBank. Only the first 30 obtained annealings or those with a >90% identity were considered. No synthetic or chromosomal sequences (other than those included in http://www.lahey.org/qnrStudies/) were included in the analyses.
Results
Overall, 1657 amino acid sequences were analysed: 224 QnrA, 499 QnrB, 1 QnrC, 102 QnrD, 13 QnrE, 758 QnrS and 60 QnrVC. Of these, 340 (20.5%) sequences presented a major error, including erroneous gene name, erroneous Qnr family attribution, erroneous allele identification, presence of partial sequences with allele assignation and/or erroneous initial codon. In addition, 449 (27.1%) Qnr sequences were present in GenBank with a partial identification or not identified as Qnr. Finally, nine new transferable Qnr alleles were detected.
Conclusions
These data highlight the frequent presence of erroneously identified qnr genes in GenBank and the need to be fully adherent to current nomenclature rules.



Impact of the ST101 clone on fatality among patients with colistin-resistant Klebsiella pneumoniae infection

Sat, 03 Feb 2018 00:00:00 GMT

Abstract
Objectives
We describe the molecular characteristics of colistin resistance and its impact on patient mortality.
Methods
A prospective cohort study was performed in seven different Turkish hospitals. The genotype of each isolate was determined by MLST and repetitive extragenic palindromic PCR (rep-PCR). Alterations in mgrB were detected by sequencing. Upregulation of pmrCAB, phoQ and pmrK was quantified by RT–PCR. mcr-1 and the genes encoding OXA-48, NDM-1 and KPC were amplified by PCR.
Results
A total of 115 patients diagnosed with colistin-resistant K. pneumoniae (ColR-Kp) infection were included. Patients were predominantly males (55%) with a median age of 63 (IQR 46–74) and the 30 day mortality rate was 61%. ST101 was the most common ST and accounted for 68 (59%) of the ColR-Kp. The 30 day mortality rate in patients with these isolates was 72%. In ST101, 94% (64/68) of the isolates had an altered mgrB gene, whereas the alteration occurred in 40% (19/47) of non-ST101 isolates. The OXA-48 and NDM-1 carbapenemases were found in 93 (81%) and 22 (19%) of the total 115 isolates, respectively. In multivariate analysis for the prediction of 30 day mortality, ST101 (OR 3.4, CI 1.46–8.15, P =0.005) and ICU stay (OR 7.4, CI 2.23–29.61, P =0.002) were found to be significantly associated covariates.
Conclusions
Besides ICU stay, ST101 was found to be a significant independent predictor of patient mortality among those infected with ColR-Kp. A significant association was detected between ST101 and OXA-48. ST101 may become a global threat in the dissemination of colistin resistance and the increased morbidity and mortality of K. pneumoniae infection.



Performance evaluation of the QMAC-dRAST for staphylococci and enterococci isolated from blood culture: a comparative study of performance with the VITEK-2 system

Sat, 03 Feb 2018 00:00:00 GMT

Abstract
Objectives
To evaluate the performance of a rapid antimicrobial susceptibility testing (AST) platform based on microfluidic chip technology, the QMAC-dRAST, which enables AST from colony isolates or positive blood culture broth (PBCB), and to compare the performance of the QMAC-dRAST for staphylococci and enterococci with that of the VITEK-2 system based on reference broth microdilution (BMD).
Methods
A total of 110 staphylococcal and enterococcal isolates from blood cultures were included. AST was performed directly using the QMAC-dRAST with PBCB. Thereafter, colony isolates derived from subculture of PBCB were used for the QMAC-dRAST, the VITEK-2 system and BMD.
Results
The overall agreement between the QMAC-dRAST with PBCB and BMD was 91.5%. There were 1.2% very major errors (VMEs), 4.3% major errors (MEs) and 5.4% minor errors (mEs). The QMAC-dRAST with colony isolates yielded 94.6% agreement and error rates of 1.0% VMEs, 1.8% MEs and 4.0% mEs. The VITEK-2 system showed 96.2% agreement and error rates of 2.3% VMEs, 0.5% MEs and 2.6% mEs. The incubation time in the QMAC-dRAST was significantly shorter than in the VITEK-2 system (median of 6 versus 10 h; P <0.0001).
Conclusions
The QMAC-dRAST system provided rapid results and represents an alternative to conventional AST methods. The QMAC-dRAST with colony isolates produced more reliable results for staphylococci and enterococci than the QMAC-dRAST with PBCB. The QMAC-dRAST system also performed comparably to BMD and the VITEK-2 system.



A mathematical model-based analysis of the time–kill kinetics of ceftazidime/avibactam against Pseudomonas aeruginosa

Sat, 03 Feb 2018 00:00:00 GMT

Abstract
Objectives
To characterize quantitatively the effect of avibactam in potentiating ceftazidime against MDR Pseudomonas aeruginosa by developing a mathematical model to describe the bacterial response to constant concentration time–kill information and validating it using both constant and time-varying concentration–effect data from in vitro and in vivo infection systems.
Methods
The time course of the bacterial population dynamics in the presence of static concentrations of ceftazidime and avibactam was modelled using a two-state pharmacokinetic/pharmacodynamic (PK/PD) model, consisting of active and resting states, to account for bactericidal activities, bacteria-mediated ceftazidime degradation and inhibition of degradation by avibactam. Ceftazidime's effect on the bacterial population was described as an enhancement of the death rate of the active population, with the effect of avibactam being to increase ceftazidime potency. Model validation was performed by comparing simulated time courses of bacterial responses with those from in vitro and in vivo experimental exposures of ceftazidime and avibactam that represented those predicted in an average patient dosed with 2 g/0.5 g ceftazidime/avibactam administered every 8 h as 2 h infusions.
Results
The two-state model successfully described the bacterial population dynamics, ceftazidime degradation and its inhibition by avibactam. For external validation, the model correctly predicted the bacterial response of P. aeruginosa isolates evaluated in in vitro hollow-fibre and in vivo neutropenic mouse thigh and lung infection models.
Conclusions
The PK/PD model and modelled strains successfully replicated the spread in activity when compared with a large selection of P. aeruginosa strains reported in the literature.



Population structure of Streptococcus pneumoniae colonizing children before and after universal use of pneumococcal conjugate vaccines in Brazil: emergence and expansion of the MDR serotype 6C-CC386 lineage

Thu, 01 Feb 2018 00:00:00 GMT

Abstract
Objectives
To determine the population structure and change in drug resistance of pneumococci colonizing children before and after the introduction of the 10-valent and 13-valent pneumococcal conjugate vaccines (PCV10/13) in Brazil.
Methods
We used MLST to analyse 256 pneumococcal isolates obtained from children aged <6 years before (2009–10; n =125) and after (2014; n =131) the introduction of the PCV10 and PCV13. Antimicrobial susceptibility and capsular types were previously determined.
Results
We identified 97 different STs. Ninety (35.2%) isolates were related to international clones. The most frequent lineages were serogroup 6-CC724 (where CC stands for clonal complex) and the MDR serotype 6C-CC386 in the pre- and post-PCV10/13 periods, respectively. Penicillin-non-susceptible pneumococci (PNSP) formed 24% and 38.9% of the pre- and post-PCV10/13 isolates, respectively (P =0.01). In the pre-PCV10/13 period, serotype 14-ST156 was the predominant penicillin-non-susceptible lineage, but it was not detected in the post-PCV10/13 period. Serotype 14-ST156 and serotype 19A-ST320 complex isolates had the highest penicillin and ceftriaxone MICs in the pre- and post-PCV10/13 periods, respectively. In turn, serotype 6C-CC386 comprised almost 30% of the PNSP and over 40% of the erythromycin-resistant isolates (MIC >256 mg/L) in the post-PCV10/13 period.
Conclusions
Although PNSP strains were polyclonal, most resistant isolates belonged to a single genotype from each period. Higher erythromycin resistance prevalence (42%) in the post-PCV10/13 period was mainly attributed to MDR serotype 6C-CC386. Ongoing surveillance of pneumococcal clonal composition is important to evaluate PCV use outcomes and to identify factors other than PCVs that drive pneumococcal drug resistance evolution.



Identification of novel variants of the colistin resistance gene mcr-3 in Aeromonas spp. from the national resistance monitoring programme GERM-Vet and from diagnostic submissions

Wed, 31 Jan 2018 00:00:00 GMT

Abstract
Objectives
To investigate Aeromonas spp. isolates for the presence of the novel resistance gene mcr-3 or variants thereof and to characterize the positive isolates by whole genome sequence analysis.
Methods
A total of 479 unrelated Aeromonas isolates were investigated by PCR for the genes mcr-1, mcr-2 and mcr-3. Positive isolates were investigated for their colistin MICs. Species assignment was based on sequence analysis of 16s rRNA and gyrB and rpoB genes. The mcr-carrying contigs obtained by WGS were analysed for the genetic environments of the mcr genes.
Results
Four (0.84%) Aeromonas isolates were positive in the mcr-3-specific PCR assay, whereas none of the isolates harboured mcr-1 or mcr-2. Each of the four mcr-3 genes encoded a novel variant, which showed amino acid identities of 95.0%–98.0% to the original Mcr-3 protein. These variants were designated Mcr-3.6 [Aeromonas allosaccharophila from golden orfe (Leuciscus idus)], Mcr-3.7 [Aeromonas media from turkey (Meleagris gallopavo)], Mcr-3.8 [Aeromonas jandaei from koi carp (Cyprinus carpio)] and Mcr-3.9 [Aeromonas hydrophila from koi carp]. The isolate harbouring the mcr-3.9 gene carried an additional mcr-3.8 gene and showed a distinctly higher colistin MIC of ≥128 mg/L than all other isolates. The genetic environments of the mcr-3 variant genes in all four isolates differed, but in part resembled the flanking regions of mcr-3.3 from Aeromonas veronii of chicken meat.
Conclusions
This study identified four novel Mcr-3 variants. The isolates carrying the respective genes dated back to 2005 suggesting that this gene has existed for more than 12 years.



Serological response to therapy following retreatment of serofast early syphilis patients with benzathine penicillin

Wed, 31 Jan 2018 00:00:00 GMT

Abstract
Background
Some syphilitic patients remain in a serologically positive state after the recommended therapy. Although we often retreat patients in clinical practice, the optimal treatment protocol remains uncertain due to the paucity of data regarding serological response to retreatment and long-term outcomes.
Methods
We examined rapid plasma reagin serological test results of 70 serofast early syphilis cases who were retreated with 2.4 million units of benzathine penicillin weekly for 3 weeks. Serological retreatment success was defined as a minimum 4-fold decrease in baseline rapid plasma reagin test antibody titre within 6 months.
Results
Thirty-four (48.6%) of the patients who failed to achieve serological cure at 6 months after initial therapy achieved serological cure at 12 months. Patients who had higher non-treponemal titres at baseline and at 6 months were more likely to exhibit serological cure after retreatment than those with lower titres.
Conclusions
Our results suggest that the incremental benefit of retreating serofast patients with early syphilis is moderate, considering the almost 1:1 ratio of serological response to serofast state at follow-up.



Epidemiology of invasive aspergillosis and triazole-resistant Aspergillus fumigatus in patients with haematological malignancies: a single-centre retrospective cohort study

Wed, 31 Jan 2018 00:00:00 GMT

Abstract
Objectives
To investigate the epidemiology and clinical relevance of triazole resistance among patients undergoing treatment for haematological malignancies who are at risk of invasive aspergillosis (IA).
Methods
This was a retrospective cohort study for which the records of consecutive patients given chemotherapy for AML or myelodysplastic syndrome (MDS) or who had received an allogeneic HSCT from 2006 to 2012 were reviewed for IA. Triazole resistance was detected by the VIPcheck™ screening method and confirmed by determining the MIC by EUCAST methodology.
Results
A total of 432 patients were included, comprising 182 (42.1%) patients who had undergone chemotherapy for AML or MDS, and 250 (57.9%) patients who had undergone an allogeneic HSCT. Probable or proven IA was diagnosed in 36 cases (8.3%, 95% CI 6.0%–11.4%). Of these, 12 (33.3%) were based on recovery of Aspergillus fumigatus from sputum, bronchoalveolar lavage or biopsy, and triazole resistance was found in 2 instances. A. fumigatus was also recovered from one or more specimens from 13 patients without probable or proven IA. Triazole resistance was documented for three patients. The survival rate of patients with IA caused by voriconazole-resistant isolates could not be assessed.
Conclusions
The overall frequency of voriconazole-resistant IA among patients at high risk was low. However, the rate of triazole resistance may have been underestimated by the low detection rate based on recovery of A. fumigatus. Alternative diagnostic tests, such as PCR-based assays, may prove better at detecting IA due to triazole-resistant A. fumigatus.



Comparison of piperacillin exposure in the lungs of critically ill patients and healthy volunteers

Mon, 29 Jan 2018 00:00:00 GMT

Abstract
Background
Severe infections of the respiratory tracts of critically ill patients are common and associated with excess morbidity and mortality. Piperacillin is commonly used to treat pulmonary infections in critically ill patients. Adequate antibiotic concentration in the epithelial lining fluid (ELF) of the lung is essential for successful treatment of pulmonary infection.
Objectives
To compare piperacillin pharmacokinetics/pharmacodynamics in the serum and ELF of healthy volunteers and critically ill patients.
Methods
Piperacillin concentrations in the serum and ELF of healthy volunteers and critically ill patients were compared using population methodologies.
Results
Median piperacillin exposure was significantly higher in the serum and the ELF of critically ill patients compared with healthy volunteers. The IQR for serum piperacillin exposure in critically ill patients was six times greater than for healthy volunteers. The IQR for piperacillin exposure in the ELF of critically ill patients was four times greater than for healthy volunteers. The median pulmonary piperacillin penetration ratio was 0.31 in healthy volunteers and 0.54 in critically ill patients.
Conclusions
Greater variability in serum and ELF piperacillin concentrations is observed in critically ill patients compared with healthy adult subjects and must be considered in the development of dosage regimens. Pulmonary penetration of antimicrobial agents should be studied in critically ill patients, as well as healthy volunteers, during drug development to ensure appropriate dosing of patients with pneumonia.



Oral doxycycline versus intravenous ceftriaxone for treatment of multiple erythema migrans: an open-label alternate-treatment observational trial

Mon, 29 Jan 2018 00:00:00 GMT

Abstract
Background
Several guidelines advocate the same treatment approaches for both early disseminated Lyme borreliosis, manifested as multiple erythema migrans (EM), and early localized Lyme borreliosis, manifested as solitary EM.
Methods
Oral doxycycline (100 mg q12h) was compared on a non-inferiority premise with intravenous ceftriaxone (2 g q24h) for 14 days in 200 adult European patients with multiple EM in an open-label alternate-treatment observational trial performed in a single-centre university hospital. Treatment outcome was assessed at 14 days and at 2, 6 and 12 months post-enrolment. Non-specific symptoms in patients and 192 control subjects without a history of Lyme borreliosis were evaluated and compared. This trial was registered at http://clinicaltrials.gov (identifier NCT01163994).
Results
At the 12 month visit, 4/82 (4.9%) multiple EM patients prescribed doxycycline and 6/88 (6.8%) multiple EM patients prescribed ceftriaxone showed incomplete response manifested predominantly as post-Lyme symptoms (1.9% difference, upper limit of 95% CI 5.1%). The upper limit of 95% CI for the difference in proportion of patients with incomplete response between doxycycline and ceftriaxone groups did not exceed the predetermined non-inferiority margin of 10%. The frequency of non-specific symptoms in patients was similar to that in controls.
Conclusions
The 14 day oral doxycycline was not inferior to the 14 day intravenous ceftriaxone in treatment of adult European patients with early disseminated Lyme borreliosis manifested as multiple EM. The frequency of non-specific symptoms in patients was similar to that in controls without a history of Lyme borreliosis.



Comparison of risk factors for, and prevalence of, antibiotic resistance in contaminating and pathogenic urinary Escherichia coli in children in primary care: prospective cohort study

Mon, 29 Jan 2018 00:00:00 GMT

Abstract
Background
All-cause antibiotic prescribing affects bowel flora antimicrobial susceptibility, and may increase risk of urinary autoinoculation with antibiotic-resistant microbes. However, little is known about relative prevalence of, or risk factors for, antimicrobial resistance among potentially pathogenic microbes thought to be contaminating and infecting urine.
Methods
Secondary analysis of 824 children under 5 years of age consulting in primary care for an acute illness and their Escherichia coli isolates cultured at ≥103 cfu/mL from the Diagnosis of Urinary Tract infection in Young children (DUTY) study. Multivariable logistic regression investigating risk factors for resistance to amoxicillin, co-amoxiclav, cefalexin, ciprofloxacin, trimethoprim, nitrofurantoin and cefpodoxime in microbes meeting the laboratory criteria for urinary tract infection: ‘pathogens’ (>105 cfu/mL, n =79) and ‘contaminants’ (103 to 105 cfu/mL, n =745).
Results
Forty-three percent of E. coli were resistant to at least one tested antibiotic, with resistance highest to amoxicillin (49.37% pathogenic versus 37.32% contaminant, P =0.04), trimethoprim (27.85% versus 16.52%, P =0.01) and co-amoxiclav (16.46% versus 21.48%, P =0.30). Multidrug resistance (to ≥3 antibiotic groups) was present in 17.07% of pathogens and 30.13% of contaminants (P =0.04). No isolates were resistant to nitrofurantoin. Recent (0–3 months) exposure to antibiotics was associated with resistance in both pathogens (aOR: 1.10, 95% CI: 1.01–4.39) and contaminants (1.69, 1.09–2.67).
Conclusions
Prevalence of resistance (including multidrug) was high, but there was no consistent relationship between isolate pathogen/contamination status and resistance. Recent all-cause antibiotic prescribing increased the probability of antimicrobial resistance in both pathogenic and contaminating urinary E. coli in children in primary care.



Pathway involving the N155H mutation in HIV-1 integrase leads to dolutegravir resistance

Wed, 24 Jan 2018 00:00:00 GMT

Abstract
Background
Dolutegravir, an integrase strand-transfer inhibitor (STI), shows a high genetic barrier to resistance. Dolutegravir is reported to be effective against viruses resistant to raltegravir and elvitegravir. In this study, we report the case of a patient treated with dolutegravir monotherapy. Failure of dolutegravir treatment was observed concomitant with the appearance of N155H-K211R-E212T mutations in the integrase (IN) gene in addition to the polymorphic K156N mutation that was present at baseline in this patient.
Methods
The impact of N155H-K156N-K211R-E212T mutations was studied in cell-free, culture-based assays and by molecular modelling.
Results
Cell-free and culture-based assays confirm that selected mutations in the patient, in the context of the polymorphic mutation K156N present at the baseline, lead to high resistance to dolutegravir requiring that the analysis be done at timepoints longer than usual to properly reveal the results. Interestingly, the association of only N155H and K156N is sufficient for significant resistance to dolutegravir. Modelling studies showed that dolutegravir is less stable in IN/DNA complexes with respect to the WT sequence.
Conclusions
Our results indicate that the stability of STI IN/DNA complexes is an important parameter that must be taken into account when evaluating dolutegravir resistance. This study confirms that a pathway including N155H can be selected in patients treated with dolutegravir with the help of the polymorphic K156N that acts as a secondary mutation that enhances the resistance to dolutegravir.



Azithromycin-resistant Neisseria gonorrhoeae spreading amongst men who have sex with men (MSM) and heterosexuals in New South Wales, Australia, 2017

Wed, 24 Jan 2018 00:00:00 GMT

Abstract
Objectives
To identify the genetic basis of resistance as well as to better understand the epidemiology of a recent surge in azithromycin-resistant Neisseria gonorrhoeae in New South Wales, Australia.
Methods
Azithromycin-resistant N. gonorrhoeae isolates (n =118) collected from 107 males, 10 females and 1 transsexual between January and July 2017 were genotyped using a previously described iPLEX method. The results were compared with phenotypic resistance profiles and available patient data.
Results
The iPLEX results revealed 10 different N. gonorrhoeae genotypes (designated AZI-G1 to AZI-G10) of which three were responsible for the majority of infections; AZI-G10 (74.6%, 88 isolates; 87 males and 1 transsexual), AZI-G4 (11.0%, 13 isolates; 7 males and 6 females) and AZI-G7 (6.8%, 8 isolates; 7 males and 1 female). The observed resistance was attributable to one of two different azithromycin resistance mechanisms; the 23S rRNA C2611T mutation was identified in 24% of isolates, whereas the majority of resistance (76%) was associated with a meningococcal-type mtrR variant. Additionally, one isolate was found to harbour both the 23S rRNA C2611T mutation and a type XXXIV mosaic penA sequence associated with cephalosporin resistance.
Conclusions
These data indicate outbreaks of azithromycin-resistant gonococci amongst networks of MSM and heterosexuals in New South Wales. The results also provide further evidence that azithromycin may soon be an ineffective treatment option for gonococcal infection and highlight the urgent need to explore alternative therapies.



Plasmids carrying antimicrobial resistance genes in Enterobacteriaceae

Tue, 23 Jan 2018 00:00:00 GMT

Abstract
Bacterial antimicrobial resistance (AMR) is constantly evolving and horizontal gene transfer through plasmids plays a major role. The identification of plasmid characteristics and their association with different bacterial hosts provides crucial knowledge that is essential to understand the contribution of plasmids to the transmission of AMR determinants. Molecular identification of plasmid and strain genotypes elicits a distinction between spread of AMR genes by plasmids and dissemination of these genes by spread of bacterial clones. For this reason several methods are used to type the plasmids, e.g. PCR-based replicon typing (PBRT) or relaxase typing. Currently, there are 28 known plasmid types in Enterobacteriaceae distinguished by PBRT. Frequently reported plasmids [IncF, IncI, IncA/C, IncL (previously designated IncL/M), IncN and IncH] are the ones that bear the greatest variety of resistance genes. The purpose of this review is to provide an overview of all known AMR-related plasmid families in Enterobacteriaceae, the resistance genes they carry and their geographical distribution.



Recombination of plasmids in a carbapenem-resistant NDM-5-producing clinical Escherichia coli isolate

Tue, 23 Jan 2018 00:00:00 GMT

Abstract
Objectives
To investigate the genetic features of five plasmids recovered from an NDM-5-producing clinical Escherichia coli strain, BJ114, and to characterize the plasmid recombination event that occurred during the conjugation process.
Methods
The genetic profiles of the five plasmids were determined by PCR, conjugation, S1-PFGE, Southern hybridization and WGS analysis. Plasmid sequences were analysed with various bioinformatic tools.
Results
Complete sequences of five plasmids were obtained. Two small plasmids, pBJ114-141 and pBJ114-46, were speculated to have recombined into a large fusion plasmid, pBJ114T-190. When conjugated to other E. coli strains, some of the fusion plasmids were able to be resolved into the original two single plasmids. A non-conjugative plasmid, pBJ114-96, exhibited a high degree of sequence identity with the phage P7-like plasmid as well as an mcr-1-bearing plasmid. Another plasmid, pBJ114-78, was found to contain multidrug resistance genes and various mobile elements.
Conclusions
The fusion plasmid recoverable from the transconjugant was found to be generated as a result of a recombination event that occurred upon interaction between a blaNDM-5-carrying plasmid and another plasmid present in the parental strain. Such recombination events presumably play a potential role in the dissemination of the blaNDM genes among different plasmids and pathogenic bacterial strains.



Evolution of drug resistance in Mycobacterium tuberculosis: a review on the molecular determinants of resistance and implications for personalized care

Fri, 19 Jan 2018 00:00:00 GMT

Abstract
Drug-resistant TB (DR-TB) remains a significant challenge in TB treatment and control programmes worldwide. Advances in sequencing technology have significantly increased our understanding of the mechanisms of resistance to anti-TB drugs. This review provides an update on advances in our understanding of drug resistance mechanisms to new, existing drugs and repurposed agents. Recent advances in WGS technology hold promise as a tool for rapid diagnosis and clinical management of TB. Although the standard approach to WGS of Mycobacterium tuberculosis is slow due to the requirement for organism culture, recent attempts to sequence directly from clinical specimens have improved the potential to diagnose and detect resistance within days. The introduction of new databases may be helpful, such as the Relational Sequencing TB Data Platform, which contains a collection of whole-genome sequences highlighting key drug resistance mutations and clinical outcomes. Taken together, these advances will help devise better molecular diagnostics for more effective DR-TB management enabling personalized treatment, and will facilitate the development of new drugs aimed at improving outcomes of patients with this disease.



PBP4 activity and its overexpression are necessary for PBP4-mediated high-level β-lactam resistance

Fri, 19 Jan 2018 00:00:00 GMT

Abstract
Background
PBP4 is typically considered unimportant for conferring high-level β-lactam resistance in Staphylococcus aureus. Mutations in PBP4 have been associated with β-lactam non-susceptibility among natural strains of S. aureus. We have previously shown that PBP4 can mediate high-level β-lactam resistance in laboratory-generated strains passaged in β-lactam antibiotics. Mutations in the pbp4 promoter that up-regulate its expression and missense mutations that surround PBP4’s active site were detected in high frequencies among passaged strains, suggesting PBP4 plays a key role in resistance. How these mutations participate in PBP4’s ability to provide high-level β-lactam resistance is unknown.
Objectives
To determine whether enzymatic activity of PBP4 is required for high-level β-lactam resistance and to investigate how the pbp4-associated mutations provide β-lactam resistance.
Methods
The catalytic activity of PBP4 was disabled through introduction of a serine to alanine point mutation in its active site (Ser-75→Ala) in a representative and well-studied passaged strain, CRB. pbp4 promoter and missense mutations detected in CRB were reconstituted in a WT strain individually and in combination. β-Lactam resistance of the resultant strains was evaluated by population analysis. Bacterial peptidoglycan composition of the pbp4 mutants was evaluated with and without antibiotic treatment using LC.
Results
PBP4 inactivation imparted complete β-lactam susceptibility of CRB. Reconstitution of PBP4 missense mutations alone did not impart β-lactam resistance, but did so in synergism with pbp4 promoter mutation. A similar synergistic interaction of pbp4 mutations was observed in enhanced peptidoglycan cross-linking upon antibiotic treatment.
Conclusions
PBP4’s activity and overexpression both contribute to high-level β-lactam resistance.



Dissemination of linezolid-dependent, linezolid-resistant Staphylococcus epidermidis clinical isolates belonging to CC5 in German hospitals

Fri, 19 Jan 2018 00:00:00 GMT

Abstract
Objectives
Linezolid-resistant Staphylococcus epidermidis (LRSE) and linezolid-dependent ST22 strains have been shown to predominate in tertiary care facilities all over Greece. We report herein the dissemination of ST22 but also ST2, ST5 and ST168 linezolid-dependent LRSE clones in four unrelated German hospitals.
Methods
Fourteen LRSE clinical isolates recovered during 2012–14 from five distantly located German hospitals were tested by for MIC determination broth microdilution and Etest, PCR/sequencing for cfr and for mutations in 23S rRNA, rplC, rplD and rplV genes, MLST, PFGE and growth curves without and with linezolid at 16 and 32 mg/L.
Results
Most (11, 78.6%) isolates had linezolid MICs >256 mg/L. Five isolates carried the cfr gene. Eight isolates belonged to ST22, two isolates each to ST168 and ST2 and one isolate each to ST5 and ST23. Ten isolates [seven belonging to ST22 and one to each of ST2, ST5 and ST168; all these STs belong to clonal complex (CC) 5] exhibited linezolid-dependent growth, growing significantly faster in linezolid-containing broth. Four isolates were non-dependent (one belonging to each of ST22, ST2, ST23 and ST168). Four isolates came from three different hospitals, whereas four and six isolates were recovered during outbreaks of LRSE in two distinct hospitals.
Conclusions
The multi-clonal dissemination of CC5 linezolid-dependent LRSE throughout German hospitals along with the clonal expansion of ST22 linezolid-dependent LRSE in Greek hospitals is of particular concern. It is plausible that this characteristic is inherent and provides a selective advantage to CC5 LRSE under linezolid pressure, contributing to their dissemination throughout hospitals in these countries.



Whole-genome analysis reveals the evolution and transmission of an MDR DH/NAP11/106 Clostridium difficile clone in a paediatric hospital

Fri, 12 Jan 2018 00:00:00 GMT

Abstract
Background
Clostridium difficile strain DH/NAP11/106, a relatively antibiotic-susceptible strain, is now the most common cause of C. difficile infection (CDI) among adults in the USA.
Objectives
To identify mechanisms underlying the evolution and transmission of an MDR DH/NAP11/106 clone.
Methods
WGS (Illumina MiSeq), restriction endonuclease analysis (REA) and antibiotic susceptibility testing were performed on 134 C. difficile isolates collected from paediatric patients with CDI over a 2 year period.
Results
Thirty-one of 134 (23%) isolates were REA group DH. Pairwise single-nucleotide variant (SNV) analyses identified a DH clone causing seven instances of CDI in two patients. During the 337 days between the first and second CDI, Patient 1 (P1) received 313 days of antibiotic therapy. Clindamycin and rifaximin resistance, and reduced vancomycin susceptibility (MIC 0.5–2 mg/L), were newly identified in the relapsed isolate. This MDR clone was transmitted to Patient 2 (P2) while P1 and P2 received care in adjacent private rooms. P1 and P2 each developed two additional CDI relapses. Comparative genomics analyses demonstrated SNVs in multiple antibiotic resistance genes, including rpoB (rifaximin resistance), gyrB and a gene encoding PBP; gyrB and PBP mutations did not consistently confer a resistance phenotype. The clone also acquired a 46 000 bp genomic element, likely a conjugative plasmid, which contained ermB (clindamycin resistance). The element shared 99% identity with the genomic sequence of Faecalibacterium prausnitzii, an enteric commensal.
Conclusions
These data highlight the emergence of MDR in C. difficile strain DH/NAP11/106 through multiple independent mechanisms probably as a consequence of profound antibiotic pressure.



Does oxidative stress contribute to adverse outcomes in HIV-associated TB?

Mon, 08 Jan 2018 00:00:00 GMT

Abstract
In HIV infection, oxidative stress is a pronounced phenomenon, with likely links to HIV-related pathologies and the progression of HIV infection per se. TB is an AIDS-defining condition. HIV-associated oxidative stress, like that associated with diabetes mellitus, might adversely impact the outcomes of TB, probably through increased propensity for generation of metabolically dormant mycobacterial persisters, alongside other mechanisms. This hypothesis might help in guiding the exploration of relevant research directions to improve the care of patients.