Antimicrobial resistance in <Emphasis Type="Italic">Enterobacteriaceae</Emphasis> from healthy broilers in Egypt: emergence of colistin-resistant and extended-spectrum β-lactamase-producing <Emphasis Type="Italic">Escherichia coli</Emphasis>

Background

Poultry remains one of the most important reservoir for zoonotic multidrug resistant pathogens. The global rise of antimicrobial resistance in Gram-negative bacteria is of reasonable concern and demands intensified surveillance.

Methods

In 2016, 576 cloacal swabs were collected from 48 broiler farms located in five governorates in northern Egypt. Isolates of Enterobacteriaceae could be cultivated on different media and were identified by MALDI-TOF MS and PCR. Escherichia coli isolates were genotyped by DNA-microarray-based assays. The antimicrobial susceptibility to 14 antibiotics was determined and resistance-associated genes were detected. The VITEK-2 system was applied for phenotypical confirmation of extended-spectrum β-lactamase-producing isolates. The determination of colistin resistance was carried out phenotypically using E-test and genotypically using PCR for detection of the mcr-1 gene.

Results

Out of 576 samples, 72 representatives of Enterobacteriaceae were isolated and identified as 63 E. coli (87.5%), 5 Enterobacter cloacae (6.9%), 2 Klebsiella pneumoniae (2.8%) and 2 Citrobacter spp. (2.8%). Seven out of 56 cultivated E. coli (12.5%) were confirmed as ESBL-producing E. coli and one isolate (1.8%) as ESBL/carbapenemase-producing E. coli. Five out of 63 E. coli isolates (7.9%) recovered from different poultry flocks were phenotypically resistant to colistin and harboured mcr-1 gene.

Conclusions

This is the first study reporting colistin resistance and emergence of multidrug resistance in Enterobacteriaceae isolated from healthy broilers in the Nile Delta region, Egypt. Colistin-resistant E. coli in poultry is of public health significance. The global rise of ESBL- and carbapenemase-producing Gram-negative bacteria demands intensified surveillance. ESBL-producing E. coli in poultry farms in Egypt are of major concern that emphasizes the possibility of spread of such strains to humans. The results also reinforce the need to develop strategies and to implement specific control procedures to reduce the use of antibiotics.

     

Serial echocardiographic evaluation of the Perimount Magna Ease prosthesis

BackgroundThe Carpentier-Edwards Perimount Magna Ease prosthesis (PME) represents the latest generation of stented bioprostheses used for surgical aortic valve replacement (SAVR). The aim of our study was to evaluate the long-term clinical outcome and hemodynamic performance of the prosthesis with a focus on the incidence and course of structural valve deterioration (SVD) by serial echocardiographic examinations.MethodsSAVR with the PME was performed in 58 consecutive patients between 2007 and 2008. Transthoracic echocardiography was performed preoperatively, at discharge and annually during a 10-year follow-up at the German Heart Center Munich.ResultsMean age at surgery was 62±14 years. At discharge (n=57), the overall mean pressure gradient (MPG) and effective orifice area (EOA) were 15.8±4.1 mmHg and 1.8±0.4 cm², respectively. Moderate patient-prosthesis mismatch (PPM) was present in 18 patients (32%) and severe PPM in 6 patients (11%) at discharge. Ten years following SAVR (n=33), the overall MPG was 16.6±7.3 mmHg and EOA was 1.3±0.4 cm².Thirty-day and late mortality was 2% (n=1) and 21% (n=12), respectively. Survival at 1, 5, and 10 years was 94.7%±3.3%, 91.1%±4.1%, and 77.3%±5.9%, respectively. Freedom from reoperation at 10 years was 88.8%±4.7%. Ten years after PME implantation the cumulative incidence of any SVD, severe SVD, and bioprosthetic valve failure (BVF) was 25%±6%, 14%±5%, and 16%±5%, respectively.ConclusionsThe PME shows an excellent hemodynamic performance over the course of 10 years with development of clinically relevant SVD as late as 6 years post implant, and a 10-year incidence of severe SVD of 14%.     

The Biogenesis of Dengue Virus Replication Organelles Requires the ATPase Activity of Valosin-Containing Protein

The dengue virus (DENV) causes the most prevalent arthropod-borne viral disease worldwide. While its incidence is increasing in many countries, there is no approved antiviral therapy currently available. In infected cells, the DENV induces extensive morphological alterations of the endoplasmic reticulum (ER) to generate viral replication organelles (vRO), which include convoluted membranes (CM) and vesicle packets (VP) hosting viral RNA replication. The viral non-structural protein NS4B localizes to vROs and is absolutely required for viral replication through poorly defined mechanisms, which might involve cellular protein partners. Previous interactomic studies identified the ATPase valosin-containing protein (VCP) as a DENV NS4B-interacting host factor in infected cells. Using both pharmacological and dominant-negative inhibition approaches, we show, in this study, that VCP ATPase activity is required for efficient DENV replication. VCP associates with NS4B when expressed in the absence of other viral proteins while in infected cells, both proteins colocalize within large DENV-induced cytoplasmic structures previously demonstrated to be CMs. Consistently, VCP inhibition dramatically reduces the abundance of DENV CMs in infected cells. Most importantly, using a recently reported replication-independent plasmid-based vRO induction system, we show that de novo VP biogenesis is dependent on VCP ATPase activity. Overall, our data demonstrate that VCP ATPase activity is required for vRO morphogenesis and/or stability. Considering that VCP was shown to be required for the replication of other flaviviruses, our results argue that VCP is a pan-flaviviral host dependency factor. Given that new generation VCP-targeting drugs are currently evaluated in clinical trials for cancer treatment, VCP may constitute an attractive broad-spectrum antiviral target in drug repurposing approaches.     

The HIV 5′ Gag Region Displays a Specific Nucleotide Bias Regulating Viral Splicing and Infectivity

Alternative splicing and the expression of intron-containing mRNAs is one hallmark of HIV gene expression. To facilitate the otherwise hampered nuclear export of non-fully processed mRNAs, HIV encodes the Rev protein, which recognizes its intronic response element and fuels the HIV RNAs into the CRM-1-dependent nuclear protein export pathway. Both alternative splicing and Rev-dependency are regulated by the primary HIV RNA sequence. Here, we show that these processes are extremely sensitive to sequence alterations in the 5’coding region of the HIV genomic RNA. Increasing the GC content by insertion of either GFP or silent mutations activates a cryptic splice donor site in gag, entirely deregulates the viral splicing pattern, and lowers infectivity. Interestingly, an adaptation of the inserted GFP sequence toward an HIV-like nucleotide bias reversed these phenotypes completely. Of note, the adaptation yielded completely different primary sequences although encoding the same amino acids. Thus, the phenotypes solely depend on the nucleotide composition of the two GFP versions. This is a strong indication of an HIV-specific mRNP code in the 5′ gag region wherein the primary RNA sequence bias creates motifs for RNA-binding proteins and controls the fate of the HIV-RNA in terms of viral gene expression and infectivity.     

Linking the Electrical Conductivity and Non-Stoichiometry of Thin Film Ce1−xZrxO2−δ by a Resonant Nanobalance Approach

Bulk ceria-zirconia solid solutions (Ce_1−xZr_xO_2−δ, CZO) are highly suited for application as oxygen storage materials in automotive three-way catalytic converters (TWC) due to the high levels of achievable oxygen non-stoichiometry δ. In thin film CZO, the oxygen storage properties are expected to be further enhanced. The present study addresses this aspect. CZO thin films with 0 ≤ x ≤ 1 were investigated. A unique nano-thermogravimetric method for thin films that is based on the resonant nanobalance approach for high-temperature characterization of oxygen non-stoichiometry in CZO was implemented. The high-temperature electrical conductivity and the non-stoichiometry δ of CZO were measured under oxygen partial pressures pO₂ in the range of 10⁻²⁴–0.2 bar. Markedly enhanced reducibility and electronic conductivity of CeO₂-ZrO₂ as compared to CeO_2−δ and ZrO₂ were observed. A comparison of temperature- and pO₂-dependences of the non-stoichiometry of thin films with literature data for bulk Ce_1−xZr_xO_2−δ shows enhanced reducibility in the former. The maximum conductivity was found for Ce_0.8Zr_0.2O_2−δ, whereas Ce_0.5Zr_0.5O_2-δ showed the highest non-stoichiometry, yielding δ = 0.16 at 900 °C and pO₂ of 10⁻¹⁴ bar. The defect interactions in Ce_1−xZr_xO_2−δ are analyzed in the framework of defect models for ceria and zirconia.     

Pulmonary vein isolation using second-generation single-shot devices: not all the same?

Introduction

Single-shot devices have been developed to simplify pulmonary vein isolation (PVI). Randomized studies of the second-generation cryoballoon (CB 2nd) demonstrated excellent results. There are limited data comparing results of circular pulmonary vein ablation catheter (PVAC) with conventional RF ablation or CB for PVI.

Objective

Using a sequential registry cohort and a prospective randomized study, we aimed to compare the acute and long-term results of CB 2nd and PVAC Gold.

Methods

In the registry, consecutive patients with paroxysmal atrial fibrillation (AF) undergoing their first PVI were included. The preferred method used was PVAC Gold in 2014 and CB 2nd in 2015. Subsequently, a randomized study (PVAC vs. CB 2nd) was performed. Ablation success was measured as freedom of AF or atrial tachycardias (AT) off antiarrhythmic drugs.

Results

In the registry cohort, PVAC Gold was used in 60 patients and CB 2nd in 56 patients (age 66?±?11 years, 52% male, LAD 43?±?6). In the randomized study, 20 patients were treated with PVAC Gold and 22 with CB 2nd (age 67?±?9; 43% men, LAD 40?±?7 mm). During a mean follow up of 13.2?±?3.6 months, success was 54% in PVAC Gold patients and 81% in CB 2nd cases (p?=?0.001). In the randomized study 12 months success was 50% versus 86%, p?<?0.05. Complications occurred rare in both groups.

Conclusions

Our registry data and the randomized study both suggest superiority of PVI using CB 2nd as compared with PVI using PVAC Gold.

     

“Device landing zone” calcification,assessed by MSCT,as a predictive factor for pacemaker implantation after TAVI

Background : After trans‐catheter aortic valve implantation (TAVI), the need for postinterventional pacemaker (PM) implantation can occur in as many as 10–50% of cases, but it is not yet clear, how this need can be predicted. The aim of this study was to assess the possible predictive factors of post TAVI PM implantation based on Computed Tomography (CT) measured aortic valve calcification and its distribution. Methods : We prospectively analyzed 81 consecutive symptomatic patients with severe AS scheduled for TAVI using the CoreValve prosthesis (Medtronic, Minneapolis, USA). In all patients, a native and contrast‐enhanced multislice cardiac CT was performed preinterventionally, estimating calcification load of the native valve cusps and of the adjacent outflow tract (so called “device landing zone”, DLZ) by the Agatston Score (AgS). Objective, computer‐evaluated, preprocedural ECG‐analysis was performed with regards to pre‐existing conduction abnormalities. Transthoracic echocardiography was performed pre and post TAVI. Results : TAVI was successful in all cases. PM implantation was deemed necessary in altogether 32 patients, out of 67 without a PM pre‐TAVI (32/67, 47%). Various parameters were tested as predictors of post TAVI PM in a multivariate logistic regression analysis model. Female sex (P = 0,005) and depressed EF (P = 0,023) showed a significant correlation. PM implantation correlated also to the DLZ calcification, as assessed by CT (P = 0,004). This model leads to an AUC (area under the ROC—receiver operator characteristics—curve) of 0.83. Conclusion : Calcium amount in the CoreValve DLZ in combination with clinical data could predict the need for post TAVI PM implantation. © 2010 Wiley‐Liss, Inc.     

Detection of recombinant and endogenous mouse melatonin receptors by monoclonal antibodies targeting the C‐terminal domain

Melatonin receptors play important roles in the regulation of circadian and seasonal rhythms, sleep, retinal functions, the immune system, depression, and type 2 diabetes development. Melatonin receptors are approved drug targets for insomnia, non‐24‐hour sleep‐wake disorders, and major depressive disorders. In mammals, two melatonin receptors (MTRs) exist, MT₁ and MT₂, belonging to the G protein‐coupled receptor (GPCR) superfamily. Similar to most other GPCRs, reliable antibodies recognizing melatonin receptors proved to be difficult to obtain. Here, we describe the development of the first monoclonal antibodies (mABs) for mouse MT₁ and MT₂. Purified antibodies were extensively characterized for specific reactivity with mouse, rat, and human MT₁ and MT₂ by Western blot, immunoprecipitation, immunofluorescence, and proximity ligation assay. Several mABs were specific for either mouse MT₁ or MT₂. None of the mABs cross‐reacted with rat MTRs, and some were able to react with human MTRs. The specificity of the selected mABs was validated by immunofluorescence microscopy in three established locations (retina, suprachiasmatic nuclei, pituitary gland) for MTR expression in mice using MTR‐KO mice as control. MT₂ expression was not detected in mouse insulinoma MIN6 cells or pancreatic beta‐cells. Collectively, we report the first monoclonal antibodies recognizing recombinant and native mouse melatonin receptors that will be valuable tools for future studies.