Prospection archéologique diachronique : l'occupation du sol sur le canton de Coutras (Gironde). Un état de la recherche

La prospection diachronique réalisée sur le canton de Coutras (Nord-Est de la Gironde en bordure des Charentes et de la Dordogne) au cours de l'année 2016 s'est articulée autour de deux objectifs. Le premier a été de mettre à jour et faire un état des recherches archéologiques menées sur le secteur par le passé (notamment par Dany Barraud). Le second a eu pour but d'alimenter la carte archéologique par la recherche de nouveaux indices de sites, sur un secteur au fort potentiel archéologique. Au terme des analyses issues de la prospection, nous constatons une forte représentation des périodes médiévale et moderne ; le matériel céramique récolté en surface des parcelles actuellement labourées témoigne d'un passé très agricole du secteur (encore très présent de nos jours). Hormis cela, aucun mobilier de l'époque gallo-romaine n'a été retrouvé : un fait très étonnant lorsque l'on tient compte du passé antique de Coutras, la station routière Corterate apparaissant sur la table de Peutinger. La prospection a toutefois permis d'identifier une occupation de la fin du Néolithique et de la protohistoire (Âge du Bronze et Âge du Fer) à la Prairie de Millet sur la commune de Coutras. La découverte de céramiques dégradées et de silex taillés correspondent à ces périodes. Ce type de mobilier a été retrouvé sur une grande surface (près de 80 000 m²) dans des proportions plus ou moins denses. Cet indice de site, n'apparaissant pas dans la carte archéologique, a faitl'objet d'une opération de sondage sur l'année 2017 dans le but de préciser le type d'occupation et d'évaluer son état de conservation.Le rapport de la prospection de 2016 a été remis au SRA de Bordeaux et consultable au centre de documentation de ce dernier

Long Term Exposure of Agricultural Soil to Veterinary Antibiotics Changes the Population Structure of Symbiotic Nitrogen-Fixing Bacteria Occupying Nodules of Soybeans (<em>Glycine max</em>)

International audienceAntibiotics can be entrained onto agricultural land through the application of animal manures, human biosolids, or recycled wastewater for irrigation. In order to evaluate the impacts on soil microorganisms of exposure to antibiotics, a series of replicated plots were initiated in 1999 at the AAFC research farm in London ON. Every spring a mixture of sulfamethazine, chlortetracycline and tylosin is incorporated directly into the soil to attain concentrations ranging from 0.1 to 10 mg/kg, and these are then seeded with soybeans. In the present study nodules were isolated from soybean plants growing in antibiotic treated and control soils in 2012, after 14 annual treatments. Rabbit polyclonal antisera distinguishing Bradyrhizobium japonicum serogroups 122, 6, 110 and 123 and B. liaoningense serogroup 135 was used for nodule strain typing. A total of 278 bradirhizobia were isolated and confirmed. Genomic DNA from isolates was subjected to RSα fingerprinting, sequencing of the 16S rDNA coding region, and the 16S rDNA- 23S rDNA intergenic spacer (IGS), and MLST analysis using six housekeeping genes. The sensitivity of isolates to the three antibiotics, singly and in combination, was evaluated. Serogroup 135 dominated nodules, with the other serotypes occasionally detected and many nodules being unreactive to the antisera. Based on genomic fingerprinting and MLST analysis the collection of 278 bradyrhizobia was very diverse. The distribution of isolates in RSa fingerprint groups was significantly different in the 3 soils treated with antibiotics compared to control soil, with an increase in proportion of strains belonging to RSa types a, b, c and q and a decrease of strains belonging to RSa types p and s. This result confirms those based on serotyping: strains belonging to B. liaoningense serogroup 135 are more abundant in soils having received antibiotics. Using RSα fingerprinting results, Shannon diversity indexes computed for control and antibiotic treated soils were equal to 1.11, 1.53, 1.43 and 1.3 for control soil, and soils having received low, intermediate and high antibiotic doses, respectively. T-tests revealed that Shannon diversity indexes were significantly different between control soil and soil having received the low (0.1 mg each drug/kg soil) dose of antibiotics (p-value=0.0045). Overall, the present study indicates that long term treatment with environmentally relevant concentrations of the veterinary antibiotics tylosin, chlortetracycline and sulfamethazine alters the composition of Bradyrhizobial populations occupying soybean nodules. The sensitivity of bradyrhizobia to the three antibiotics was not associated with the treatment from which they were recovered, indicating that variation in nodule occupancy was due to an indirect effect rather than direct antibiotic selection

Evaluation de la qualité microbiologique des eaux de baignade via la mesure ampérométrique d’activités enzymatiques

BIOMEEAUBEvaluation de la qualité microbiologique des eaux de baignade via la mesure ampérométrique d’activités enzymatiques. XV. Colloque du Groupe Français de Bioélectrochimi

Amperometric detection of extended-spectrum β-lactamase activity : application to the characterization of resistant <em>E.coli</em> strains

EA MERS CT3International audienceThe amperometric detection of extended-spectrum β-lactamase (ESBL) with carbon screen-printed sensors was investigated in the presence of the Nitrocefin, a commercially-available β-lactamase chromogenic cephalosporin substrate. Using an ESBL isolated from a clinical sample, it was shown for the first time that the intensity of a specific anodic pic current (EP = [similar]+0.3 V vs. Ag/AgCl) resulting from the catalytic hydrolysis of the β-lactam ring was proportional to the amount of ESBL. The proof-of-principle of a novel susceptibility assay for the rapid and accurate identification of ESBL- producing bacteria was then demonstrated. The detection scheme relied on (i) the culture of the sample in a medium containing the cefotaxime supplemented or not with the clavulanic acid inhibitor to allow the specific determination of ESBL producers (ii) followed by the incubation of the bacteria with the Nitrocefin and (iii) the measurement of the enzyme product by cyclic voltammetry. The amperometric assay was further applied to the characterization of E. coli strains and to the quantification of the ESBL producers. A detection limit of 5 × 104 cfu mL−1 ESBL-producing E. coli was achieved after a 10 min incubation time. In contrast to the approved routine assays, the electrochemical approach, which did not require isolated colonies to be performed, provided quantified results regarding ESBL activity within a few hours. Finally, owing to its cost-effectiveness, portability and simplicity, this test holds great promise for clinical and environmental applications

A nitrocefin-based amperometric assay for the rapid quantification of extended-spectrum β-lactamase-producing Escherichia coli in wastewaters

International audienceA sensitive and inexpensive amperometric assay based on the electrochemical detection of the beta-lactamase activity using the nitrocefin as substrate was developed for the rapid and quantitative detection of extended spectrum beta-lactamase-producing Escherichia colt (ESBL-EC) in urban wastewaters. The specific detection of ESBL-EC was achieved by culturing the filtered sample in a medium containing the cefotaxime supplemented or not with the potassium clavulanate inhibitor. This step was followed by the incubation of each subculture filtrate with the nitrocefin substrate which hydrolysis was monitored by amperometry using disposable carbon screen-printed sensors. Current intensities i(Cef) and i(Clav) correspond to the intensity of the anodic current measured (similar to+ 0.2 V vs. Ag/AgCl) for the sample incubated with the cefotaxime without and with potassium clavulanate, respectively. The intensity value i = i(Cef) iClav was chosen as the analytical response. ESBL-EC calibration plots were established with artificially contaminated wastewater samples. This assay allowed the detection of ESBL-EC amounts as low as 10 cfu in treated effluents and 100 cfu in raw wastewaters with short time analysis of 5.5 h and 4.5 h, respectively. The amperometric method was applied to the analysis of 38 wastewater samples and the results were in good agreement with CFU counts on a selective chromogenic medium for 24 h. Owing to its rapidity, convenience, low-cost and portability, this assay is a promising tool to obtain quantitative data on antimicrobial-resistant E. coli in wastewater effluents. Furthermore, this assay might be used to improve wastewater treatment plant processes in order to minimize the release of antibiotic resistant bacteria into the aquatic environment

A voltammetric test for the rapid discrimination of β-lactamase-producing <em>Enterobacteriaceae</em> in blood cultures

International audienceThe accurate identification of β-lactamases produced by Enterobacteriaceae is a major challenge in clinical laboratories in order to optimize antimicrobial treatment and patient care. We describe here a rapid voltammetric-based method to detect and to discriminate β-lactamase activity in Enterobacteriaceae i.e., penicillinase, cephalosporinase (inducible or overproduced), extended-spectrum beta-lactamase and carbapenemase producers. After a 2-h growth step of the sample under three separate conditions: 1) LB (Luria-Bertani) medium, 2) LB supplemented with 4 μg/mL cefotaxime and 3) LB supplemented with 4 μg/mL cefotaxime and 100 μg/mL potassium clavulanate, the β-lactamase activity was measured by incubating a 0.5 mM nitrocefin solution for 15 min followed by the voltammetric detection of the hydrolyzed nitrocefin with disposable carbon screen printed sensors. The development and the calibration of the method were carried out by analyzing pure cultures of fifty-seven strains with well characterized β-lactam-resistance phenotypes. Thanks to the combination of the three currents (i(1), i(2), i(3)) recorded for each tested bacteria, the proposed procedure allowed to distinguish the different classes of β-lactamase producers. In the second part of the study, the method was applied to the analysis of one hundred and fifteen samples Enterobacteriaceae-positive blood culture samples of bacteraemic patients. Overall data showed that the voltammetric method offered a sensitivity of 100% and a specificity of 80%. Interestingly, all of sixteen samples infected by a third-generation cephalosporins-resistant bacteria (i.e. ESBL and overproduced cephalosporinase producers) were detected. This study clearly demonstrated that the voltammetric assay is an efficient alternative technique for the rapid discrimination of β-lactamases-producing Enterobacteriaceae in blood culture. In contrast to the approved routine assays, the electrochemical test did not require isolated colonies to be performed and was thus carried out in less than 3 h which could allow early administration of an appropriate antibiotic therapy

A thin layer-based amperometric enzyme immunoassay for the rapid and sensitive diagnosis of respiratory syncytial virus infections

EA MERS CTInternational audienceA simple electrochemical sandwich immunoassay involving a polystyrene microarray slide coated with monoclonal capture antibodies and carbon screen-printed sensors (SPS) was designed for the rapid diagnosis of respiratory syncytial virus (RSV). The detection of the antibody-antigen complex formation relied on the use of a horseradish peroxidase conjugate. Its chronoamperometric measurement detection was performed by confining a droplet of H2O2/3.3',5,5'-tetramethylbenzidine enzyme substrate/mediator solution within a thin layer between one spot of the microarray and the surface of one screen-printed electrochemical cell. The accumulation of the enzyme product in the thin film of liquid enhanced the electrochemical response which allowed the development of a rapid (25 min) and sensitive thin layer-based amperometric (TLA) enzyme immunoassay. The method was successfully compared to commercially-available immunofluorescent and real-time PCR assays for RSV testing in respiratory secretion clinical samples. This suggests that owing to its rapidity, convenience, low-cost, portability and ability to provide quantified results, the reported concept could be a promising point-of-care diagnostic tool to screen patients with suspected respiratory infection or other types of infectious diseases

Fast non-contact surface roughness measurements up to the micrometer range by dual-wavelength digital holographic microscopy

We present fast high-roughness and non-contact surface measurements by digital holographic microscopy (DHM). By using single- and dual-wavelength operation modes, coupled with advanced image stitching and non-measured points management methods, the technique enables two-dimensional roughness measurements up to the micrometer (N6). The sample is mechanically scanned over a surface up to 5 × 0.3 mm2 with 17 holograms each acquired in less than 500 µs, the corresponding phase images stitched together by software, and therefore providing multiple profiles measurement in the ISO definition in less than 30 s. The approach is validated by inspection of several different roughness standards and our technique is demonstrated to be in agreement with two existing well-known techniques in the field

Occurence of ArmA and RmtB Aminoglycoside Resistance 16S rRNA Methylases in Extended-Spectrum β-Lactamases Producing Escherichia coli in Algerian Hospitals

EABIOMEThe aim of this study was to characterize the extended-spectrum-β-lactamases (ESBLs) producing clinical strains of Escherichia coli isolated between January 2009 and June 2012 from Algerian hospitals and to determine the prevalence of 16S rRNA methylase among them. Sixty-seven ESBL-producers were detected among the 239 isolates included: 52 CTX-M-15-producers, 5 CTX-M-3-producers, 5 CTX-M-1-producers, 2 CTX-M-14-producers, 2 SHV-12-producers and one TEM-167-producer. Among the ESBL-producing strains twelve harboured 16S rRNA methylase genes: 8 rmtB and 4 armA. rmtB was located on a IncFIA plasmid and armA was located either on a IncL/M or a IncFIA plasmid. RmtB-producing isolates were genotypically related and belonged to the sequence type ST 405 whereas ArmA-producing isolates belonged to ST10, ST 167 and ST 117. This first description of 16S rRNA methylases among E. coli in Algerian hospitals pointed out the necessity to establish control measures to avoid their dissemination

Rapid amperometric detection of <em>Escherichia coli</em> in wastewater by measuring beta-D glucuronidase activity with disposable carbon sensors

International audienceAn assay on the indirect amperometric quantification of the beta-D-Glucuronidase (GLUase) activity was developed for the rapid and specific detection of Escherichia coli (E. coli) in complex environmental samples. The p-aminophenyl beta-D-glucopyranoside (PAPG) was selected as an electrochemical substrate for GLUase measurement and the p-aminophenol (PAP) released during the enzymatic hydrolysis was monitored by cyclic voltammetry with disposable carbon screen-printed sensors. The intensity of the measured anodic peak current was proportional to the amount of GLUase, and therefore to the number of E. coli in the tested sample. Once the substrate concentration and pH values optimized, a GLUase detection limit of 10 ng mL(-1) was achieved. Using a procedure involving a filtration step of the bacteria followed by their incubation with the substrate solution containing both the nonionic detergent Triton X-100 as permeabilization agent and the culture media Luria broth to monitor the growth, filtered bacterial cells ranging from 5 x 10(4) to 10(8) UFC/membrane were detected within 3 h. The amperometric assay was applied to the determination of fecal contamination in raw and treated wastewater samples and it was successfully compared with conventional bacterial plating methods and uidA gene quantitative PCR. Owing to its ability to perform measurements in turbid media, the GLUase amperometric method is a reliable tool for the rapid and decentralized quantification of viable but also nonculturable E. coli in complex environmental samples