Detection of AmpC and ESBL-producing Enterobacterales isolated from urinary tract infections in Tunisia

Urinary tract infections (UTIs) are the most frequent human infections in community and hospitals. This study aimed to determine the distribution of bacterial uropathogens among urinary tract infections diagnosed within the regional hospital Houcine Bouzaiene (Gafsa, South West Tunisia) during a survey of 54 days from the 8th of November to the 31st of December 2017. Enterobacterales strains were tested for antimicrobial resistance by disk diffusion method and extended-spectrum beta-lactamase (ESBL) production was tested by double-disc synergy test. Strains were further subjected to a molecular assessment of ESBL and AmpC beta-lactamase production by PCR. Overall, 173 bacterial isolates were studied, out of which 91.3% were Enterobacterales. Escherichia coli was the dominant pathogen, followed by Klebsiella pneumoniae. High to moderate resistance rates were observed, ranging from 66% to 90.7% for penicillin, from 6.7% to 18.6% for cephalosporins and from 16.2% to 25.4% for fluoroquinolones. Enterobacterales with decreased susceptibility to third-generation cephalosporins (3rd GC) carried several resistance genes: bIaCTX-M group 1 and group 9, and ACC and FOX AmpC beta-lactamase genes. Overall, ESBLs and AmpC beta-lactamases were detected in 57% and 14% of the 3rd GC-resistant isolates, respectively. This study proved the high potential of K. pneumaniae species to develop resistance against commonly used antibiotics. Thus, rigorous monitoring of the antibiotic resistance of clinical pathogens have to be implemented in Tunisia. Our results are very relevant to evaluate efficiency of the Tunisian therapeutic strategies against UTIs and adapt them to the emerging problem of antimicrobial resistance

Mutations in phospholipase C eta-1 ( PLCH1 ) are associated with holoprosencephaly

Funder: NIHR Cambridge Biomedical Research centreBackground: Holoprosencephaly is a spectrum of developmental disorder of the embryonic forebrain in which there is failed or incomplete separation of the prosencephalon into two cerebral hemispheres. To date, dominant mutations in sonic hedgehog (SHH) pathway genes are the predominant Mendelian causes, and have marked interfamilial and intrafamilial phenotypical variabilities. Methods: We describe two families in which offspring had holoprosencephaly spectrum and homozygous predicted-deleterious variants in phospholipase C eta-1 (PLCH1). Immunocytochemistry was used to examine the expression pattern of PLCH1 in human embryos. We used SHH as a marker of developmental stage and of early embryonic anatomy. Results: In the first family, two siblings had congenital hydrocephalus, significant developmental delay and a monoventricle or fused thalami with a homozygous PLCH1 c.2065C>T, p.(Arg689*) variant. In the second family, two siblings had alobar holoprosencephaly and cyclopia with a homozygous PLCH1 c.4235delA, p.(Cys1079ValfsTer16) variant. All parents were healthy carriers, with no holoprosencephaly spectrum features. We found that the subcellular localisation of PLCH1 is cytoplasmic, but the p.(Cys1079ValfsTer16) variant was predominantly nuclear. Human embryo immunohistochemistry showed PLCH1 to be expressed in the notorcord, developing spinal cord (in a ventral to dorsal gradient), dorsal root ganglia, cerebellum and dermatomyosome, all tissues producing or responding to SHH. Furthermore, the embryonic subcellular localisation of PLCH1 was exclusively cytoplasmic, supporting protein mislocalisation contributing to the pathogenicity of the p.(Cys1079ValfsTer16) variant. Conclusion: Our data support the contention that PLCH1 has a role in prenatal mammalian neurodevelopment, and deleterious variants cause a clinically variable holoprosencephaly spectrum phenotype

Mutations in phospholipase C eta-1 ( PLCH1 ) are associated with holoprosencephaly

Funder: NIHR Cambridge Biomedical Research centreBackground: Holoprosencephaly is a spectrum of developmental disorder of the embryonic forebrain in which there is failed or incomplete separation of the prosencephalon into two cerebral hemispheres. To date, dominant mutations in sonic hedgehog (SHH) pathway genes are the predominant Mendelian causes, and have marked interfamilial and intrafamilial phenotypical variabilities. Methods: We describe two families in which offspring had holoprosencephaly spectrum and homozygous predicted-deleterious variants in phospholipase C eta-1 (PLCH1). Immunocytochemistry was used to examine the expression pattern of PLCH1 in human embryos. We used SHH as a marker of developmental stage and of early embryonic anatomy. Results: In the first family, two siblings had congenital hydrocephalus, significant developmental delay and a monoventricle or fused thalami with a homozygous PLCH1 c.2065C>T, p.(Arg689*) variant. In the second family, two siblings had alobar holoprosencephaly and cyclopia with a homozygous PLCH1 c.4235delA, p.(Cys1079ValfsTer16) variant. All parents were healthy carriers, with no holoprosencephaly spectrum features. We found that the subcellular localisation of PLCH1 is cytoplasmic, but the p.(Cys1079ValfsTer16) variant was predominantly nuclear. Human embryo immunohistochemistry showed PLCH1 to be expressed in the notorcord, developing spinal cord (in a ventral to dorsal gradient), dorsal root ganglia, cerebellum and dermatomyosome, all tissues producing or responding to SHH. Furthermore, the embryonic subcellular localisation of PLCH1 was exclusively cytoplasmic, supporting protein mislocalisation contributing to the pathogenicity of the p.(Cys1079ValfsTer16) variant. Conclusion: Our data support the contention that PLCH1 has a role in prenatal mammalian neurodevelopment, and deleterious variants cause a clinically variable holoprosencephaly spectrum phenotype

Développement de biocapteurs électrochimiques fonctionnalisés avec des empreintes moléculaires pour la détection de protéines

This work aims to design selective electrochemical biosensors based on molecularly imprinted polymers (MIP) for protein detection. The first part is carried to polydopamine-MIP elaboration, for a model protein ''immunoglobulin G (IgG)'' detection. Two different strategies are investigated: the first is based on the dopamine electropolymerization with the free protein and the second consists of protein covalently grafting on the transducer surface previously functionalized by self-assembled monolayers (SAMs). The electrochemical results as well as the mathematical modelling have shown a higher impression rate with a low dissociation constant for the second strategy (IF2 = 80%, Kd2 = 3.14 ± 0.43 10-11 M) compared to the first one (IF1 = 50 %, Kd1 = 1.65 ± 0.45 x 10-10 M). The obtained results confirm that covalent protein grafting to the electrode surface could generate more homogeneous and accessible surface cavities for protein recognition, improving then the sensor metrological performance.The second approach was then adopted to study the interactions between others three proteins (having different molecular weights and isoelectric points) with polydopamine MIP in the presence and absence of the redox probe [Fe (CN)6] 3-/4-. These are bovine serum albumin (BSA), human serum albumin (HSA) and prostate specific antigen (PSA). The modeling of electrochemical measurements showed that the presence of the redox probe considerably reduces the interaction between the protein and its imprint. In addition, the MIP – protein interaction depends on the intrinsic properties of the protein. Indeed, due to its low molecular weight and its richness in positively charged groups, PSA is more cross-linked than BSA and HAS within the polydopamine-based MIP. To better understand the interaction mechanism between the protein and its MIP, the effect of pH and ionic strength was studied on the MIP ’s design and the further recognition process of the analyte by its imprints. The selected protein for this study is the BSA. The combination of electrochemical results, zeta potential measurements and infrared spectroscopy with Fourier transform analyzes have shown that an ionic strength of 167 mM and a pH of 7.4, allow having a better protein structural stability and a high electroactivity of the polymeric matrix in polydopamine, ameliorating thus the recognition of the protein by its MIP.In conclusion, this work allowed us to optimize the conditions for designing of a dopamine-based molecularly imprinted polymer (MIP) for the detection of proteins by electrochemical measurements. This MIP-based biosensor has good selectivity, high specificity and low detection limit. The adopted approach can be applied to the quantification of other proteins such as viral proteins, bacterial proteins, immunoglobulins as well as protein biomarkers of many pathologies. The integration of microfluidic system with this MIP-based biosensor could constitute an analytical tool performing a rapid and in-situ diagnosis in several fields including biomedical, food and environmental.Les travaux de cette thèse visent la conception de biocapteurs électrochimiques sélectifs à base de polymère à empreintes moléculaires (MIP) pour la détection des protéines. Dans la première partie, une empreinte moléculaire à base de dopamine a été élaborée pour la détection d’une immunoglobuline G (IgG), considérée comme protéine modèle. Deux différentes stratégies ont été testées : la première est basée sur l’électropolymérisation de la dopamine en présence de la protéine libre et la seconde consiste à greffer la protéine d’une manière covalente sur la surface de l’électrode préalablement fonctionnalisée par des monocouches autoassemblées (SAMs). Les résultats des mesures électrochimiques ainsi que les modélisations mathématiques ont montré un taux d’impression élevé et une faible constante de dissociation pour la seconde stratégie (IF2 = 80%, Kd2 = 3.14 ± 0.43 10-11 M) par rapport aux valeurs de la première stratégie (IF1 = 50 %, Kd1 = 1.65 ± 0.45 x 10-10 M). Les résultats obtenus confirment que le greffage covalent de la protéine engendre la création de cavités surfaciques homogènes, plus accessibles pour la protéine, améliorant ainsi les performances métrologiques de ce capteur. La second approche a ensuite été adoptée pour étudier les interactions de trois types de protéines avec l’empreinte de polydopamine en présence et en absence du couple redox [Fe(CN)6]3-/4-. Il s’agit de l’albumine du sérum bovin (BSA), de l’albumine du sérum humain (HSA) et de l’antigène spécifique de la prostate (PSA) ayant des poids moléculaires et des points isoélectriques différents. La modélisation des résultats des mesures électrochimiques a montré que la présence du couple redox diminue de façon considérable l’interaction entre la protéine et son empreinte. Cette interaction dépend également des propriétés intrinsèques de la protéine. En effet, en raison de son faible poids moléculaire et de sa richesse en groupements chargés positivement, le PSA est plus réticulé que la BSA et la HSA au sein de l’empreinte de polydopamine.Pour mieux comprendre le mécanisme d’interaction entre une protéine et son empreinte, l’effet du pH et de la force ionique a été étudié sur la réalisation de l’empreinte et la détection d’une protéine modèle, la BSA. Les résultats électrochimiques, les mesures de potentiel zêta et les analyses par spectroscopie infra-rouge à transformée de Fourier ont montré qu’une force ionique de 167 mM et un pH de 7,4, permettent d’avoir une meilleure stabilité structurelle de la protéine et une électroactivité élevée de la matrice polymérique en polydopamine, améliorant ainsi la reconnaissance de la protéine par ses empreintes moléculaires.En conclusion, ce travail nous a permis d’optimiser les conditions d’élaboration d’une empreinte moléculaire à base de dopamine pour détecter les protéines par des mesures électrochimiques. Ce biocapteur à base de MIP présente une bonne sélectivité, une grande spécificité et une faible limite de détection. L’approche adoptée peut être appliquée à la détection d’autres protéines telles que les protéines virales, les protéines bactériennes, les immunoglobulines ainsi que les biomarqueurs protéiques de nombreuses pathologies. L’intégration d’un système microfluidique à ce biocapteur à base d’empreintes moléculaires pourrait constituer un outil analytique capable d’effectuer un diagnostic rapide et in-situ dans plusieurs domaines dont le biomédical, l’agroalimentaire et l’environnemental

Deep Multi-modal Object Detection for Autonomous Driving

International audienc

Design and efficiency enhancement of FTO/PC60BM/CsSn0.5Ge0.5I3/Spiro-OMeTAD/Au perovskite solar cell utilizing SCAPS-1D Simulator

The poisoning potential of lead, which is the main component of the absorber layer of lead halide (Pb) perovskites, as well as the stability problems of the manufactured devices, constitute a major obstacle to the industrialization of this technology. As a result, recent research is concentrating on lead-free metal halide perovskites. Unfortunately, current lead-free perovskites suffer from poor performance, hence the interest of our study. The research presented here shows that optimizing several variables related to the performance of each layer of a perovskite solar cell (PSC) constructed from lead-free inorganic materials provides an efficiency of 18.13%. We designed a structure with outstanding performance using the FTO/PC _60 BM/CsSn _0.5 Ge _0.5 I _3 /Spiro-OMeTAD/Au configuration. The impact of various relevant factors, such as the thickness and defect density of the absorber layer their doping densities, the back contact work, and the operating temperature, have been thoroughly investigated to boost the performance of the proposed device. The performance of cesium-tin-germanium triiodide (CsSn _0.5 Ge _0.5 I _3 ) solar cells with different electron transport materials, including ZnO, TiO _2 , CdS, C _60 ; Cd _0.5 Zn _0.5 S, IGZO, has also been examined. It has been demonstrated that using ZnO as an electron transport layer improves electron extraction and, therefore, performance. The best outcomes are obtained after optimizing all the factors mentioned above, namely: Jsc of 28.70 mA/cm ^2 , Voc of 1.115 V, FF of 87.86%, and PCE of 18.13%. Additionally, the explored structure may be an excellent candidate for the future development of lead-free perovskite solar cells

Contribution to the Understanding of the Interaction between a Polydopamine Molecular Imprint and a Protein Model: Ionic Strength and pH Effect Investigation

Several studies were devoted to the design of molecularly imprinted polymer (MIP)-based sensors for the detection of a given protein. Here, we bring elements that could contribute to the understanding of the interaction mechanism involved in the recognition of a protein by an imprint. For this purpose, a polydopamine (PDA)-MIP was designed for bovine serum albumin (BSA) recognition. Prior to BSA grafting, the gold surfaces were functionalized with mixed self-assembled monolayers of (MUDA)/(MHOH) (1/9, v/v). The MIP was then elaborated by dopamine electropolymerization and further extraction of BSA templates by incubating the electrode in proteinase K solution. Three complementary techniques, electrochemistry, zetametry, and Fourier-transform infrared spectrometry, were used to investigate pH and ionic strength effects on a MIP’s design and the further recognition process of the analytes by the imprints. Several MIPs were thus designed in acidic, neutral, and basic media and at various ionic strength values. Results indicate that the most appropriate conditions, to achieve a successful MIPs, were an ionic strength of 167 mM and a pH of 7.4. Sensitivity and dissociation constant of the designed sensor were of order of (3.36 ± 0.13) µA·cm−2·mg−1·mL and (8.56 ± 6.09) × 10−11 mg/mL, respectively

Novel pathogenic mutations and further evidence for clinical relevance of genes and variants causing hearing impairment in Tunisian population

Introduction: Hearing impairment (HI) is characterized by complex genetic heterogeneity. The evolution of next generation sequencing, including targeted enrichment panels, has revolutionized HI diagnosis. Objectives: In this study, we investigated genetic causes in 22 individuals with non-GJB2 HI. Methods: We customized a HaloplexHS kit to include 30 genes known to be associated with autosomal recessive nonsyndromic HI (ARNSHI) and Usher syndrome in North Africa. Results: In accordance with the ACMG/AMP guidelines, we report 11 pathogenic variants; as follows; five novel variants including three missense (ESRRB-Tyr295Cys, MYO15A-Phe2089Leu and MYO7A-Tyr560Cys) and two nonsense (USH1C-Gln122Ter and CIB2-Arg104Ter) mutations; two previously reported mutations (OTOF-Glu57Ter and PNPT1-Glu475Gly), but first time identified among Tunisian families; and four other identified mutations namely WHRN-Gly808AspfsX11, SLC22A4-Cys113Tyr and two MYO7A compound heterozygous splice site variants that were previously described in Tunisia. Pathogenic variants in WHRN and CIB2 genes, in patients with convincing phenotype ruling out retinitis pigmentosa, provide strong evidence supporting their association with ARNSHI. Moreover, we shed lights on the pathogenic implication of mutations in PNPT1 gene in auditory function providing new evidence for its association with ARNSHI. Lack of segregation of a previously identified causal mutation OTOA-Val603Phe further supports its classification as variant of unknown significance. Our study reports absence of otoacoustic emission in subjects using bilateral hearing aids for several years indicating the importance of screening genetic alteration in OTOF gene for proper management of those patients. Conclusion: In conclusion, our findings do not only expand the spectrum of HI mutations in Tunisian patients, but also improve our knowledge about clinical relevance of HI causing genes and variants