Detection of metallo-beta-lactamase producing Pseudomonas aeruginosa using a modified IMP-lysate assay

Since the increasing prevalence of carbapenem-resistant Pseudomonas aeruginosa spp., accurate detection of metallo-β-lactamase (MBL) such as blaVIM type enzyme producing isolates became very important. However, phenotypic MBL detection methods previously reported are not highly sensitive or highly specific. This study aimed to evaluate the performance of a modified IMP-lysate test, the doubledisk- synergy-test (DDST) and the combined-disk-test (CDT) for detecting MBL blaVIM gene in P. aeruginosa. The reference technique was PCR molecular test. The study used 12 blaVIM2 producer isolates, 13 MBL-negative controls which included 4 imipenem-susceptible strains and 9 imipenemresistant strains harbouring blaSHV-2a genes collected from two Tunisian hospitals and P. aeruginosa ATCC27853 and P. aeruginosa COL-1 as negative and positive controls respectively. CDT showed 100% of sensitivity. The highest level of specificity was shown by IMP-lysate test (76.92%). To evaluate efficiencies of methods, the study noted that the highest Youden Index (YI) was shown by IMP-lysate method (0.7), followed by DDST (0.6) than CDT (0.2). Since MBL-Etest and PCR were expensive and not adaptable for extension use in clinical microbiology laboratories, a modified IMP-lysate MBL hydrolytic activity can be chosen by laboratories with limited resources as an inexpensive, simple, and accurate test to detect . P. aeruginosa producing blaVIM enzyme.Key words: Metallo-beta-lactamase, VIM, phenotypic detection, pseudomonas aeruginosa

Biological activities of Peganum harmala leaves

Ethyl acetate, chloroform, butanol and methanol extracts of the leaves of Peganum harmala were tested for antibacterial, antioxidant and antiviral activities. The antibacterial activity was evaluated by the determination of minimum inhibitory concentration (MIC) using the solid medium technique. The antiviral activity was evaluated against human cytomegalovirus (HCMV) strain AD-169 (ATCC Ref. VR 538) and Coxsackie B virus type 3 (CoxB-3) using diagnostic method ‘shell-vial’ culture. The antioxidant activity was evaluated using ammonium thiocyanate method. Among tested extracts, methanol and chloroform extracts displayed a higher antibacterial activity against gram-positive than gram-negative bacteria. The methanol extract demonstrated the highest antioxidant activity and good antiviral activity against HCMV.Key words: Peganum harmala, antimicrobial, antiviral, antioxidant, activities, leave

Induction and flow cytometry identification of mixoploidy through colchicine treatment of Trigonella foenum-graecum L.

Polyploidy plays an important role in plant evolution and constitutes an important mechanism of diversification and genetic variations creation. The objective of this study was to produce a polyploid cultivar of Trigonella foenum-graecum L. (2n=16) and to evaluate its cytological potentialities. Polyploidy induction was carried out by using a 0.05% colchicine solution. Ploidy level was determined by an analysis of flow cytometry, which is suitable, quick and easy for the identification of ploidy level. Quantification of DNA by spectrophotometric methods showed that treated plants presented higher contents of cellular DNA than diploid plants, and this quantification was checked by fluorescence method and comparison of the DNA profiles showed a more significant thickness of the DNA band in the extract from the treated plants than the diploid plants. In addition, the tested plants were classed as diploid and mixoploid by flow cytometry. The cytological characteristics such as the stomata size and pollen grain diameter of the mixoploid were significantly larger than those of the diploid. Polyploidy induction is an effective method to increase plant performance.Key words: Fenugreek, colchicine, flow cytometry, polyploidy, mixoploidy

Molecular characterization of enterovirus detected in cerebrospinal fluid and wastewater samples in Monastir, Tunisia, 2014-2017

Background: Enteroviruses (EVs) are considered the main causative agents responsible for aseptic meningitis worldwide. This study was conducted in the Monastir region of Tunisia in order to know the prevalence of EV infections in children with meningitis symptoms. Detected EV types were compared to those identified in wastewater samples. Methods: Two hundred CSF samples collected from hospitalized patients suspected of having aseptic meningitis for an EV infection between May 2014 and May 2017 and 80 wastewater samples collected in the same time-period were analyzed. EV detection and genotyping were performed using PCR methods followed by sequencing. Phylogenetic analyses in the 3'-VP1 region were also carried-out. Results: EVs were detected in 12% (24/200) CSF and in 35% (28/80) wastewater samples. EV genotyping was reached in 50% (12/24) CSF-positive samples and in 64% (18/28) sewage. Most frequent types detected in CSF were CVB3, E-30 and E-9 (25% each). In wastewater samples, the same EVs were identified, but also other types non-detected in CSF samples, such as E-17,CVA9 and CVB1 from EV species B, and EV-A71 and CVA8 from EV-A, suggesting their likely lower pathogenicity. Phylogenetic analysis showed that within the same type, different strains circulate in Tunisia. For some of the EV types such as E-9, E-11 or CVB3, the same strains were detected in CSF and wastewater samples. Conclusions: Epidemiological studies are important for the surveillance of the EV infections and to better understand the emergence of certain types and variants.Y. Rmadi was supported by the grant of the Ministry of Higher Education and Research of Tunisia.S

Cytotoxic effect of nanoparticles synthesized from Salvia officinalis L. and Ricinus communis aqueous extracts against vero cell line and evaluation of their antioxidant activities

The development of safe carriers for the use of plant extracts in industrial and health fields constitute a matter of serious concern. The development of plant derived nanoparticles may help to overcome such barriers. However, the major concern is still the safety of these carriers. The present study describes the synthesis of nanoparticles derived from Salvia officinalis L. and Ricinus communis and the evaluation of their cytotoxic and antioxidant effects. It is shown in this study that the aqueous extracts of S. officinalis L. and R. communis have the potentials to reduce silver nitrate ions to silver nanoparticles. The biosynthesized nanoparticles were analyzed by spectrophotometer and granulometric tests. The biological activities of these extracts and nanoparticles were carried out. S. officinalis L. leaf extract had the strongest antioxidant activity, followed by R. communis leaf and fruit extracts. Altogether, the synthesized nanoparticles were safe and may serve as antioxidant products in many fields.Key words: Plant material, biosynthesis of nanoparticle, cytotoxicity, cell culture, free radicals

Sudden unexpected death related to enterovirus myocarditis: histopathology, immunohistochemistry and molecular pathology diagnosis at post-mortem

BACKGROUND: Viral myocarditis is a major cause of sudden unexpected death in children and young adults. Until recently, coxsackievirus B3 (CVB3) has been the most commonly implicated virus in myocarditis. At present, no standard diagnosis is generally accepted due to the insensitivity of traditional diagnostic tests. This has prompted health professionals to seek new diagnostic approaches, which resulted in the emergence of new molecular pathological tests and a more detailed immunohistochemical and histopathological analysis. When supplemented with immunohistochemistry and molecular pathology, conventional histopathology may provide important clues regarding myocarditis underlying etiology. METHODS: This study is based on post-mortem samples from sudden unexpected death victims and controls who were investigated prospectively. Immunohistochemical investigations for the detection of the enteroviral capsid protein VP1 and the characterization and quantification of myocardial inflammatory reactions as well as molecular pathological methods for enteroviral genome detection were performed. RESULTS: Overall, 48 sudden unexpected death victims were enrolled. As for controls, 37 cases of unnatural traffic accident victims were studied. Enterovirus was detected in 6 sudden unexpected death cases (12.5 %). The control samples were completely enterovirus negative. Furthermore, the enteroviral capsid protein VP1 in the myocardium was detected in enterovirus-positive cases revealed by means of reverse transcriptase-polymerase chain reaction (RT-PCR). Unlike control samples, immunohistochemical investigations showed a significant presence of T and B lymphocytes in sudden unexpected death victims. CONCLUSIONS: Our findings demonstrate clearly a higher prevalence of viral myocarditis in cases of sudden unexpected death compared to control subjects, suggesting that coxsackie B enterovirus may contribute to myocarditis pathogenesis significantly

Multidrug-resistant Acinetobacter baumannii strains carrying the blaOxA-23 and the blaGES-11 genes in a neonatology center in Tunisia

Multidrug-resistant and difficult-to-treat Acinetobacter baumannii may be responsible for nosocomial infections. The production of carbapenem-hydrolyzing class D β-lactamases (CHDLs) and extended-spectrum β-lactamase (ESBLs) of the GES type possessing a carbapenemase activity has been increasingly reported worldwide in A. baumannii. The aim of this study was to analyze the resistance mechanisms of two carbapenem resistant A. baumannii clinical isolates recovered in a neonatology center in the center-east of Tunisia.Two carbapenem resistant A. baumannii isolates were recovered. The first isolate co-harbored the blaGES-11 ESBL gene and the blaOxA-23 CHDL gene. Analyses of the genetic location indicated that the blaGES-11 gene was plasmid located (Gr6). However, the blaOxA-23 gene was located on the chromosome. The second strain had only the blaOxA-23 CHDL gene, which was plasmid located.This study showed the first description of the GES-type β-lactamase in A. baumannii in Tunisia

Cosavirus, Salivirus and Bufavirus in Diarrheal Tunisian Infants

International audienceThree newly discovered viruses have been recently described in diarrheal patients: Cosa-virus (CosV) and Salivirus (SalV), two picornaviruses, and Bufavirus (BuV), a parvovirus. The detection rate and the role of these viruses remain to be established in acute gastroen-teritis (AGE) in diarrheal Tunisian infants. From October 2010 through March 2012, stool samples were collected from 203 children <5 years-old suffering from AGE and attending the Children's Hospital in Monastir, Tunisia. All samples were screened for CosV, SalV and BuV as well as for norovirus (NoV) and group A rotavirus (RVA) by molecular biology. Positive samples for the three screened viruses were also tested for astrovirus, sapovirus, ade-novirus, and Aichi virus, then genotyped when technically feasible. During the study period, 11 (5.4%) samples were positive for one of the three investigated viruses: 2 (1.0%) CosV-A10, 7 (3.5%) SalV-A1 and 2 (1.0%) BuV-1, whereas 71 (35.0%) children were infected with NoV and 50 (24.6%) with RVA. No mixed infections involving the three viruses were found, but multiple infections with up to 4 classic enteric viruses were found in all cases. Although these viruses are suspected to be responsible for AGE in children, our data showed that this association was uncertain since all infected children also presented infections with several enteric viruses, suggesting here potential water-borne transmission. Therefore, further studies with large cohorts of healthy and diarrheal children will be needed to evaluate their clinical role in AGE