Impacts of the human pharmaceutical diclofenac in the aquatic environment

An increasing number of pharmaceuticals have been found in the aquatic environment and the issue has become a human and environmental health concern. Many pharmaceuticals are not fully degraded in wastewater treatment plants (WWTPs) and are continuously released in the aquatic environment resulting in concentrations in the low µg/l range in the receiving waters. Diclofenac is a widely used non-steroidal anti-inflammatory drug (NSAID) and is persistent in the aquatic environment. This pharmaceutical has been frequently reported in wastewater effluents, surface waters, groundwaters and even drinking water. NSAIDs are known to inhibit the cyclooxygenase activity, an enzyme present in many species of the animal kingdom responsible for the synthesis of prostanoids, and chronic exposure to environmental diclofenac may have detrimental effects on metabolism of non-target organisms including microbes and fish. In this thesis, microbiology, genomics and metabolomics approaches were used to investigate the effects of diclofenac on aquatic microbes and fish. In the first study of the thesis (chapter 3), the biodegradation of selected NSAIDs was investigated, together with their potential toxicity to aquatic microbes. Aerobic biodegradation experiments were conducted using activated sludge and wastewater effluents as microbial inocula and diclofenac, ketoprofen or naproxen as sole carbon source (1-10 mg/l) in order to isolate and identify the bacterial degraders. Changes in the bacterial populations were monitored by optical density and PCR-DGGE. The analytical techniques solid phase extraction (SPE) and ultraperformance liquid chromatography-mass spectrometry (UPLC-TOF-MS) were optimised to quantify the pharmaceuticals in environmental samples. High recovery rates were obtained with 94% for diclofenac; 92% for ketoprofen and 85% for naproxen and with detection capabilities down to 3-7 ng/l. Results from the biodegradation experiments showed that ketoprofen and naproxen were eliminated at up to 99 and 55% respectively over a 40 days period. Consistently with previous studies, diclofenac showed no significant degradation. In all the enrichments, a significant decrease in the bacterial abundance was observed as a consequence of NSAIDs exposure and attempts to isolate the bacterial degrading populations were unsuccessful. Given the apparent micro-toxicity of these NSAIDs, the standardised test Microtox@ was carried out with Vibrio fischeri. The EC50 (15 min) estimated ranged from 13.5 mg/l + 2.3 for diclofenac to 42.1 mg/l + 3.9 for naproxen. Further toxicological tests were performed with diclofenac on bacterial strains isolated from activated sludge. Growth inhibitory effects were observed from 50-70 mg/l for Micrococcus luteus, Zoogloea ramigera and Comamonas denitrificans. Pseudomonas putida seemed more tolerant to diclofenac exposure and toxic effects were observed from 90 mg/l. These studies showed that diclofenac was the most toxic NSAID but toxicological effects in bacteria only occurred at concentrations at least 1,000 times higher than those found in the environment. However, chronic exposure to lower concentrations may cause similar interferences and affect the degradation potential of naturally occurring microbial populations. The second study (chapter 4) investigated the biological effects of sub-chronic exposure to waterborne diclofenac (0.5, 1, 5 and 25 µg/l) in female juvenile rainbow trout Oncorhynchus mykiss. After 21-day exposure, mRNA expression levels of cytochrome p450 1a1 (cyp1a1), cyclooxygenase (cox) 1 and 2, and p53 were investigated in the liver, kidney and gills using RT-PCR and QPCR. These genes were selected as they are likely targets for diclofenac in mammals. Histopathological investigations were carried out in the small intestine, liver and kidney because diclofenac has been reported to induce toxicity responses in these tissues. Fish bile was also analysed by SPE and UPLC-TOF-MS to evaluate the bioconcentration potential of diclofenac and look for evidences of diclofenac metabolism. Results showed a significant reduction of both cox1 and cox2 expression in the liver, gills and kidney from 1 μg diclofenac/l. In contrast diclofenac induced an increase in mRNA levels for cyp1a1 in the liver and gills but a significant reduction of cyp1a1 expression in the kidney from 1 µg/l. There were no clear effects of diclofenac on the mRNA levels of p53. Diclofenac exposure caused tissue damages at exposure concentrations as low as 1 µg/l. Histopathological injuries included inflammation, hyperplasia and fusion of the villi in the small intestine and tubule necrosis in the kidney. There were no obvious changes in the liver of diclofenac-exposed fish. The analysis of bile revealed a bioconcentration potential between 509 + 27 and 657 + 25. A reactive metabolite of diclofenac was also detected at the highest exposure concentration which may be responsible for the severe injuries found in those fish. Sub-chronic exposure to environmental concentrations of diclofenac altered gene expression and it is possible that long term exposure to environmental diclofenac lead to significant impacts on fish health. In the final part of this thesis (chapters 5 and 6) effects on the metabolite composition of biofluids were analysed in diclofenac-exposed fish. This work entailed developing and validating appropriate methodologies to analyse fish bile and blood plasma. Methanol extraction and UPLC-TOF-MS were optimised to analyse the plasma metabolome but the methodologies were not suitable to detect low abundance molecules such as eicosanoids due to the interferences (ion suppression) in the samples matrix. Multivariate data analysis failed to detect the endogenous metabolites of the plasma affected by the chemical exposure. The only discriminating metabolite was found after analysis of the plasma samples from control vs. 25 µg/l treatment groups and identified as the exogenous compound diclofenac. To analyse the bile, the developed SPE methodology was carried out in order to separate the metabolites between a free steroids (fatty acids, eicosanoids, etc.) fraction and a conjugated steroids (bile salts) fraction. Due to high levels of taurocholic acid masking other metabolites in the conjugated fraction, some bile samples were hydrolysed to deconjugate these metabolites. The non-hydrolysed and hydrolysed bile fractions were analysed by UPLC-TOF-MS in positive and negative ionization. Multivariate data analysis using principal component analysis (PCA) and partial least square discriminant analysis (PLS-DA) revealed significant perturbations in the bile metabolite profile of diclofenac-exposed rainbow from the lowest exposure concentration (0.5 µg/l). Over 50 metabolites were elevated or reduced as a result of the 21-day exposure, suggesting that diclofenac affected several metabolic pathways. One metabolite was identified as a lipooxygenase product. This suggests that the inhibition of prostanoids synthesis can cause a shift in the arachidonic cascade and increase the synthesis of other eicosanoids. Most of the other discriminative metabolites remain unidentified and FT-MS analysis will be performed to obtain a structural identity. The metabolomics study further highlights the concern of environmental diclofenac in non-target organisms and the need to investigate the metabolic pathways affected

Efficacité comparée des insecticides de natures différentes dans la gestion des insectes ravageurs du niébé au Centre du Bénin

Objectif: Le niébé est une importante plante alimentaire dont la production et la valorisation sont d’une grande priorité pour l’amélioration de la sécurité alimentaire de la population. L’objectif de cette étude est de comparer en milieu paysan, l'efficacité du champignon entomopathogène Beauveria bassiana (isolat Bb 115), du mélange Neem (Topbio) - virus MaviMNPV et de l’insecticide chimique Decis (deltaméthrine) dans la gestion des insectes ravageurs du niébé.Méthodologie et résultats: L'essai a été conduit dans trois villages de la zone cotonnière du centre du Bénin (commune de Glazoué). C’est en effet, une zone de grande production du niébé. L’essai a été conduit suivant un dispositif de bloc aléatoire complet avec quatre traitements et trois répétitions par village de la zone. Les traitements comprennent 1) témoin (niébé non traité), 2) mélange Topbio - MaviMNPV, 3) B. bassiana et 4) Decis. L’application du champignon B. bassiana a été faite à la dose de 75 g m.a. (matière active)/ha. Le Décis et le TopBio à 1L /ha. Quant à la suspension virale, la dose de 2.109 OB/ha a été utilisée. Les effets des différents pesticides utilisés sur les densités de populations de M. vitrata, Megalurothrips sjostedti, Clavigralla tomentosicollis, que sur leur dégâts sur les organes reproductifs (boutons floraux, fleurs, gousses) ont été évalués. Des organes fructifères ont été prélevés hebdomadairement pour évaluer le nombre de larves mortes ou vivantes et aussi leurs dégâts sur les boutons floraux, fleurs et gousses. Les rendements en grains ont été également évalués. Bien qu’il y ait une grande variation dans les effets occasionnés par les biopesticides testés, ils ont considérablement réduit la densité de population de Maruca vitrata et des autres espèces d'insectes, Megalurothrips sjostedti et Clavigralla tomentosicollis ainsi que le niveau de dommages sur les organes fructifères. Comparativement au témoin, le rendement global en grains a été amélioré dans le niébé traité. Ainsi, à Glazoué, dans la zone centrale, des rendements meilleurs de 933,03±8,7Kg/ha ont été obtenus avec le champignon B. bassiana.Conclusion et applications des résultats: Ces résultats révèlent clairement que les biopesticides utilisés dans notre étude peuvent réduire significativement les densités des ravageurs ciblés. Cette étude suggère la possibilité de l'utilisation de biopesticides comme des alternatives aux insecticides chimiques dans la lutte contre les ravageurs du niébé.Mots clés : Niébé, insectes ravageurs, insecticide chimique, insecticide biologique, lutte intégré

Localisations primitives inhabituelles du kyste hydatique

L’Echinococcus granulosus, parasite appartenant à la famille des Cestodes est responsable, à travers sa forme larvaire, d’une maladie : l’hydatidose. Celle-ci se manifeste typiquement au niveau hépatique ou pulmonaire ; cependant une fois sur dix, l’embryon va se loger dans n’importe quel tissu ou organe du corps humain. Les auteurs rapportent quatre cas de localisation inhabituelle de l’hydatidose : rénale, musculaire, pancréatique et thyroïdienne

Applications for next-generation sequencing in fish ecotoxicogenomics

The new technologies for next-generation sequencing (NGS) and global gene expression analyses that are widely used in molecular medicine are increasingly applied to the field of fish biology. This has facilitated new directions to address research areas that could not be previously considered due to the lack of molecular information for ecologically relevant species. Over the past decade, the cost of NGS has decreased significantly, making it possible to use non-model fish species to investigate emerging environmental issues. NGS technologies have permitted researchers to obtain large amounts of raw data in short periods of time. There have also been significant improvements in bioinformatics to assemble the sequences and annotate the genes, thus facilitating the management of these large datasets.The combination of DNA sequencing and bioinformatics has improved our abilities to design custom microarrays and study the genome and transcriptome of a wide variety of organisms. Despite the promising results obtained using these techniques in fish studies, NGS technologies are currently underused in ecotoxicogenomics and few studies have employed these methods. These issues should be addressed in order to exploit the full potential of NGS in ecotoxicological studies and expand our understanding of the biology of non-model organisms

Kyste De L’ouraque Infecte : A Propos De Trois Cas Cliniques Au Chu De Parakou Au Benin

Bacground: The urachus is a fibrous remnant of the allantois. That connects the bladder dome to the anterior abdominal wall. After birth it is obliterated in general. In some cases it may persist as cyst , fistula , sinus or diverticulum. This persistence of the urachus often is signaled by a complication. Infection is the most common complication and the most dangerous is his degeneration. The infection can be misunderstood and confused with other pathologies of the navel. Methods: Our study reports three clinical cases of infected urachal cyst. Result: these report cases illustrate the need to think of this diagnosis before the painful of the navel and abdomen. The ultrasound examination can help for the diagnosis

Scenarii Bioclimatiques A L’horizon 2050 Dans Le Departement De L’oueme Au Benin (Afrique De L’ouest)

This study aims to determine the future bioclimatic atmospheres by 2050 according to the scenarios A1B and B1, in order to determine if the human health of the populations in the department Ouémé in Benin would be subjected to more or less harsh environments. To do this, this study was conducted using descriptive statistics methods, and bioclimatic index calculation (UTCI). The data used are the meteorological data (temperature, relative humidity, insolation and wind) on a monthly scale over the period 1971-2014 and the data from 2020 to 2025 from the ReMO database. The results of this study make it possible to remember that the A1B scenario presents a distinct singularity, because it describes more bioclimatic atmospheres than the B1 scenario. Whatever the variations, the December- March period will be dominated by a hot atmosphere, while April-October by a more comfortable atmosphere. Differences between bioclimatic atmospheres by 2050 and the current one will reach +9.2 in February for the A1B scenario and +8.4 for the B1 scenario. This variation of future bioclimatic atmospheres simulated by means of the REMO data and UTCI will not be without effects on the health of children from 0 to 5 years old in the Department of Ouémé and therefore in the face of this future configuration. adaptations are proposed to the different actors in the study area

Les Perforations D’ulcere Gastro-Duodenal Au Centre National Hospitalier Et Universitaire (Cnhu) De Cotonou (Benin)

Introduction: Duodenal or gastric ulcer perforations constitute a medical and surgical emergency. Purpose: Expose its epidemiological, diagnostic and therapeutic aspects. Method: About a retrospective study from January 1st 2006 to March 31st, 2012 in visceral surgery at CNHU of Cotonou, records of 86 cases of duodenal or gastric ulcer perforation were analyzed. Results: On average 13.8 cases / year, perforation of peptic ulcer was the most etiological cause of acute generalized peritonitis (25.7%). The average age was 34.2 years, the sex ratio of 16.2. Average admission stay was 6 days and it was observed: undocumented epigastralgia (42cas; 48.8%), no pathological medical history (32 cases; 37.2%). Peritoneal syndrome was in 81 cases (94.2%). Abdomen radiography without preparation (71 patients / 86; 82.6%) showed a pneumoperitoneum (52 cases / 71; 77.6%). The treatment was nonoperative (2 cases / 86; 2.3%) with a favorable outcome. Surgical patients had duodenal perforation (49 cases / 84; 57%), gastric perforation (35 cases / 84; 43%). Suturing with or without other technical devices was achieved in all cases. Postoperative complications were present in 28.5% of cases and parietal abscesses were documented as a priority (14 cases / 84; 16.7%). Mortality rate was 4.7% (4/86) and the average hospital stay was 11.6 days. Conclusion: The perforations of peptic ulcer were common in young men. Clinical examination was especially helpful in the diagnosis by abdomen radiography without preparation. The treatment was medical and surgical and the outcome was mostly favorable