Mobility of trace metals and microbiological pollution from dredged sediments to the Gulf of Gabes, Tunisia.

Sediments are periodically dredged from the major ports in the Gulf of Gabes (GG) during maintenance operations. These sediments are dumped near the coats of Sfax and Gabes cities. In the present study, 6 trace metals (Cd, Cu, Cr, Ni, Pb, and Zn) concentrations were assessed in both sediments and column water taken from the different basins of Sfax port. This study is the first to focus on the microbial and microalgae contaminations of Sfax port sediments. The spatial distributions of trace metals in dredged sediments from the different basins of Sfax port show that the maximum concentrations of Cd (13.75 µg/g), Cu (892.5 µg/g), and Zn (1447 µg/g) exceeded the Geode standard thresholds. The same elements, also detected in the water column, exceeded the toxicity thresholds for phytoplankton, shellfish, and algae (Cd 0.095 µg/l, Cu 4.52 µg/l, and Zn 37 µg/l). The presence of coliforms, indicators of sewage pollution, as pathogenic germs (Pseudomonas), was shown through microbiological examinations of the sediments and water column. The microalgae enumeration showed the abundance of dinoflagellate cysts (stressed form) which attests to the presence of severe conditions in Sfax port. Different toxic species were identified as Prorocentrum lima and Alexandrium minutum. The same trace metal sources and abundance in Sfax port and GG sediments suggest the hypothesis of their transfer from their disposal sites in coastal areas to deeper depths in the GG and until Boughrara lagoon (BL). As trace metals, microalgae species were also transferred by hydrodynamic currents inside the GG where they found suitable conditions to their proliferation causing the seawater coloration phenomenon, eutrophication, and degradation of the aquatic system.

Rapid Detection of Beta-Lactamases Genes among Enterobacterales in Urine Samples by Using Real-Time PCR.

The objective of this study was to develop and evaluate newly improved, rapid, and reliable strategies based on real-time PCR to detect the most frequent beta-lactamase genes recorded in clinical Enterobacterales strains, particularly in Tunisia (blaSHV12 , blaTEM , blaCTX-M-15 , blaCTX-M-9 , blaCMY-2 , blaOXA-48 , blaNDM-1 , and blaIMP ) directly from the urine. Following the design of primers for a specific gene pool and their validation, a series of real-time PCR reactions were performed to detect these genes in 78 urine samples showing high antibiotic resistance after culture and susceptibility testing. Assays were applied to DNA extracted from cultured bacteria and collected urine. qPCR results were compared for phenotypic sensitivity. qPCR results were similar regardless of whether cultures or urine were collected, with 100% sensitivity and specificity. Out of 78 multiresistant uropathogenic, strains of Enterobacterales (44 E. coli and 34 K. pneumoniae strains) show the presence of the genes of the bla group. In all, 44% E. coli and 36 of K. pneumoniae clinical strains harbored the bla group genes with 36.4%, 52.3%, 70.5%, 68.2%, 18.2%, and 4.5% of E. coli having blaSHV-12 , blaTEM , blaCTX-M 15 , blaCTX-M-9 , blaCMY-2 , and blaOXA-48 group genes, respectively, whereas 52.9%, 67.6%, 76.5%, 35.5%, 61.8, 14.7, and 1.28% of K. pneumoniae had blaSHV-12 , blaTEM , blaCTX-M 15 , blaCTX-M-9 , blaCMY-2 , blaOXA-48 , and blaNDM-1 group genes, respectively. The time required to have a result was 3 hours by real-time PCR and 2 to 3 days by the conventional method. Resistance genes of Gram-negative bacteria in urine, as well as cultured bacteria, were rapidly detected using qPCR techniques. These techniques will be used as rapid and cost-effective methods in the laboratory. Therefore, this test could be a good candidate to create real-time PCR kits for the detection of resistance genes directly from urine in clinical or epidemiological settings.

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 ß-lactamase (ESBL) production was tested by double-disc synergy test. Strains were further subjected to a molecular assessment of ESBL and AmpC ß-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 penicillins, 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: blaCTX-M group 1 and group 9, and ACC and FOX AmpC ß-lactamase genes. Overall, ESBLs and AmpC ß-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.

A New Lavender (Lavandula multifida L.) Ecotype from Arid Tunisia, with Differential Essential Oil Composition and Higher Antimicrobial Potential.

The lavender Lavandula multifida L., a medicinal plant grown in arid regions of Tunisia, was recently considered an endangered species; thus, its habitats regressed to some difficult zones in terms of access, such as the watershed of Oued Agareb in central-eastern Tunisia. This species was recorded only in deep and narrow shady Wadi of the watershed and benefited from protection against overgrazing, erosion and sunlight. L. multifida was rarely observed in an open area, such as a plateau or large-bed valley. The plant's metabolism is linked to its response to environmental conditions, which is of particular interest to understanding the components of the considered population of L. multifida. Consequently, biochemical and antimicrobial analyses have been evaluated. Using gas chromatography-mass spectrometry (GC-MS) analysis reveals that among the 58 compounds identified in L. multifida essential oil extracted from aboveground plant tissues, camphor was the major component (15.68%), followed by 1,8-cineole (14.14%) and alpha-pinene (13.82%). Moreover, it has been observed that Escherichia coli was more susceptible than Staphylococcus aureus to the antimicrobial properties of L. multifida essential oil, while in the case of camphor, S. aureus was more susceptible than E. coli. The protected population of L. multifida exhibits a distinctive vegetative development and growth cycle, resulting in specific secondary metabolites and distinguished antimicrobial activity.

Elimination of Escherichia coli and Salmonella in Clam by Using Zeolite in a Station of Depuration.

The application of natural zeolite for water and wastewater treatment has been carried out and is still a promising technique in environmental cleaning processes. Natural zeolite can be used to improve the purification process of clams (Ruditapes decussatus). Thus, our study aimed at improving the clam purification system in order to reduce Escherichia coli and eliminate Salmonella in samples artificially contaminated with this bacterium using a natural zeolite to replace the biological filter. The results showed that zeolite used in a depuration system improved the clam purification process. Moreover, natural zeolite exhibited high performance in the adsorption of bacteria and allowed to reduce the Escherichia coli abundance in 24 h, thus ensuring purified clams conformity with the ISO 16649-3 standard. These results indicate the beneficial effects of using zeolite in the adsorption of bacteria and the reduction in the abundance of Escherichia coli and set the Salmonella from marine organisms.

Antimicrobial effect of the Tunisian Nana variety Punica granatum L. extracts against Salmonella enterica (serovars Kentucky and Enteritidis) isolated from chicken meat and phenolic composition of its peel extract.

Punica granatum L. is widely recognized for its potency against a broad spectrum of bacterial pathogens. The purpose of this study was to explore the inhibitory and the bactericidal activities of Punica granatum against Salmonella strains. The effect of extracts obtained from different parts (peels, seeds, juice and flowers) of pomegranate and using different solvents against Salmonella enterica serovars Kentucky and Enteritidis isolated from chicken meat was thus investigated. Salmonella strains were identified with the standard API-20E system and confirmed by real time PCR. The obtained results showed that the highest antibacterial activity against Salmonella strains was observed with the peels ethanolic extract giving MIC values ranging from 10.75 to 12.5mg/mL. The ethanolic extract of P. granatum Nana peels at 0.8 and 1.6mg/g significantly inhibited the growth of Salmonella Kentucky in chicken meat stored at 4°C. The phenolic composition of the ethanolic peel extract was explored by HPLC coupled to both DAD and ESI/TOF-MS detections. The obtained results allowed the detection of 21 phytochemical compounds among which various phenolic compounds have been identified on the basis of their UV and MS spectra as well as with literature data. Among the detected compounds, anthocyanins, ellagitannins, ellagic acid derivatives and flavanols were further characterized through MS-MS analysis. Our results showed thus that the Tunisian variety Nana pomegranate constitutes a good source of bioactive compounds with potent antimicrobial activity on the growth of Salmonella strains suggesting that the studied pomegranate cultivar could be a natural remedy to minimize the emergence of Salmonella enterica strains which is often involved in food borne illness.

Isolation of a novel amylase and lipase-producing Pseudomonas luteola strain: study of amylase production conditions.

An amylase and lipase producing bacterium (strain C2) was enriched and isolated from soil regularly contaminated with olive washing wastewater in Sfax, Tunisia. Cell was aerobic, mesophilic, Gram-negative, motile, non-sporulating bacterium, capable of growing optimally at pH 7 and 30°C and tolerated maximally 10% (W/V) NaCl. The predominant fatty acids were found to be C(18:1)ω7c (32.8%), C(16:1)ω7c (27.3%) and C16:0 (23.1%). Phylogenetic analysis of the 16S rRNA gene revealed that this strain belonging to the genus Pseudomonas. Strain C2 was found to be closely related to Pseudomonas luteola with more than 99% of similarity. Amylase optimization extraction was carried out using Box Behnken Design (BBD). Its maximal activity was found when the pH and temperature ranged from 5.5 to 6.5 and from 33 to 37°C, respectively. Under these conditions, amylase activity was found to be about 9.48 U/ml.

Fungal diversity in adult date palm (Phoenix dactylifera L.) revealed by culture-dependent and culture-independent approaches.

Endophytic flora plays a vital role in the colonization and survival of host plants, especially in harsh environments, such as arid regions. This flora may, however, contain pathogenic species responsible for various troublesome host diseases. The present study is aimed at investigating the diversity of both cultivable and non-cultivable endophytic fungal floras in the internal tissues (roots and leaves) of Tunisian date palm trees (Phoenix dactylifera). Accordingly, 13 isolates from both root and leaf samples, exhibiting distinct colony morphology, were selected from potato dextrose agar (PDA) medium and identified by a sequence match search wherein their 18S-28S internal transcribed spacer (ITS) sequences were compared to those available in public databases. These findings revealed that the cultivable root and leaf isolates fell into two groups, namely Nectriaceae and Pleosporaceae. Additionally, total DNA from palm roots and leaves was further extracted and ITS fragments were amplified. Restriction fragment length polymorphism (RFLP) analysis of the ITS from 200 fungal clones (leaves: 100; roots: 100) using HaeIII restriction enzyme revealed 13 distinct patterns that were further sequenced and led to the identification of Alternaria, Cladosporium, Davidiella (Cladosporium teleomorph), Pythium, Curvularia, and uncharacterized fungal endophytes. Both approaches confirmed that while the roots were predominantly colonized by Fusaria (members of the Nectriaceae family), the leaves were essentially colonized by Alternaria (members of the Pleosporaceae family). Overall, the findings of the present study constitute, to the authors' knowledge, the first extensive report on the diversity of endophytic fungal flora associated with date palm trees (P. dactylifera).

Genetic diversity of food-isolated Salmonella strains through Pulsed Field Gel Electrophoresis (PFGE) and Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR).

All over the world, the incidence of Salmonella spp contamination on different food sources like broilers, clams and cow milk has increased rapidly in recent years. The multifaceted properties of Salomnella serovars allow the microorganism to grow and multiply in various food matrices, even under adverse conditions. Therefore, methods are needed to detect and trace this pathogen along the entire food supply network. In the present work, PFGE and ERIC-PCR were used to subtype 45 Salmonella isolates belonging to different serovars and derived from different food origins. Among these isolates, S. Enteritidis and S. Kentucky were found to be the most predominant serovars. The Discrimination Index obtained by ERIC-PCR (0.85) was slightly below the acceptable confidence value. The best discriminatory ability was observed when PFGE typing method was used alone (DI = 0.94) or combined with ERIC-PCR (DI = 0.93). A wide variety of profiles was observed between the different serovars using PFGE or/and ERIC-PCR. This diversity is particularly important when the sample origins are varied and even within the same sampling origin.

Influence of Zeolite on fatty acid composition and egg quality in Tunisian Laying Hens.

BACKGROUND: The health benefits of omega-3 and omega-6 polyunsaturated fatty acids (PUFA) are generally recognized. Unfortunately, in most Mediterranean countries, the recommended daily intake of these compounds is rarely met. Therefore, enrichment of commonly occurring foods can boost intake of these fatty acids. In this regard, eggs are an interesting target, as they form an integral part of the diet. RESULT: Zeolite (Clinoptilolites) was added to Laying Hens feed at concentrations 1% or 2% and was evaluated for its effects on performance of the production and on egg quality. The Laying Hens were given access to 110 g of feed mixtures daily that was either a basal diet or a 'zeolite diet' (the basal diet supplemented with clinoptilolite at a level of 1% or 2%). It was found that zeolite treatment had a positive and significatif (p<0.05) effect on some parameters that were measured like egg height and eggshell strength. While dietary zeolite supplementation tended to/or has no significant effects on total egg, eggshell, yolk and albumen weights. It was found also that zeolite mainly increases level of polyunsaturated fatty acids in egg. CONCLUSION: This study showed the significance of using zeolite, as a feed additive for Laying Hens, as part of a comprehensive program to control egg quality and to increase level of polyunsaturated fatty acids on egg.

Effect of zeolite (clinoptilolite) as feed additive in Tunisian broilers on the total flora, meat texture and the production of omega 3 polyunsaturated fatty acid.

BACKGROUND: Increasing consumer demand for healthier food products has led to the development of governmental policies regarding health claims in many developed countries. In this context, contamination of poultry by food-borne pathogens is considered one of the major problems facing the progress of the poultry industry in Tunisia. RESULT: Zeolite (Clinoptilolites) was added to chicken feed at concentrations 0,5% or 1% and was evaluated for its effectiveness to reduce total flora in chickens and its effects on performance of the production. The broilers were given free and continuous access to a nutritionally non-limiting diet (in meal form)that was either a basal diet or a' zeolite diet' (the basal diet supplemented with clinoptilolite at a level of 0,5% or 1%). It was found that adding zeolite in the broiler diet significantly (p < 0,05) reduced total flora levels, as compared to the control, on the chicken body. In addition, it was found that zeolite treatment had a positive effect on performance production and organoleptic parameters that were measured and mainly on the increase level of Omega 3 fatty acid. CONCLUSION: This study showed the significance of using zeolite, as a feed additive for broilers, as part of a comprehensive program to control total flora at the broiler farm and to increase level of Omega 3 fatty acid on the chicken body.