A fast and accurate method for specific detection and quantification of the bloom-forming microalgae Karlodinium veneficum in the marine environment.

Karlodinium veneficum is a toxic benthic globally distributed dinoflagellate which has direct impacts on human health and the environment. Early and accurate detection of this harmful algal bloom-forming species could be useful for potential risks monitoring and management. In the present work, a real-time PCR targeting the internal transcribed spacer ribosomal DNA region for the specific detection and absolute quantification of K. veneficum was designed. Then, the assay conditions were adjusted and validated. The developed qPCR was highly specific for the target species and displayed no cross-reactivity with closely related dinoflagellates and/or other microalgal species commonly distributed along the Tunisian coast. Its lowest detection limit was 5 rDNA copies per reaction, which is often considered satisfying. qPCR assay enumeration accuracy was evaluated using artificially inoculated environmental samples. The comparison of the cell abundance estimates obtained by qPCR assay with the theoretical estimates showed no statistically significant difference across a range of concentrations. We suggest that the qPCR approach developed in the present study may be a valuable tool to investigate the distribution and seasonal dynamics of K. veneficum in marine environments.

Development of a novel TaqMan qPCR assay for rapid detection and quantification of Gymnodinium catenatum for application to harmful algal bloom monitoring in coastal areas of Tunisia.

Gymnodinium catenatum is a dinoflagellate known to cause paralytic shellfish poisoning (PSP), commonly associated with human muscular paralysis, neurological symptoms, and, in extreme cases, death. In the present work, we developed a real-time PCR-based assay for the rapid detection of the toxic microalgal species, G. catenatum, in environmental bivalve mollusc samples as well as seawater samples. G. catenatum-specific primers and probe were designed on the ITS1-5.8S-ITS2 rDNA region. Hydrolysis probe qPCR assay was optimized. ITS1-5.8S-ITS2 rDNA region copy numbers per G. catenatum cell genome were estimated to be 122.73 ± 5.54 copies/cell, allowing cell quantification. The application of the optimized qPCR assay for G. catenatum detection and quantification in field samples has been conducted, revealing high sensitivity (detection of around 1.3105 cells/L of seawater samples. Thus, the designed hydrolysis probe qPCR assay could be considered an efficient tool for phytoplankton monitoring whilst ensuring accuracy and sensitivity and providing cost and time savings.

Implant placement in a focal cemento-osseous dysplasia: A modified protocol with a successful outcome.

Cemento-osseous dysplasia (COD) is defined as a condition in which normal bone is replaced by fibrous connective tissue and cementum-like deposits. It is generally asymptomatic and occurs mostly in female patients. Radiologically, it appears as an opaque, lobulated mass. When facing such lesions, it is advised to avoid performing any surgical procedures due to the decreased vascularization and healing potential. The main reported complications being poor healing, sequestrum formation, risk of infection, and fracture of the jaw. Treating posterior mandibular edentulism in presence of a COD can be a challenging situation especially when the patient requires an implant-supported fixed rehabilitation. The aim of this paper is to describe a three-stage modified protocol for implant placement in a compromised site presenting a focal cemento-osseous dysplasia.

q-PCR-based assay for the toxic dinoflagellate Karenia selliformis monitoring along the Tunisian coasts.

Karenia selliformis is a marine dinoflagellate responsible for fish-kill events. Its presence has been reported along the Tunisian coasts (south-eastern Mediterranean Sea) since the 1990s. In the present study, a quantitative-PCR assay, based on the internal transcribed spacer (ITS) molecular marker, was developed to detect and quantify K. selliformis in environmental bivalve mollusk samples and in seawater samples. The assay was optimized, and its specificity was confirmed using cross-reactivity experiments against microalgal species commonly found on the Tunisian coasts and/or closely related to K. selliformis. Calibration curves were performed by tenfold dilutions of plasmid DNA harboring target sequence and genomic DNA, attaining a limit of detection of around 5 copies of target DNA per reaction, far below one K. selliformis cell per reaction. The field application of the developed assay showed a powerful detection capability. Thus, the designed assay could contribute to the deployment of in-field diagnostic tools for K. selliformis blooms monitoring.

Lateral maxillary sinus floor elevation in the presence of a sinus osteoma: A case report.

Eventhough the lateral sinus floor elevation is a well-documented procedure, many factors can increase its difficulty. The presence an osteoma can be very challenging and must be managed with caution taking in consideration the lesions's size and extension.

Glycated hemoglobin influence on periodontal status, pathogens and salivary interleukins in type II diabetic Tunisian subjects with chronic periodontitis.

BACKGROUND/PURPOSE: Studies have shown that there is a possible correlation between the amount of glycated hemoglobin and the periodontal status. The goal of this study was to investigate the relationship between glycated hemoglobin (HbA1c) and the prevalence of gingival pathogens and circulating interleukin levels in type II diabetic Tunisian subjects. MATERIAL AND METHODS: The research included four groups; 30 healthy subjects (H group), 30 non-diabetic subjects suffering from chronic periodontitis (CP group). Type-II diabetic patients were divided according to HbA1c level into 30 adequately-controlled type-II diabetes subjects (HbA1c ≤ 7 percent (ATIID&CP group)) and 30 inadequately-controlled type-II diabetes subjects and HbA1c > 7 percent (ITIID&CP group). Clinical periodontal condition parameters and assessment of salivary interleukin IL-1beta, IL-6 and IL-10 were assessed. Quantitative Polymerase Chain Reaction used for detection of Subgingival biofilm of periodontal pathogens. RESULTS: Clinical parameters analyzed were positively associated with HbA1c levels (p < 0.05). A. Actinomycetemcomitans were found in 80 percent of ITIID&CP, 65 percent of CP and almost absent in H group. Porphyromonas gingivalis was present in 100 percent of CP, 85 percent of ITIID&CP, 50 percent of ATIID&CP and 3 percent of H group. T. Denticola had an equivalent occurrence. While Tannerella forsythia was scarce in ITIID&CP groups, but abundant in the H group. ITIID&CP had the highest IL-6 and IL-1beta/IL-10 ratios. CONCLUSION: HBA1c levels affect periodontal status, pathogens and salivary interleukins in Type-II diabetic Tunisians with chronic periodontitis, compared with stable and chronic periodontitis groups and can interact with periodontal infections and increase the inflammatory state.

Zinc biosorption by Dunaliella sp. AL-1: Mechanism and effects on cell metabolism.

Phycoremediation is being considered as an eco-friendly and safe technology for toxics eradication from contaminated aquatic systems. The zinc biosorption capacity of Dunaliella sp. AL-1 was demonstrated. Zinc impacted cell growth and photosynthetic pigments accumulation showing exposure time and concentration-dependent effects. The investigation of the antioxidant protective response to zinc exposition proved a stimulation of guaiacol peroxidase (GPX) activity and an increased rate of total phenolics, flavonoids, condensed tannins and glutathione (GSH). The Box-Behnken design was used to optimize zinc removal conditions by Dunaliella sp. AL-1 strain. The maximum experimental zinc uptake was obtained when zinc concentration, algae dose, initial pH, and contact time were set at 25 mg/L, 0.5 g/L, 7.59 and 13 h 43 min, respectively. Under completely optimized conditions, the fraction of zinc removed intracellularly was much lower than the adsorbed on the cell surface. FTIR analysis Dunaliella sp. AL-1 biomass demonstrated that several functional groups as OH, CH2, CO, PO, COO and CO may participate in the biosorption process. A comparative proteomic analysis through nano-HPLC coupled to LC-MS/MS, was performed from pre- and post-zinc treatments cells. Among 199 identified proteins, 60 were differentially expressed of which 41 proteins were down-regulated against 19 up-regulated ones. Target proteins have been demonstrated to be implicated in different metabolic processes mainly photosynthesis and antioxidant defenses.

Extracellular neutral protease from Arthrospira platensis: Production, optimization and partial characterization.

Proteases are industrially important catalysts. They belong to a complex family of enzymes that perform highly focused proteolysis functions. Given their potential use, there has been renewed interest in the discovery of proteases with novel properties and a constant thrust to optimize the enzyme production. In the present study, a novel extracellular neutral protease produced from Arthrospira platensis was detected and characterized. Its proteolytic activity was strongly activated by ß-mercaptoethanol, 5,5-dithio-bis-(2-nitrobenzoic acid) and highly inhibited by Hg2+ and Zn2+ metal ions which support the fact that the studied protease belongs to the cysteine protease family. Using statistical modelling methodology, the logistic model has been selected to predict A. platensis growth-kinetic values. The optimal culture conditions for neutral protease production were found using Box-Behnken Design. The maximum experimental protease activities (159.79 U/mL) was achieved after 13 days of culture in an optimized Zarrouk medium containing 0.625 g/L NaCl, 0.625 g/L K2HPO4 and set on 9.5 initial pH. The extracellular protease of A. platensis can easily be used in the food industry for its important activity at neutral pH and its low production cost since it is a valuation of the residual culture medium after biomass recovery.

Saliva diagnostic utility in patients with type 2 diabetes: Future standard method.

BACKGROUND: The purpose of the present study was to assess saliva reliability in diagnosis and monitoring type 2 diabetes instead of blood. METHODS: Blood and unstimulated whole saliva were collected from 300 type 2 diabetic subjects and 300 healthy controls in fasting. Then, the salivary flow rate was calculated. All parameters including glucose, urea, amylase, total protein, albumin, C-reactive protein (CRP), immunoglobulin A (IgA), potassium, calcium and chloride were assessed in the supernatant, using an autoanalyzer. Oral exam was conducted by a single examiner on full mouth excluding third molars. Statistical analysis was performed by the SPSS 20.0 version. RESULTS: Saliva screening showed that glucose, urea, amylase, total protein, potassium, calcium and chloride were significantly higher in patients compared to controls (p < 0.05). Whereas, the IgA level and salivary flow rate were significantly reduced in patients (p < 0.05). No significant difference was found in albumin and CRP levels (p > 0.05). There was a significant positive correlation between salivary and plasma glucose levels (r = 0.887, and r = 0.900, p < 0.001), as well as, salivary and blood urea (r = 0.586, and r = 0.688, p < 0.001) in patients and controls, respectively. CONCLUSIONS: From this study, saliva could be suggested as a useful diagnostic tool for type 2 diabetes.

Peri-implant Soft Tissue Management: Cairo Opinion Consensus Conference.

Peri-implant soft tissues play a role of paramount importance, not only on the esthetic appearance, but also on the maintenance and long-term stability of implants. The present report presents the conclusions from the Consensus Conference of the South European North African Middle Eastern Implantology & Modern Dentistry Association (SENAME) (4-6 November 2016, Cairo, Egypt). The conference focused on the topic of the soft tissue around dental implants, and in particular, on the influence of implant configurations on the marginal soft tissues, soft tissue alterations after immediate, early or delayed implant placement and immediate loading, the long-term outcomes of soft tissue stability around dental implants, and soft tissue augmentation around dental implants. Thirty world experts in this field were invited to take part in this two-day event; however, only 29 experts were in the final consensus voting process.

Investigation of carbonic anhydrase 6 gene polymorphism rs2274327 in relation to the oral health status and salivary composition in type 2 diabetic patients.

OBJECTIVE: The aim of the present study was to investigate the oral manifestations and salivary composition in type 2 diabetics with periodontitis and to evaluate their association with CA6 gene polymorphism rs2274327. METHODS: Oral examination was performed by a single periodontist for 300 type 2 diabetics. Whole unstimulated saliva and blood were collected. The salivary pH, buffer capacity and flow rate were later measured. Immunoglobulin A and electrolytes were assessed using an autoanalyzer. CA6 gene polymorphism rs2274327 was screened by PCR-RFLP assay. The statistical analysis was performed using the SPSS 20.0 version. RESULTS: The salivary pH, buffer capacity and flow rate were significantly lower in the patients carrying TT genotype compared to CC and CT genotype carriers (p < .05). Furthermore, the DMFT index, OHI-s, PI, PPD and CAL were significantly higher in the subjects with TT genotype (p < .05). Carrying at least one T allele seemed to increase the risk of dental caries (OR = 2.59, p < .001), xerostomia (OR = 2.11, p=.003) and taste impairment (OR = 1.97, p < .05). CONCLUSION: CA6 gene polymorphism rs2274327 seemed to increase the risk of developing, dental caries, periodontitis, xerostomia and taste impairment in type 2 diabetics.

The Incidence of Complications of Dental Trauma and Associated Factors: A Retrospective Study.

OBJECTIVE: The aim of this work was to study the incidence of complications of dental traumatisms and look for associations between factors related to trauma and the occurrence of complications. Materials and Methods. It is a longitudinal retrospective study on a sample of 125 traumatized teeth. The sample is taken from patients consulting the dentistry service at the hospital Sahloul Sousse between 2014 and 2017. Criteria for including a patient were presence of a permanent incisor affected by a subluxation, intrusion, lateral luxation, extrusion, or avulsion injuries associated or not with concomitant dentoalveolar injuries. Data were collected using a questionnaire. The information about etiology of trauma, delay of consultation, orientation of the patient, kind of injury, and emergency treatment and complications were obtained from the patients' records. RESULTS: The incidence of complications was 8%: external root resorption was present in 70% of cases, surface resorption was observed in 10% of cases, and replacement resorption in 10%, ankylosis in 10%. About pulpal complications, pulp necrosis was found after 4 weeks of follow-up, as well as the internal root resorption after one year. The most common cause of the trauma was the fall (40%). The majority of patients came for emergency consultation within "1 to 3 days," and the coronary fracture without pulp exposure was the first diagnosis (20.60%). Statistical analysis showed no significant relationship between the cause of the trauma and the complication (P=0.577) and between the delay of consultation and complication (P=0.577) and between the delay of consultation and complication (P=0.577) and between the delay of consultation and complication (. CONCLUSION: Treatment of dental injuries is usually delayed and not given as much attention as general medical treatment that can explain the occurrence of pulpal and periodontal complications. Immediate consultation and treatment could improve long-term prognosis of the injured tooth.

Ridge preservation techniques to avoid invasive bone reconstruction: A systematic review and meta-analysis: Naples Consensus Report Working Group C.

PURPOSE: To analyse and compare the dimensional changes of unassisted extraction sockets with alveolar ridge preservation (ARP) techniques and investigate any factors that impact the resorption of the alveolar bone. MATERIALS AND METHODS: A systematic search was conducted to identify randomised clinical trials (RCTs). All data were extracted, and a meta-analysis was performed for the changes in all buccolingual ridge width, midbuccal and midlingual ridge height, and mesial and distal ridge height, and horizontal width at reference points apical to the crestal area. RESULTS: Based on 14 RCTs, the effectiveness of ARP in reducing the dimensions of the postextraction alveolar socket was confirmed. The clinical magnitude of this effect was 1.95 mm in the buccolingual ridge width, 1.62 mm in the midbuccal ridge height, and 1.26 mm on the midlingual ridge height. Additionally, 0.45 mm and 0.34 mm for mesial and distal ridge height, and 1.21 mm, and 0.76 mm for ridge width changes at points 3 and 5 mm apical to the crest were noted. Meta-regression analyses revealed that the reflection of flaps and primary wound coverage during ARP may have detrimental effects on bone remodelling, while no statistical significance was observed for any of the bone graft substitutes or the percentage of molar sockets. CONCLUSIONS: Regardless of the protocol, ARP can only minimise ridge resorption. ARP is most effective on horizontal ridge width, providing the most benefit coronally (approximating the crest), followed by the midbuccal ridge height.

[Orthodontic intrusion using mini-screws]. / Ingression orthodontique sur mini-vis d'ancrage.

INTRODUCTION: Dental intrusion has long been considered one of the most difficult movements to induce in orthodontics. Using conventional mechanics, the main difficulty lies in the need to ensure anchorage control, which is highly complicated to achieve, so as to avoid parasitic movements. In this framework, mini-screws have proven to offer a very effective means of anchorage, allowing greater control over intrusion of the anterior and posterior teeth and a simpler biomechanical movement opening up new therapeutic perspectives for the orthodontist. OBJECTIVE: The aim of this study was to describe the clinical and biomechanical application of mini-screws for dental intrusion.

Insight into Arthrospira platensis Δ9 desaturase: a key enzyme in poly-unsaturated fatty acid synthesis.

Membrane-bound Δ9 desaturase perform oxygenated desaturation reactions to insert the first double bonds within fatty acyl chains between C9 and C10 positions of most saturated substrates. Arthrospira platensis, a blue green microalga, is an important source of polyunsaturated fatty acids (PUFA) such as oleic, linoleic and linolenic acids lending benefits and functions in dietetics and therapeutic uses. In this paper, we report homology modeling and docking studies of a Δ9 desaturase from Arthrospira platensis strain. The protein model showed high topology resemblance compared to membrane-bound desaturases with a cytoplasmic core displaying the catalytic site and a transmembrane domain created by four α-helices. The cytoplasmic cap contained the three conserved-histidine boxes typical for all membrane bound desaturases. The protein model was used to perform protein-protein docking and the dimer structure was generated. The two monomers are tightly related with hydrophobic interactions between the transmembrane domain helices. The study highlighted also the potent role of a particular 53 residues sequence located at the N terminal end of the enzyme.

Proteomic Analysis of the Chlorophyta Dunaliella New Strain AL-1 Revealed Global Changes of Metabolism during High Carotenoid Production.

The green microalgae Dunaliella genus is known for the production of high added value molecules. In this study, strain AL-1 was isolated from the Sebkha of Sidi El Hani (Sousse, Tunisia). This isolate was identified both morphologically and genetically via 18S rRNA gene sequence as a member of the genus Dunaliella. Strain AL-1 was found to be closely related to Dunaliella salina, Dunaliella quartolecta and Dunaliella polymorpha with more than 97% similarity. Response surface methodology was used to maximize carotenoid production by strain AL-1 by optimizing its growth conditions. The highest carotenoid content was obtained at salinity: 51, light intensity: 189.89 µmol photons·m-2·s-1, and nitrogen: 60 mg·L-1. Proteomic profiling, using two-dimensional gel electrophoresis, was performed from standard and optimized cultures. We detected 127 protein spots which were significantly differentially expressed between standard and optimized cultures. Among them 16 protein spots were identified with mass spectrometry and grouped into different functional categories using KEGG (Kyoto Encyclopedia of Genes and Genomes) such as photosynthetic Calvin cycle, regulation/defense, energy metabolism, glycolysis, and cellular processes. The current study could be of great interest in providing information on the effect of stressful conditions in microalgae carotenoid production.

Cyanobacteria as source of marine bioactive compounds: Molecular specific detection based on Δ9 desaturase gene.

The blue-green microalga, Arthrospira sp., isolated from the sea of Kssour Essef in Mahdia (Tunisia), was purified and then identified both morphologically and genetically based on 16S rRNA gene sequence. Following physicochemical analysis, the prokaryotic microalga tested represented a competitive source of pigments and showed a considerable rate in protein (64%) which was confirmed by FTIR measurement. The lipid content (4%) was quantified by the gravimetric method and the intracellular lipid bodies were detected with the Nile red staining. Using gas chromatography coupled with flame ionization detector, the fatty acid profile revealed the presence of 27.4% and 32.88% of monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs), respectively. Given the richness of the isolated microalga in unsaturated fatty acids, we have developed a SYBR Green real time PCR method for the specific identification of Arthrospira sp. Δ9 desaturase gene. This current method will be of great value for carrying out high-throughput studies like cloning, heterologous expression and structure-function relationship analysis.

Age assessment in canine and premolar by cervical axial sections of cone-beam computed tomography.

Age estimation in adults is a challenge in both anthropological and forensic situations compared to sub-adults age estimation. The present study explored, for the first time, the cervical zone of single rooted teeth. The deposition of secondary dentin is responsible for a decrease in the volume of the dental pulp cavity with aging, and therefore is often used as an age indicator. The current study aimed at estimating the chronological age among adults by measuring the pulp/dentin area ratio (physiological ratio) by axial sections at cervical region of maxillary canine and mandibular second premolar. The sample consisted of 120 cone beam CT images of 120 Tunisians whose age ranged from 22 to 67, from the database of a private clinic of radiology. The first axial section of chosen teeth without enamel was selected. Linear regression models were derived for canine, premolar and for all variables to predict the age. They indicated that dentin deposition on canine and premolar have almost the same correlation with age (r=-0.838 and -0.837 respectively). The residual standard errors (RSE), when these regression equations applied for the entire sample, were ranged between 8.27, 8.29 and 7.06 for canine, premolar and for all variables respectively. Tested for younger ages (from 22 to 44years) the RSE decreased considerably and thus ranged between 4.32, 4.72 and 4.05. The outcomes of this study show that the physiological ratio is a useful variable for assessing age with a satisfying accuracy.

Enhanced lipid and biomass production by a newly isolated and identified marine microalga.

BACKGROUND: The increasing demand for microalgae lipids as an alternative to fish has encouraged researchers to explore oleaginous microalgae for food uses. In this context, optimization of growth and lipid production by the marine oleaginous V2-strain-microalgae is of great interest as it contains large amounts of mono-unsaturated (MUFAs) and poly-unsaturated fatty acids (PUFAs). METHODS: In this study, the isolated V2 strain was identified based on 23S rRNA gene. Growth and lipid production conditions were optimized by using the response surface methodology in order to maximize its cell growth and lipid content that was quantified by both flow cytometry and the gravimetric method. The intracellular lipid bodies were detected after staining with Nile red by epifluorescence microscopy. The fatty acid profile of optimal culture conditions was determined by gas chromatography coupled to a flame ionization detector. RESULTS: The phenotypic and phylogenetic analyses showed that the strain V2 was affiliated to Tetraselmis genus. The marine microalga is known as an interesting oleaginous species according to its high lipid production and its fatty acid composition. The optimization process showed that maximum cell abundance was achieved under the following conditions: pH: 7, salinity: 30 and photosynthetic light intensity (PAR): 133 µmol photons.m-2.s-1. In addition, the highest lipid content (49 ± 2.1% dry weight) was obtained at pH: 7, salinity: 37.23 and photosynthetic light intensity (PAR): 188 µmol photons.m-2.s-1. The fatty acid profile revealed the presence of 39.2% and 16.1% of total fatty acids of mono-unsaturated fatty acids (MUFAs) and poly-unsaturated fatty acids (PUFAs), respectively. Omega 3 (ω3), omega 6 (ω6) and omega 9 (ω9) represented 5.28%, 8.12% and 32.8% of total fatty acids, respectively. CONCLUSIONS: This study showed the successful optimization of salinity, light intensity and pH for highest growth, lipid production and a good fatty acid composition, making strain V2 highly suitable for food and nutraceutical applications.