Evaluation of Lactobacillus plantarum and PRGF as a new bioactive multi-layered scaffold PU/PRGF/gelatin/PU for wound healing.

The effect of tissue engineering strategies in combination with Lactobacillus plantarum and platelet-rich growth factor (PRGF) with the aim of creating an appropriate wound dressing can be useful in wound healing and infection prevention in patients suffering from acute and chronic skin damages. Therefore, in this study, a new approach was employed to create a bioactive multilayer electrospun scaffold composed of polyurethane (PU), PRGF, and gelatin fibers, then human adipose-derived mesenchymal stem cells (hAMSCs), fibroblast cells (HU-02) and L. plantarum were cultured on the scaffold. The physicochemical properties, biocompatibility, and antibacterial activity of the scaffold were evaluated. In addition, the expression of the migration and proliferation genes of fibroblast cells were investigated by real-time PCR (polymerase chain reaction). Mitochondrial activity assays revealed that PRFG and L. plantarum had a significant positive effect on the viability of target co-cultured cells.Fluorescent and SEM (scanning electron microscopy) images presented the cells and bacterial proliferation and adhesion in hydrophilic scaffolds within 21 days. The sustained release of PRGF from scaffolds with a zero-order pattern was confirmed. RT-PCR analysis revealed that PRGF elevated the expression of VEGF genes up to fourfold, but L. plantarum had a better effect on DDR2 gene expression compared to the TCPS group. Antibacterial tests showed that L. plantarum has a bacterial load reduction of more than 70% in CFU/mL. The present scaffold is an appropriate model for cell attachment, migration, proliferation, and infection prevention.

Impact of Nutritional Profile on Gut Microbiota Diversity in Patients with Celiac Disease.

The pathogenesis of celiac disease (CD) is significantly influenced by gut microbiota. Daily nutritional profile influences the diversity of gut microbiota. This study was aimed to compare the abundance of gut microbiota in CD patients compared to normal control (NC), and to investigate the impact of nutritional factors on their fecal microbiota diversity. In this study, a selected panel of intestinal bacteria was assessed in 31 confirmed CD patients adhering to gluten-free diet (GFD) for more than 6 months and in 20 NC subjects. Stool samples were collected from each participant, DNA was extracted, and absolute quantitative real-time PCR (qPCR) was carried out. The gut microbiota including Bacteroidetes, Bifidobacterium, Clostridium, Staphylococcus, Enterobacteiaceae, Firmicutes, and Lactobacillus were assessed. The quantities of fruits, vegetables, meat, liquids, sugar and gluten-free candy/bread consumption were evaluated using a questionnaire. The proportion of Bifidobacterium, Firmicutes, and Lactobacillus in CD cases was significantly lower than NC (P < 0.005). Significant correlation coefficients between Bifidobacterium and Lactobacillus (P < 0.001), and also Firmicutes and Lactobacillus (P < 0.001) were recorded. Moreover, a significant association between medium amount of meat and bean consumptions and low abundance of Lactobacillus and Firmicutes (P = 0.024 and P = 0.027, respectively), and also high amount of bean consumptions and low abundance of Lactobacillus (P = 0.027) in CD were observed. The results showed that meat and bean consumptions could reduce the beneficial bacteria including Firmicutes and Lactobacillus in CD patients. Therefore, changes in the gut microbiota abundance may contribute to dietary changes and unimproved CD symptoms.

Multiple Candida strains causing oral infection in COVID-19 patients under corticosteroids and antibiotic therapy: An observational study.

Introduction: The occurrence of oral candidiasis (OC) is expected in patients with COVID-19, especially those with moderate to severe forms of infection who are hospitalized and may be on long-term use of broad-spectrum antibiotics or prolonged corticosteroid therapy. We aimed to characterize clinical conditions, the prevalence profile of Candida species, and outcomes of COVID-19 patients with OC. Methods: In this observational study, oral samples were obtained from COVID-19 patients suspected of OC admitted to Razi teaching hospital. Patients with OC were monitored daily until discharge from the hospital. Species identification was performed by a two-step multiplex assay named YEAST PLEX, which identifies 17 clinically important uncommon to common yeast strains. Results: Among the 4133 patients admitted with COVID-19, 120 (2.90%) suffered from OC. The onset of signs and symptoms of OC in patients was, on average (2.92 ± 3.596 days) with a range (of 1-29 days). The most common OC presentation was white or yellow macules on the buccal surface or the tongue. In (39.16%) of patients suffering from OC multiple Candida strains (with two or more Candida spp.) were identified. The most common Candida species were C. albicans (60.57%), followed by C. glabrata (17.14%), C. tropicalis (11.42%), C. kefyr (10.83%) and C. krusei (3.42%). Notably, OC caused by multiple Candida strains was more predominant in patients under corticosteroid therapy (P <0.0001), broad-spectrum antibiotics therapy (P = 0.028), and those who used nasal corticosteroid spray (P <0.0001). The majority of patients who recovered from OC at the time of discharge were patients with OC by single Candida species (P = 0.049). Discussion: Use of corticosteroids and antimicrobial therapy in COVID-19 patients increases risk of OC by multiple Candida strains.

Immunological Properties of Exotoxin A Toxoid - Detoxified Lipopolysaccharide - Gold Nanoparticles Conjugate Against Pseudomonas aeruginosa Infection.

BACKGROUND: Pseudomonas aeruginosa is an important opportunistic pathogen, especially in patients with compromised host defense. OBJECTIVE: To prepare the conjugate of detoxified lipopolysaccharide (D-LPS) and exotoxin A toxoid (T-ETA) from P. aeruginosa in gold nanoparticles (Au NPs) in a mice model. METHODS: LPS and ETA were purified from P. aeruginosa PAO1. D-LPS was conjugated withT-ETA via the amidation method. Au NPs were bound to D-LPS-T-ETA conjugate via electrostatic interaction. Mice were immunized with D-LPS, D-LPS-Au NPs, T-ETA, T-ETA-Au NPs, D-LPS-T-ETA, D-LPS-T-ETA-Au NPs, D-LPS-Au NPs+T-ETA-Au NPs, Au NPs, and phosphate-buffered saline (PBS), and specific IgG titers were determined by the ELISA and the whole-cell ELISA methods. Mice in the vaccinated and control groups were exposed to a 2×LD50 of P. aeruginosa and mortality rates were recorded for one week. RESULTS: The results showed that vaccination by D-LPS, D-LPS-Au NPs, T-ETA, T-ETA-Au NPs, D-LPS-T-ETA, D-LPS-T-ETA-Au NPs and D-LPS-Au NPs+T-ETA-Au NPs induced specific IgG. Mice received the D-LPS-T-ETA-Au NPs conjugate showed significant protection against bacterial challenge. CONCLUSION: These data indicate that D-LPS-T-ETA-Au NPs conjugate has a significant immunogenicity potential to be applied as a new vaccine against Pseudomonas infections.

Oral health-related quality of life (OHRQoL) in cardiovascular patients referring to Fatima Zahra Hospital in Sari, Iran.

BACKGROUND: Cardiovascular Disease (CVD) is one of the leading causes of mortality and morbidity and significantly impacts the health-related quality of life. Oral infections have been linked to cardiovascular diseases such as thrombosis, cardiac infarction, stroke, and peripheral vascular disease. This study aims to evaluate the effects of oral health on the quality of life in cardiovascular patients. METHODS: The oral health-related quality of life was measured using the OHIP-14 questionnaire. Demographic information, questions regarding smoke consumption, wearing removable prostheses, nine questions regarding xerostomia, and the existence of other systemic diseases were asked from 240 participants with cardiovascular diseases. The DMFT index was clinically examined in each patient. Also, the Plaque, Gingival, and Sulcular Bleeding Indices were measured on the Ramfjord teeth. Data analysis was conducted using SPSS version 16. The independent t test, Mann-Whitney test, the variance analysis, and the Kruskal-Wallis test were used to compare variables in the present study. Also, regression models were used to eliminate the effect of confounding variables. RESULTS: Gender variables, removable prosthesis, xerostomia, DMFT, and SBI were the main determinants of quality of life in CVD patients. The mean ADD-OHIP14 of participants in the study was calculated at 21.34 ± 17.40, and the SC-OHIP14 was 6.11 ± 5.07. The mean OHRQoL was higher in females than in males, and this difference was statistically significant. OHRQoL was significantly lower in patients wearing a removable prosthesis than in those without one. The relationship between age and xerostomia was significant in this study, and patients with xerostomia had a lower quality of life than those without xerostomia. Also, the mean DMFT index in subjects with xerostomia was 23.69 ± 7.76, which was statistically significant compared to those without xerostomia. CONCLUSION: Cardiovascular patients experienced a decreased OHRQoL. Prevention or treatment of these problems seems to justify improving the quality of life in these patients.

Characterization of Clinical Isolates of Mycobacterium simiae Using Drug Susceptibility Tests and Molecular Analyses.

Mycobacterium simiae is an emerging nontuberculous mycobacterium (NTM) and an opportunistic pathogen which is described mainly in Asia and presents in the environment that can cause pulmonary infection. The objective of this study is to characterize M. simiae clinical isolates using mycobacterial interspersed repetitive unit variable-number tandem repeats (MIRU-VNTR) typing for the differentiation of the strains. A total of 169 clinical isolates of NTM were recovered from patients suspected of having tuberculosis (TB)-like and related infections. After isolation and identification of mycobacterial strains by conventional biochemical and PCR-based tests, M. simiae strains were confirmed using restriction fragment length polymorphism (RFLP)-based identification assay. Furthermore, drug susceptibility and MIRU-VNTR typing was performed using on the clinical isolates of M. simiae. Out of 169 NTM strains, 92 (54.4%) isolates were identified as M. simiae. Antibiotic susceptibility experiments indicated that all 92 M. simiae isolates were resistant to first line antimycobacterial agents. Moreover, 8 (8.6%) M. simiae isolates were resistant to ciprofloxacin; and 6 (6.5%) were resistant to both amikacin and kanamycin, while the remaining were susceptible to second line antimycobacterial agents. MIRU-VNTR analysis showed that the M. simiae isolates were classified in four distinct M. simiae clusters and two single types. The minimum spanning analysis revealed that the isolates were grouped in three complexes. The data suggested that MIRI-VNTR typing is useful for typing of M. simiae isolates, however, MIRU-16 locus was absolutely absent in M. simiae.

Metagenomic investigation of bacteria associated with dental lesions: a cross-sectional study

BACKGROUND: Dental caries is considered as one of the most significant global health problem over the world. Dental caries initiates from bacterial shifts within the supragingival biofilm, then a polymicrobial biofilm is formed on the surface of tooth, and finally various bacterial species aggregate in a complex-organized manner. The exploiting variability in 16S rRNA gene sequence has been considered as a cost-efficient high-throughput characterization approach in human oral microbiome investigations. The aim of this study is to characterize bacterial species associated with superficial dental biofilm, underlying carious dentine and root caries lesion by16S rRNA gene-based metagenomic analysis. MATERIAL AND METHODS: Herein, the bacterial communities in carious dentin lesion, biofilm and root canal samples of 30 subjects (aged 4-76 years) admitted to a clinic in Tehran during 2017 were investigated using a culture independent approach. Total genomic DNA of each tissue was subjected to metagenomic identification of bacteria using a nested PCR assay and 16S rRNA library construction method. RESULTS: 31 samples collected from 30 consenting patients (29 samples from 29 patients ant two biofilm samples from one patient). Bioinformatics analyses of a-800bp sequences of the second step of Nested-PCR revealed presence of 156 bacterial isolates in carious (n = 45), biofilm (n = 81) and root canal (n = 30) specimens. Prevotella spp., Lactobacillus vaginalis, and streptococcus spp. showed higher prevalence in carious dentin, root and biofilm samples, respectively. CONCLUSIONS: Exploring the dental microbiota and comparing them in health or diseased conditions is critical step in the determination of human general health. The method applied in this study could identify bacteria related to the three dental lesions. However, due to lack of data for comparison in Genbank or because of the sequence similarity lower than 98% for most identified bacteria, the use of more powerful approaches like NGS platforms or typing of multiple loci (MLST) in future studies is recommended

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Implementation of a novel self-induced promoter for the expression of pharmaceutical peptides in Escherichia coli: YY(3-36) peptide.

Background Increasing the expression rate of recombinant mammalian hormones in Escherichia coli by combining efficient promoters and signal sequences is a never ending process. A self-induced promoter will have some beneficial gains compared to the classical T7 promoter or its variants with isopropyl ß-D-1-thiogalactopyranoside (IPTG) as the inducer. Obesity is the prime suspect in widespread frequency of diabetes type II and cardiovascular diseases worldwide. YY (tyrosine-tyrosine) peptide is a local acting hormone, controlling appetite. Excitingly, it was has been shown that a truncated version of the YY peptide, YY(3-36) peptide, has potential as a worthy biopharmaceutical agent in the fight against obesity. Materials and methods To develop an economical expression system for the large scale production of the peptide in Gram-negative bacteria, we introduced a promoter sequence upstream of a chimeric gene for the extracellular expression of this peptide with the assistance of a signal sequence of asparaginase II from E. coli. This system has the advantage of producing a complete sequence of a truncated YY peptide, YY(3-36), without any extra tags that would require further removal with the assistance of expensive specific proteases and reduced the downstream steps, significantly. Results Recombinant production of YY(3-36) peptide under a self-induced promoter proves the efficacy of the asparaginase II signal sequence as a communicator of foreign peptides and proteins into the extracellular space of E. coli. Conclusions The application of fusion protein expression of biopharmaceuticals, especially mammalian hormones, in prokaryotic systems with the help of native signal sequences makes some common tags with expensive proteases for the removal of the attached protein Tag redundant.

Detection of tetracycline resistance genes, aminoglycoside modifying enzymes, and coagulase gene typing of clinical isolates of Staphylococcus aureus in the Southwest of Iran.

OBJECTIVES: The aim of the present study was to determine the aminoglycoside modifying enzymes (AMEs) encoded genes, tetracycline resistance genes, and the coa based typing of Staphylococcus aureus isolates in the Southwest of Iran. MATERIALS AND METHODS: Antimicrobial susceptibility of isolates was carried out by agar disk diffusion methods. Two sets of multiplex PCR mixture were used for detection of AME genes and tet genes. All of the isolates were typed with the coagulase gene typing method. Of the 121 isolates, 29.75% and 47.93% were resistant to at least one aminoglycosides and tetracyclines, respectively. RESULTS: The aac(6')-Ie-aph(2") was the most frequent gene (97.22%), and aph (3')-IIIa and ant (4')-Ia genes were detected in 61.11% and 11.11% of aminoglycoside resistant isolates, respectively. The tetK and tetM genes were detected in 82.75% and 56.9% of tetracycline resistant isolates, respectively. Overall 31.4% of isolates were MRSA. Totally 17 distinct coa gene RFLP patterns, numbered C1 to C17, were observed. The C5 was the most frequent coa type with 31 isolates. CONCLUSION: The aac(6')-Ie-aph(2") and aph (3')-IIIa genes were the most important genes contributing to aminoglycosides resistance, while resistance to tetracyclines was mediated by tetK and tetM genes. Interestingly all S. aureus with C5 as the most prevalent coa-type were resistant to at least one of the aminoglycoside antibiotics and tetracycline simultaneously. Moreover, 30 out of 31 isolates with this coa type were MRSA, indicating the importance of the C5 coa-type in MRSA strains and also in isolates that were resistant to aminoglycosides and tetracycline.

Biofilm formation in clinical isolates of nosocomial Acinetobacter baumannii and its relationship with multidrug resistance

Objective: To check biofilm formation by Acinetobacter baumannii (A. baumannii) clinical isolates and show their susceptibility to different antibiotics and investigate a possible link between establishment of biofilm and multidrug resistance. Methods: This study was performed on clinical samples collected from patients with nosocomial infections in three hospitals of Tehran. Samples were initially screened by culture and biochemical tests for the presence of different species of Acinetobacter. Iden-tifications were further confirmed by PCR assays. Their susceptibilities to 11 antibiotics of different classes were determined by disc diffusion method according to Clinical and Laboratory Standards Institute guidelines. The ability to produce biofilm was investigated using methods:culture on Congo red agar, microtiter plate, and test tube method. Results: From the overall clinical samples, 156 specimens were confirmed to contain A. baumannii. The bacteria were highly resistant to most antibiotics except polymyxin B. Of these isolates, 10.26% were able to produce biofilms as shown on Congo red agar. However, the percentage of bacteria with positive biofilm in test tube, standard microtiter plate, and modified microtiter plate assays were 48.72%, 66.66%, and 73.72%, respec-tively. At least 92%of the biofilm forming isolates were multidrug resistant. Conclusions: Since most of the multidrug resistant strains produce biofilm, it seems necessary to provide continuous monitoring and determination of antibiotic susceptibility of clinical A. baumannii. This would help to select the most appropriate antibiotic for treatment.

Comparing the Coronal Flaring Efficacy of Five Different Instruments Using Cone-Beam Computed Tomography.

INTRODUCTION: Fearless removal of tooth structure during canal preparation and shaping has negative effects on the prognosis of treatment. On the other hand, sufficient pre-enlargement facilitates exact measurement of the apical size. The present in vitro study aimed to compare the efficacy of Gates-Glidden drills, K3, ProTaper, FlexMaster and RaCe instruments in dentin removal during coronal flaring using cone-beam computed tomography (CBCT). METHODS AND MATERIALS: A total of 40 mandibular molars were selected and the coronal areas of their mesiobuccal and mesiolingual root canals were randomly prepared with either mentioned instruments. Pre- and post-instrumentation CBCT images were taken and the thickness of canal walls was measured in 1.5- and 3-mm distances from the furcation area. Data were analyzed using the one-way ANOVA. Tukey's post hoc tests were used for two-by-two comparisons. RESULTS: At 1.5-mm distance, there was no significant difference between different instruments. However, at 3-mm distances, Gates-Glidden drills removed significantly more dentin compared to FlexMaster files (mean=0.18 mm) (P<0.02); however, two-by-two comparisons did not reveal any significant differences between the other groups. CONCLUSION: All tested instruments can be effectively used in clinical settings for coronal pre-enlargement.

A CBCT Assessment of Apical Transportation in Root Canals Prepared with Hand K-Flexofile and K3 Rotary Instruments.

INTRODUCTION: Apical transportation changes the physical shape and physiologic environment of the root canal terminus. The aim of the present experimental study was to determine the extent of apical transportation after instrumentation with hand K-Flexofile and K3 rotary instruments by means of cone-beam computed tomography (CBCT). METHODS AND MATERIALS: Forty mesiobuccal root canals of maxillary first molars, with 19-22 mm length and 20-40(°) canal curvature, were selected and assigned into two preparation groups. The first group was prepared with K-Flexofile with passive step-back technique and the second group was prepared with K3 rotary instruments. Pre and post instrumentation CBCT images were taken under similar conditions. The amount of root canal transportation was evaluated by Mann-Whitney U test and the chi-square test was used for the qualitative evaluation. RESULTS: The amounts of apical canal transportation with the K3 and K-Flexofile instruments were 0.105±0.088 and 0.150±0.127 mm, respectively with no statistically significant differences. In the manual technique, 25% of the canals had no apical transportation; while 30% of the canals in the K3 group were transportation free. CONCLUSION: Both systems were able to preserve the initial curvature of the canals and both had sufficient accuracy. Preparation with K3 rotary instruments resulted in apical transportation similar to that of K-Flexofile.

Cone-beam computed tomography for evaluation of apical transportation in root canals prepared by two rotary systems.

INTRODUCTION: Due to the importance of apical transportation during root canal preparation, the aim of the current study was to use cone-beam computed tomography (CBCT) to assess the extent of apical transportation caused by ProTaper and Mtwo files. METHODS AND MATERIALS: Forty extracted maxillary first molars with 19-22 mm length and 20-40 degrees of curvature were selected. The mesiobuccal canals were prepared using either Mtwo or ProTaper rotary files (n=20). CBCT images were obtained before and after canal preparation to compare the apical transportation in different cross-sections of mesial and distal surfaces. The apical transportation values were analyzed using the SPSS software. The results were compared with student's t-test and Mann-Whitney U test. RESULTS: There was no significant difference in the extent of apical transportation between Mtwo and ProTaper systems in different canal cross-sections. The apical transportation value was less than 0.1 mm in most of the specimens, which was clinically acceptable. CONCLUSION: Considering the insignificant difference between the two systems, it can be concluded that both system have low rates of apical transportation and can be assuredly used in clinical settings.

Histopathologic Responses of the Dental Pulp to Calcium-Enriched Mixture (CEM) and Mineral Trioxide Aggregate (MTA) in Diabetic and Non-Diabetic Rats.

Diabetes mellitus is a chronic disease which affects the healing ability of the pulp and periodontium. The aim of the present study was to assess the histopathologic response of dental pulp to pulp capping using MTA or CEM cement in diabetic rats. Thirty two Wistar male rats aged between 8 and 10 weeks (weight: 200-250g) were divided into two groups of diabetic (n=16) and healthy (n=16) animals and then subdivided into MTA and CEM subgroups. In each group, 10 MTA treated, 10 CEM treated and 12 intact (without any intervention) teeth were analyzed. Intact teeth were considered as a baseline inflammation control. Then, class I cavity was made in the maxillary first molars teeth with pinpoint pulpal exposure. Either MTA or CEM cement was then placed over exposed pulp as pulp capping agent and the cavities were restored using resin- modified glass ionomer cement. Both teeth of rats in subgroups remained intact without any intervention. After four weeks, the rats were sacrificed and the teeth were subjected to histological evaluation in terms of inflammation intensity, dentin bridge formation and dentin bridge continuity. The CEM cement treated diabetic rats exhibited a significant higher inflammatory response when compared to healthy control group (P=0.004) whereas, MTA treated diabetic rats did not exhibit a significant higher inflammatory response in comparison to healthy controls. There was no significant difference between MTA and CEM cement in the induction of dentin bridge formation in diabetic and healthy controls. This preliminary study suggests that MTA is a superior dental material than CEM cement for pulp therapy in subjects with diabetes.

Emulsification potential of a newly isolated biosurfactant-producing bacterium, Rhodococcus sp. strain TA6.

An indigenous biosurfactant producing bacterium, Rhodococcus sp. strain TA6 was isolated from Iranian oil contaminated soil using an efficient enrichment and screening method. During growth on sucrose and several hydrocarbon substrates as sole carbon source, the bacterium could produce biosurfactants. As a result of biosurfactant synthesis, the surface tension of the growth medium was reduced from 68mNm(-1) to values below 30mNm(-1). The biosurfactant was capable of forming stable emulsions with various hydrocarbons ranging from pentane to light motor oil. Preliminary chemical characterization revealed that the TA6 biosurfactant consisted of extracellular lipids and glycolipids. The biosurfactant was stable during exposure to high salinity (10% NaCl), elevated temperatures (120°C for 15min) and within a wide pH range (4.0-10.0). The culture broth was effective in recovering up to 70% of the residual oil from oil-saturated sand packs which indicates the potential value of the biosurfactant in enhanced oil recovery.

Sensitivity to detergents and plasmid curing in Enterococcus faecalis.

This research reports the sensitivity of a clinical isolate of Enterococcus faecalis to sodium N-lauroylsarcosinate (sarkosyl) and sodium dodecyl sulfate (SDS), as well as the efficiency of these detergents in curing the strain. Compared to Escherichia coli, Enterococcus faecalis was very sensitive to both detergents, with minimum inhibitory concentrations (MIC) for the latter being 100 times lower than for Escherichia coli. The clinical isolate of Enterococcus faecalis used in this study exhibited plasmid-borne resistance to kanamycin (MIC 2 mg/ml) and tetracycline (MIC 50 mug/ml); 3% curing was observed after growth in the presence of sarkosyl but no curing was observed after growth in the presence of either SDS or acridine orange. In contrast, 35% curing of plasmid-bearing Escherichia coli was observed after growth in the presence of either SDS or acridine orange, but none was observed after growth in the presence of sarkosyl.