Oral Isolates Research Articles

HPLC-MS identification and expression of Candida drug-resistance proteins from African HIV-infected patients.

The objective of this study was to elucidate the proteomic mechanisms of drug resistance in HIV-infected African patients. Cell membrane fractions from forty oral Candida isolates isolated from African HIV-positive patients were analysed using HPLC-MS with the aim of identifying proteins associated with their pathogenicity and drug resistance. Heat shock proteins that mediate the fungicidal activity of salivary peptides were found in all tested Candida fractions, with pH-responsive proteins associated with increased pathogenicity only being present in the three most commonly isolated species. ABC multidrug transporter efflux pumps and estrogen binding proteins were only found in C. albicans fractions, while ergosterol biosynthesis proteins were identified in four species. The combination of various adherence, invasion, upregulation and efflux pump mechanisms appear to be instrumental for the Candida host colonization and drug resistance emergence in HIV-infected individuals.

Kan ve Oral Kavite Örneklerinden Soyutlanan Candida albicans Suşlarının Fosfolipaz Aktivitelerinin Araştırılması

Objective: The aim of the present study was to investigate and compare the phospholipase B1, B2, C1 and D1 activities in C.albicans strains isolated from blood cultures and oral cavity specimens. Method: Phospholipase activity of the strains was examined by plate method and reverse transcriptase polymerase chain reaction (RT-PZT). Results: Twenty-six (86.7%) strains isolated from blood and 24 (80.0%) from oral cavities revealed phospholipase activity by plate method. No statistically significant difference (χ2 =0.48; p=0.49) was observed between the groups. However statistical difference was determined between the mean Pz values (0.6138±0.9823; 0.6988±0.9910) (p=0.007). PLB1 expression was detected in all (100%), PLB2 in 29 (96.7%) and 30 (100.0%), PLC1 in 27 (90%) and 22 (73.3%), PLD1 in 27 (90.0%) and 21 (70.0%) of blood and oral strains, respectively. While no significant difference was detected between PLB1 expression values of the groups (t=-0.307; p=0.760), PLB2 and PLC1 expressions were found to be significantly higher in blood (p=0.043) and oral cavity isolates (p<0.001), respectively. No difference was observed between PLD1 expressions (p=0.732). PLB1, PLB2, PLC1 and PLD1 expressions were 100%, 81.7%, 71.6% and 76.7% in agreement with the plate method. The agreement was 83.3% when all the phospholipase genes were considered. No correlation was detected between the Pz values and phospholipase expressions (p=0.602; p=0.555; p=0.241; p=0,096, respectively). Conclusion: The high rates of phospholipase activity of the C.albicans isolates in our study, support the important roles of these enzymes in virulence. Our results may indicate that phospholipase enzymes encoded by PLB2 and PLC1 genes play more important roles for invasive and oral cavity infections, respectively; however large scale studies are needed on this issue.

The genotypes and virulence attributes of C. albicans isolates from oral leukoplakia.

Background There is a debate as to whether some types of oral leucoplakias (OL) are caused by Candida species, and whether they contribute to the malignant transformation, associated with a minority of such lesions. As no detailed population analysis of yeast isolates from OL is available, we evaluated the virulence attributes, and genotypes of 35 C. albicans from OL, and compared their genotypes with 18 oral isolates from healthy individuals.Material and Methods The virulence traits evaluated were esterase, phospholipase, proteinase, haemolysin and coagulase production, and phenotypic switching activity, and yeast adherence and biofilm formation. DNA from OL and control yeasts were evaluated for A, B or C genotype status.Results Phospholipase, proteinase, and coagulase activity and biofilm formation was observed in 80%, 66%, 97 % and 77 % of the isolates, respectively. Phenotypic switching was detected in 8.6%, while heamolytic, and esterase activity and adherence were noted in all isolates.Conclusions The genotype A was predominant amongst both the OL and control groups. Due to the small sample size of our study a larger investigation to define the role of candidal virulent attributes in the pathogenicity of OL is warranted, and the current data should serve as a basis until then. Key words:C. albicans, oral cavity, leukoplakia, virulence factors, genotyping.

Comparison of the fungicidal efficacy of photodynamic therapy with methylene blue, silver nanoparticle, and their conjugation on oral Candida isolates using cell viability assay.

Background and Purpose: This study aimed to evaluate the effect of common photodynamic therapy and photodynamic therapy by the silver nanoparticle, methylene blue, and their combination on biofilm and plankton cells of standard oral Candida isolates using cell viability assay. Materials and Methods: In this in vitro study, biofilm and plankton cells of Candida species(i.e. C .albicans and C. parapsilosis) and plankton cells of Candida glabrata were treated with methylene blue, silver nanoparticle, and their combination once alone and then with the irradiation of total dose of 1.92 J/cm² for 60 sec. The minimum inhibitory concentration and antifungal activity of each approach were evaluated using the XTT assay. Results: After photodynamic therapy, methylene blue showed antifungal effect only on Candida albicans, while the antifungal effect of silver nanoparticles was increased on all Candida species. On the other hand, photodynamic therapy with the combination of methylene blue and silver nanoparticles did not have any effect on C. albicans. However, it reduced the minimum inhibitory concentration value of C. parapsilosis, and the most antifungal effect was observed on C. glabrata. Conclusion: Photodynamic therapy with photosensitizers can serve as a treatment modality in Candida infections of the oral cavity. Antifungal effect of photodynamic therapy was strain- and photosensitizer-dependent.

Comparative Study of Ethanolic Wild African Nutmeg (Pycnanthus angolensis (Welw.) Stem Bark Extract Potentials and Selected Conventional Toothpaste against Hidden Resident Mouth Cavity Microfora

The aim of the study is to evaluate and compare the antibacterial activity of ethanolic stem extract of (Wild African nutmeg) Pycnanthus angolensis (Welw.) and some commercially available toothpaste against bacteria isolated from the hidden resident mouth cavity microfora. Bacteria were isolated from swabs of apparently healthy individuals and were identified using Staining procedure biochemical tests and the use of Bergey’s manual of bacteria identification The assay for antibacterial activity of Pycnanthus angolensis stem bark extract and the four toothpastes were determined using agar well diffusion method. The Gram positive bacteria isolated were Streptococcus sangus, Streptococcus ratti, Stomatococcus mucilaginous., Peptostreptococcus sp., and Streptococcus mutans and the Gram negative bacteria were Veillonella atypical, Veillonella parvula, Veillonella dispar and Acidiaminococcus sp. Oral B toothpaste showed maximum efficacy of inhibition with inhibition zone diameter as wide as 20 mm at 100 mg/ml. Percentage frequency distribution of antibacterial activity of conventional toothpaste (Close-up) against hidden resident mouth cavity microfora depicts Acidaminococcus sp.13%, Veillonella parvula (10%), Veillonella dispar (12%), Peptostreptococcus sp.(12%), Stomatococcus mucilaginous.(9%), Streptococcus ratti (13%), Veillonella atypical (11%), Streptococcus sangus (9%) and Streptococcus mutans (11%), Percentage frequency distribution of antibacterial activity of conventional toothpaste (Oral B toothpaste) against hidden resident mouth cavity microfora reveals Acidaminococcus sp.(11%,) Veillonella dispar (11%), Veillonella parvula (10%), Peptostreptococcus sp. (12%), Stomatococcus mucilaginous.(15%), Streptococcus ratti (11%), Veillonella atypical (8%), Streptococcus sangus (10%), and Streptococcus mutans (12%), Percentage frequency distribution of antibacterial activity of conventional toothpaste (MyMy toothpaste) against hidden resident mouth cavity microfora depicts Acidaminococcus sp.(12%), Veillonella dispar (9%), Veillonella parvula (8%), Peptostreptococcus sp.(10%), Stomatococcus mucilaginous.(16%), Streptococcus ratti (9%), Veillonella atypical (15%),Streptococcus sangus (9%) and Streptococcus mutans (12%), Percentage frequency distribution of antibacterial activity of conventional toothpaste (Olive toothpaste) against hidden resident mouth cavity microfora shows Acidaminococcus sp.(9%), Veillonella dispar (10%), Veillonella parvula (10%), Peptostreptococcus sp.(12%), Stomatococcus mucilaginous.(13%), Streptococcus ratti (10%) ,Veillonella atypical (17%), Streptococcus sangus (7%), and Streptococcus mutans (12%). Pycnanthus Angolensis stem bark extract inhibited the growth of the oral bacterial isolates with of zones of inhibition diameter ranging from 6 mm to 17 mm at a concentration of 100mg/ml. Secondary metabolite (Phytochemical) screening shows the presence of flavonoids, tannins, saponins, alkaloids, reducing sugars, steroid, phenol, terpenoid, pyrrolozidine alkaloid, glycoside and cardiac glycoside with glycoside and terpenoid most present. However, anthraquinones and volatile oil were absent. With menial antibacterial activity, P. angolensis can be use in the formulation of herbal toothpaste. It should be advocated that Pycnanthus angolensis should be added to our convention toothpaste to improve the functional ingredient of the toothpaste and Plant-based traditional knowledge has become a recognized tool in search for new sources of drugs. It is clear that the use of these herbal plants can offer a platform for further research.

Reliability of MALDI-TOF mass spectrometry to identify oral isolates of Streptococcus salivarius and Lactobacillus spp

ObjectiveThe aim of this study is to evaluate the performance of MALDI-TOF mass spectrometry in identifying bacteria isolated in the oral cavity known to be of probiotic interest. DesignWe evaluated Bruker MALDI Biotyper for the identification of 92 clinical oral isolates of probiotic interest (31 Streptococcus salivarius and 61 Lactobacillus spp.) by comparing direct colony method with on-plate formic acid extraction. Isolates were previously identified by use of biochemical methods and molecular biology. ResultsUsing the manufacturer’s suggested genus and species level cutoff scores, the direct colony method identified 42 (45.7%) isolates at the genus level and 35 (38%) at the species level while the on-plate extraction method correctly identified 90 (97.8%) isolates at the genus level and 82 (89.1%) at the species level. The difference between the two methods was statistically significant at the genus and species levels (P ≤ 0.0001). After dividing the isolates into two subgroups, the analysis was repeated. The direct colony method identified correctly all isolates of Streptococcus salivarius at the species level. In contrast, the direct colony method allowed the identification of only 11 (18%) lactobacilli at the genus level and 4 (6.6%) at the species level. The on-plate extraction method was statistically (P ≤ 0.0001) more efficient since 59 (96.7%) lactobacilli were identified at the genus level and 51 (83.6%) at the species level. ConclusionsMALDI Biotyper can efficiently identify Streptococcus salivarius regardless of the preparative method but on-plate extraction is superior to direct colony method for the identification of lactobacilli.

Phytocompounds vs. Dental Plaque Bacteria: In vitro Effects of Myrtle and Pomegranate Polyphenolic Extracts Against Single-Species and Multispecies Oral Biofilms.

In the last decades, resistant microbial infection rate has dramatically increased, especially infections due to biofilm-producing strains that require increasingly complex treatments and are responsible for the increased mortality percentages compared with other infectious diseases. Considering that biofilms represent a key factor for a wide range of chronic infections with high drug tolerance, the treatment of biofilm-causing bacterial infections represents a great challenge for the future. Among new alternative strategies to conventional antimicrobial agents, the scientific interest has shifted to the study of biologically active compounds from plant-related extracts with known antimicrobial properties, in order to also evaluate their antibiofilm activity. In this regard, the aim of this study has been to assess the antibiofilm activity of polyphenolic extracts from myrtle leaf and pomegranate peel against oral pathogens of dental plaque, an excellent polymicrobial biofilm model. In particular, the in vitro antibiofilm properties of myrtle and pomegranate extracts, also in binary combination, were highlighted. In addition to inhibiting the biofilm formation, the tested polyphenolic extracts have been proven to destroy both preformed single-species and multispecies biofilms formed by Streptococcus mutans, Streptococcus oralis, Streptococcus mitis, and Rothia dentocariosa oral isolates, suggesting that the new natural sources are rich in promising compounds able to counteract biofilm-related infections.

Choosing the Right Essential Oil for a Mouthwash: Chemical, Antimicrobial and Cytotoxic Studies.

Thirteen commercial essential oils were assessed for their possible inclusion in a mouthwash formulation based on their inhibitory effect against potentially pathogenic anaerobic oral bacterial isolates from subgingival plaque, and their cytotoxicity towards gingival cells. The essential oils, originating from species belonging to seven major aromatic plant families, were chosen to provide the necessary diversity in chemical composition that was analyzed in detail by GC and GC/MS. Multivariate statistical analysis, performed using the in vitro microbiological/toxicological assays and compositional data, revealed that the major components of the essential oils were probably not the main carriers of the activities observed. A formulation of 'designer' mouthwashes is proposed based on the selective action of certain essential oils towards specific bacterial isolates (e. g., Citrus bergamia vs. Parvimonas micra), and non-toxicity to gingival cells at antimicrobially active concentrations.

Intra-species variation within Lactobacillus rhamnosus correlates to beneficial or harmful outcomes: lessons from the oral cavity

BackgroundThe origin of most of the Lactobacillus rhamnosus genome sequences lodged in NCBI can be traced to food and faecal isolates followed by blood and tissue sites but with minimal representation from oral and vaginal isolates. However, on the L. rhamnosus phylogenetic tree no apparent clade is linked to the origin of isolation or to the relevant clinical source, except for a distinct clade exclusively shared by L. rhamnosus isolates from early stages of dental pulp infection (LRHMDP2 and LRHMDP3) and from bronchoalveolar lavage (699_LRHA and 708_LRHA) from a critical care patient. These L. rhamnosus strains, LRHMDP2, LRHMDP3, 699_LRHA and 708_LRHA isolated from different continents, display closest genome neighbour gapped identity of 99.95%. The aim of this study was to define a potentially unique complement of genes of clinical relevance shared between these L. rhamnosus clinical isolates in comparison to probiotic L. rhamnosus strains.ResultsIn this analysis we used orthologous protein identification tools such as ProteinOrtho followed by tblastn alignments to identify a novel tyrosine protein phosphatase (wzb)-tyrosine-protein kinase modulator EpsC (wzd)- synteny exopolysaccharide (EPS) cluster. This EPS cluster was specifically conserved in a clade of 5 clinical isolates containing the four L. rhamnosus clinical isolates noted above and Lactobacillus spp. HMSC077C11, a clinical isolate from a neck abscess. The EPS cluster was shared with only two other strains, L. rhamnosus BPL5 and BPL15, which formed a distant clade on the L. rhamnosus phylogenetic tree, with a closest genome neighbour gapped identity of 97.51% with L. rhamnosus LRHMDP2 and LRHMDP3.Exclusivity of this EPS cluster (from those identified before) was defined by five EPS genes, which were specifically conserved between the clade of 5 clinical isolates and L. rhamnosus BPL5 and BPL15 when compared to the remaining L. rhamnosus strains. Comparative genome analysis between the clade of 5 clinical isolates and L. rhamnosus BPL5 and BPL15 showed a set of 58 potentially unique genes characteristic of the clade of 5.ConclusionThe potentially unique functional protein orthologs associated with the clade of 5 clinical isolates may provide understanding of fitness under selective pressure.

Spectrum and resistance determinants of oral streptococci clinical isolates.

The profiles of oral streptococci sensitivity to antibacterial drugs may reflect information about the presence of macroorganism resistance determinants. The aim of the work was to isolate the spectrum of oral streptococci from the microbiota of the oral cavity of patients and to determine their sensitivity to a wide range of antibiotics. A total of 342 microbial streptococcal isolates were isolated from saliva samples and a periodontal pocket and tested for antibiotic sensitivity. Species identification of streptococci was carried out using biochemical API test systems. Evaluation of antibiotic resistance was performed using E-tests. Real-time PCR was used to identify the presence of tetracycline and macrolide resistance genes. The study identified six types of oral streptococci: S. oralis, S. salivarius, S. mitis, S. sanguinis, S. anginosus and S. mutans. All streptococci were sensitive to linezolid and meropenem. The proportion of penicillin-resistant streptococci in the subgroup S. oralis / mitis / mutans was 47,8% versus 23,5% in the subgroup S. salivarius / sanguinis / anginosus (p = 0.020). Significant levels of resistance were revealed to macrolides (erythromycin) - 47,9%, tetracyclines (tetracycline) - 44,4% and quinolones (ofloxacin) - 41%. Multiple drug resistance (MDR) was detected in 31,9% of oral streptococcal isolates, a combination of erythromycin, tetracycline and ofloxacin resistance was prevalent in 79 isolates (23,1%). The most common genotypes of macrolides and tetracycline resistant oral streptococci (in 127 streptococcal isolates with combined resistance) were ermB-mefE + and tetM + tetQ-, respectively. Thus, S. oralis / mitis / mutans group streptococci predominated in the structure of antibiotic-resistant oral streptococci, including MDR. So, being in one of the most densely populated biotopes of a macroorganism, oral streptococci can mediate the transfer of resistance determinants to more pathogenic and clinically significant microorganisms, which requires careful monitoring of their level of susceptibility to antimicrobial agents.

The effect of benzyl isothiocyanate on Candida albicans growth, cell size, morphogenesis, and ultrastructure

Candida albicans is a commensal yeast that may become pathogenic and even lethal to the host. Over the last few decades, antifungal resistance has increased, promoting screening of the antifungal potential of old and new substances. This study investigates the antifungal potential of isothiocyanates (ITCs) against C. albicans oral isolates. A preliminary susceptibility disk diffusion test (DD) was performed using allyl isothiocyanate (AITC), benzyl isothiocynanate (BITC) and phenyl ethyl isothiocyanate (PEITC) at a fixed concentration range (0.001-0.1M). Because C. albicans isolates were more susceptible to BITC and PEITC, their effect on cell size and on germ tube formation (GTF) were tested. The most promising molecule, BITC, was further tested for effects on cell viability, oxidative stress and for ultrastructure. ITCs, especially the aromatic ones, had a significant type-, dose- and isolate-dependent anti-Candida activity. Although BITC and PEITC had similar activity against the yeast cells, BITC had a more pronounced effect on cell size and GTF. Furthermore, BITC appears to induce oxidative stress and promote changes in the cell ultrastructure, interfering with cell wall structure. Our work showed that aromatic ITCs have the potential to effect C. albicans cells in multiple ways, including size, shape and GTF (BITC and PEITC), oxidative stress, and ultrastructure (BITC). Overall, our results suggest that BITC may be effectively used against C. albicans to modulate its growth, and control or suppress its invasive potential.

Correlative antimicrobial assay of various foreign toothpaste against oral microbial isolates

The in-vitro antimicrobial assay of four (4) foreign toothpastes sold in Nigeria viz; Sensodyne®, Raiya®, Mentadent® and Colgate® were carried out using agar well diffusion method against eight (8) oral clinical isolates which include the strains of Streptococcus mutans, Brahamella catarrhalis, Veillonella spp, Micrococcus luteus, Lactobacillus spp, Staphylococcus aureus, Streptococcus pneumoniae, and Candida albicans. The statistical analysis was done using SPSS windows, by applying mean value, T-test, analysis of variance (ANOVA) with Post-hoc least significant differences (LSD) method (α = 0.05). The result showed that based on the mean diameter of the zone of microbial inhibition produced by the toothpaste in agar well diffusion method, Mentadent® showed maximum zone of inhibition against the entire test organisms with Streptococcus pneumoniae being the most inhibited (sig< 0.01) compared to all other toothpaste formulations. Colgate® gave the least antimicrobial activity, whereas Micrococcus luteus was the least inhibited among the eight (8) isolates. Although all the toothpaste selected for this study had antimicrobial properties, Mentadent® distinguish itself because of its improved formulation of 2 % zinc citrate. There is an urgent need for toothpaste manufacturers to periodically review their formulations in order to improve the oral health of its users.

Correlative antimicrobial assay of various foreign toothpaste against oral microbial isolates

Correlative antimicrobial assay of various foreign toothpaste against oral microbial isolates

Comprehensive analysis of transcriptional profiles in oral epithelial-like cells stimulated with oral probiotic Lactobacillus spp.

ObjectiveThe mechanisms of action of probiotics can vary among species and among strains of a single species; thus, they can affect host cells in a complex manner. In the present study, Lactobacillus spp. were evaluated for their ability to adhere to gingival epithelial-like cells. Comprehensive analyses of transcriptional profiles of mouse gingival epithelial GE1 cells treated with L. rhamnosus L8020 were performed to assess the putative in vivo probiotic potential of this strain. MethodsFive Lactobacillus spp., isolated from the oral cavity, traditional Bulgarian yoghurt, and the feces of a healthy human, were each co-cultured with GE1 cells. Adhesion assays with serial dilution plating and DNA microarray analysis were performed to identify differentially expressed genes (DEGs) in GE1 cells grown in co-culture with L. rhamnosus L8020. ResultsThe oral isolates L. rhamnosus L8020, L. casei YU3, and L. paracasei YU4 demonstrated significantly greater adhesion compared with the non-oral isolates. In total, 536 genes in GE1 cells exhibited more than twofold upregulation or downregulation, compared with the 0 h timepoint, during co-culture with L. rhamnosus L8020. Gene ontology enrichment analysis revealed that DEGs were differentially enriched in a time-dependent manner. Early responses involved widespread changes in gene expression. ConclusionsThis study reveals changes in expression of genes involved in the epithelial physical barrier and immune response in gingival epithelial-like cells co-cultured with L. rhamnosus L8020. Further investigations regarding the molecular mechanisms by which L. rhamnosus L8020 serves as a probiotic may provide evidence to support clinical use.

THE EVALUATION OF THE COLONIZATION AND BIOFILM FORMATION CAPACITY OF THE CANDIDA SPECIES ISOLATED FROM DENTURE WEARER PATIENTS

Aim: C Aim: Candida species, although being a member of the oral flora, may exhibit pathological features under certain conditions. The aim of this study was to determine the distribution and biofilm production of Candida species that were isolated from denture related stomatitis (DRS) patients and to compare with healthy subjects. andida species, although being a member of the oral flora, may exhibit pathological features under certain conditions. The aim of this study was to determine the distribution and biofilm production of Candida species that were isolated from denture related stomatitis (DRS) patients and to compare with healthy subjects. Materials and Methods: The 56 non-smoker, systemically healthy, maxillary removable denture wearer subjects included in this study and diagnosed as DRS(N=27) and healthy(N=29) . Samples from the palatal mucosal surface of patients were collected by sterile swabs during 20 seconds. Cultivation and selective isolation of Candida species were performed on CHROMagar (CHROMagar ® Candida, CHROMagar, Paris, France) at 37ºC for 2 days. Isolated Candida species were identified with API ID 32 C (bioMérieux ® , France). Biofilm formation by Candida species were determined by a visual tube method. Results: The prevalence of the yeasts in the healthy group was found to be 37.94%, while in the DRS group it was 100% (p<0.001). The isolation rate of C. albicans in patients with DRS significantly higher (p<0.001). Biofilm formation was observed in a total of 37 oral yeast isolates, 9 isolates from healthy denture wearers and 28 isolates from DRS patients. The number of C. albicans and C. glabrata isolates showing biofilm formation ability in the DRS group was significantly higher than the healthy group (p<0.05). Conclusions: Within the limitations of this study, our results suggest that Candida species play a major role on the development of DRS. While C. albicans was the most isolated species in DRS, C. glabrata was as important as C. albicans on the pathogenesis of DRS. keywords: Biofilms , Candida, denture stomatitis, mycology, oral pathology HAREKETLİ PROTEZ KULLANAN HASTALARDA KANDİDA TÜRLERİNİN KOLONİZASYONUNUN VE BİYOFİLM OLUŞTURMA KAPASİTELERİNİN DEĞERLENDİRİLMESİ Öz Amaç: Candida türleri, oral floranın doğal üyesi olmakla birlikte, belirli koşullar altında patolojik özellikler sergileyebilir. Bu çalışmanın amacı, protez stomatiti (PS) olan hastalardan ve protez kullanan sağlıklı bireylerden elde edilen Candida türlerinin dağılımını ve biyofilm üretime kapasitelerini karşılaştırmaktır Gereç ve Yöntem: Sigara içmeyen, sistemik olarak sağlıklı, maksiller tam damak protez kullanan 56 kişi bu çalışmaya dahil edildi. Katılımcılar PS (N = 27) ve sağlıklı (N = 29) olmak üzere gruplandırıldı. Hastaların palatal mukoza yüzeyinde 20 saniye boyunca steril swablar gezdirilerek sürüntü örnekleri elde edildi. Candida türlerinin kültivasyon ve seçici izolasyonu 2 gün boyunca 37 ° C'de CHROMagar (CHROMagar® Candida, CHROMagar, Paris, Fransa) besiyerinde gerçekleştirildi. İzole edilen Candida türlerinin identifikasyonu API ID 32 C (bioMérieux®, Fransa) ile gerçekleştirildi. Candida türlerinin biyofilm oluşturma potansiyelleri görsel tüp yöntemi ile saptandı Bulgular: Sağlıklı gruptaki mayaların prevalansı % 37.94 iken, PS grubunda % 100'dü ( p <0.001). PS'li hastalarda C. albicans 'ın izolasyon oranı anlamlı derecede daha yüksekti ( p <0.001). Sağlıklı katılımcılardaki 9 PS katılımcılardaki 28 izolatta biyofilm oluşumu gözlendi. PS grubunda biyofilm oluşumunu gösteren C. albicans ve C. glabrata izolatlarının sayısı sağlıklı gruba göre anlamlı olarak yüksekti ( p <0.05). Sonuç: Bu çalışmanın sınırları dahilinde, sonuçlarımız Candida türlerinin PS'nin gelişiminde önemli bir rol oynadığını göstermektedir. C. albicans , PS'de en sık izole edilen tür olsa da C. glabrata da PS'nin patogenezinde C. albicans kadar önemlidir. Anahtar Kelimeler: Biyofilm , Kandida, protez stomatitisi, mikoloji, oral patoloji

Antifungal susceptibility of oral isolates of Candida species from chronic kidney disease patients on chronic dialysis

Chronic renal disease patients under chronic dialysis (CRDD) have a multifactorial immunological deterioration with an increased risk of Candida infections. Incidence of Candida infections is increasing. Choice of suitable antifungal agents is limited due to the resistance of some species to several antifungals. Aim of the present study was to identify the distribution and antifungal susceptibility patterns of oral isolated Candida species from infected and colonized patients, as well as to investigate the risk factors for oral infection in patients on dialysis. Cross-sectional study, approved by the institutional bioethics committees was performed in CRDD patients. Demographic, clinic data, and oral mucosa samples were obtained. Infection diagnosis was established clinically and confirmed with exfoliative cytology, each sample was plated on CHROMagar Candida and incubated at 36°C for 2 days. Yeast species were identified by carbohydrate assimilation ID 32C AUX system and the apiweb database. For the antifungal susceptibility test, the M44 A-3 method (CLSI) using fluconazole (FCZ), miconazole (MCZ), nystatin (NYS), and voriconazole (VCZ). Study included 119 participants, the main cause of CRD was nephropathy due to DM2 (58%), and three-fourths of the patients were under hemodialysis. Candida prevalence was 56.3% of 67 colonized or infected patients, 88 isolates were obtained. Principal identified species were C. albicans (51.1%), C. glabrata (25%), and C. tropicalis (14.8%). C. glabrata showed a reduced response to FCZ in 50% of isolates and C. albicans had a reduced response in 16% of the isolates. Antifungal agent with the least efficacious response or with the lowest susceptibility in the isolates of these patients was MCZ, followed by VCZ and FCZ, whereas NYS induced the best antifungal response.

Linden tea from Serbia – an insight into the phenolic profile, radical scavenging and antimicrobial activities

Linden tea has been used in Serbian folk medicine for centuries to induce sweating for colds, relieve throat irritation and cough, reduce blood pressure, as well as a diuretic, spasmolytic, and as a sedative. The main aim of this study was to determine the comprehensive metabolite profile, free radical scavenging activity, and antimicrobial activities against 23 human and plant pathogens of both commercial and field-collected linden tea samples from Serbia. Developed high-performance thin-layer chromatography (HPTLC) method allowed identification of eight major metabolites in investigated samples, while forty-six metabolites were tentatively identified using high resolution mass spectrometry (HRMS): 14 phenolic acids and their derivatives, 15 glycosides, 9 flavonoids, and 8 procyanidins. Both commercial and field-collected linden tea samples showed similar total phenolic content and radical scavenging activity. Seven compounds such as p-hydroxybenzoic acid 4-O-hexoside, caffeic acid cinnamyl ester, pinocembrin, galangin, luteolin 7-O-glucuronide (and its isomer), isorhamnetin hexosyl hexoside were found in Tilia samples for the first time. Radical reagent 2,2-Diphenyl-1-picrylhydrazyl· (DPPH·) was used in DPPH-HPTLC assay which identified chlorogenic acid, caffeic acid, astragalin, quercitrin, tiliroside, and rutin as compounds that exhibit radical scavenging activity. Tilia samples showed inhibitory effect overwhelming only on Gram-positive bacteria, especially on Bacillus subtilis with the lowest MIC values observed, as well as towards Staphylococcus aureus and oral cavity isolates of Streptococcus pyogenes and Streptococcus mutans. Vaginal isolate of Candida glabrata showed more susceptibility than Candida albicans isolate. Generally, extract from Tilia cordata Miller (L4) showed the highest antimicrobial activity against the most of the tested pathogens, among all field-collected Tilia samples.

PepO is a target of the two-component systems VicRK and CovR required for systemic virulence of Streptococcus mutans

ABSTRACT Streptococcus mutans , a cariogenic species, is often associated with cardiovascular infections. Systemic virulence of specific S. mutans serotypes has been associated with the expression of the collagen- and laminin-binding protein Cnm, which is transcriptionally regulated by VicRK and CovR. In this study, we characterized a VicRK- and CovR-regulated gene, pepO, coding for a conserved endopeptidase. Transcriptional and protein analyses revealed that pepO is highly expressed in S. mutans strains resistant to complement immunity (blood isolates) compared to oral isolates. Gel mobility assay, transcriptional, and Western blot analyses revealed that pepO is repressed by VicR and induced by CovR. Deletion of pepO in the Cnm+ strain OMZ175 (OMZpepO) or in the Cnm− UA159 (UApepO) led to an increased susceptibility to C3b deposition, and to low binding to complement proteins C1q and C4BP. Additionally, pepO mutants showed diminished ex vivo survival in human blood and impaired capacity to kill G. mellonella larvae. Inactivation of cnm in OMZ175 (OMZcnm) resulted in increased resistance to C3b deposition and unaltered blood survival, although both pepO and cnm mutants displayed attenuated virulence in G. mellonella. Unlike OMZcnm, OMZpepO could invade HCAEC endothelial cells. Supporting these phenotypes, recombinant proteins rPepO and rCnmA showed specific profiles of binding to C1q, C4BP, and to other plasma (plasminogen, fibronectin) and extracellular matrix proteins (type I collagen, laminin). Therefore this study identifies a novel VicRK/CovR-target required for immune evasion and host persistence, pepO, expanding the roles of VicRK and CovR in regulating S. mutans virulence.

Tolerance and Persister Formation in Oral Streptococci.

The aim of this study was to analyze the potential influence of long-term exposure in subinhibitory concentrations of chlorhexidine on the emergence of tolerant and/or persistent cells in oral streptococci. The two oral streptococcal isolates S. mutans ATCC25175 and S. sobrinus ATCC33402 were incubated, after long-term subinhibitory exposure to chlorhexidine, in liquid growth media containing high concentrations of chlorhexidine. A distinct subpopulation of more chlorhexidine-tolerant cells could be detected in streptococci that had been previously exposed to subinhibitory concentrations of chlorhexidine but not in the control strains. These more biocide-tolerant and persisting microbial subpopulations might also arise in vivo. Therefore, the rational and proper use of antimicrobials in dentistry, especially when used over a long period of time, is crucial.

Chemical composition and in vitro activity of Origanum vulgare L., Satureja hortensis L., Thymus serpyllum L. and Thymus vulgaris L. essential oils towards oral isolates of Candida albicans and Candida glabrata

AbstractThe purpose of this research was to investigate the chemical composition of essential oils (EOs) from:Origanum vulgareL.,Satureja hortensisL.,Thymus serpyllumL. andThymus vulgarisL. (Lamiaceae) cultivated in Poland, and to study their antifungal activity towards clinical isolates of oralCandidaspp. The hydrodistilled essential oils were analyzed using the GC-MS method. The antifungal activity was evaluatedin vitroagainst oral isolates and reference strains ofCandida albicansandC. glabrata, using the broth microdilution method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Clinical and Laboratory Standards Institute (CLSI) guidelines, allowing for estimation of minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC). GC-MS analysis revealed that carvacrol was the main EO compound in oregano and summer savory, while thymol and linalool were the major ingredients of thyme and wild thyme oils, respectively. The EOs possessed fungicidal activity againstC. albicansandC. glabrata, including oral isolates, with MIC = 125 – 2000 mg/L, MFC = 250 – 4000 mg/L and MFC/MIC = 1 – 4, depending on the yeast and plant species. The most active was thyme oil – with MIC = 125 – 500 mg/L, MFC = 250 – 500 mg/L and MFC/MIC = 1 – 2.