A SARS-CoV-2 full genome sequence of the B.1.1 lineage sheds light on viral evolution in Sicily in late 2020.

To investigate the influence of geographic constrains to mobility on SARS-CoV-2 circulation before the advent of vaccination, we recently characterized the occurrence in Sicily of viral lineages in the second pandemic wave (September to December 2020). Our data revealed wide prevalence of the then widespread through Europe B.1.177 variant, although some viral samples could not be classified with the limited Sanger sequencing tools used. A particularly interesting sample could not be fitted to a major variant then circulating in Europe and has been subjected here to full genome sequencing in an attempt to clarify its origin, lineage and relations with the seven full genome sequences deposited for that period in Sicily, hoping to provide clues on viral evolution. The obtained genome is unique (not present in databases). It hosts 20 single-base substitutions relative to the original Wuhan-Hu-1 sequence, 8 of them synonymous and the other 12 encoding 11 amino acid substitutions, all of them already reported one by one. They include four highly prevalent substitutions, NSP12:P323L, S:D614G, and N:R203K/G204R; the much less prevalent S:G181V, ORF3a:G49V and N:R209I changes; and the very rare mutations NSP3:L761I, NSP6:S106F, NSP8:S41F and NSP14:Y447H. GISAID labeled this genome as B.1.1 lineage, a lineage that appeared early on in the pandemic. Phylogenetic analysis also confirmed this lineage diagnosis. Comparison with the seven genome sequences deposited in late 2020 from Sicily revealed branching leading to B.1.177 in one branch and to Alpha in the other branch, and suggested a local origin for the S:G118V mutation.

Conocimiento sobre odontología legal y la normativa sanitaria por parte de los odontólogos y estomatólogos de la Comunidad Valenciana / Knowledge of legal dentistry and of health regulations by the Dentists and Stomatologists of the Valencian Community

Objetivos: el objetivo principal de este estudio es analizar los conocimientos que poseen los odontólogos y los estomatólogos colegiados en Alicante, Castellón y Valencia, las 3 provincias que conforman la Comunidad Valenciana, acerca de odontología legal y sobre la legislación y normativa sanitaria actual, e intentar cuantificar la relación entre el nivel de formación, la especialidad, la experiencia laboral, el lugar y la figura en el trabajo con el grado de conocimiento.Material y métodosse diseñó y validó una encuesta anónima, de 33 preguntas tipo test. Esta encuesta incluía cuestiones sobre el perfil profesional y preguntas clave acerca del derecho dental actual, ciencias forenses y peritación.Resultadosel nivel de conocimiento es moderado, con solo el 63% de las preguntas respondidas correctamente. Ni el nivel de formación ni la experiencia laboral se correlacionan significativamente con el grado de conocimiento. Los dentistas especializados en odontología general y estética resultaron ser más conocedores de cuestiones éticas. Con respecto al lugar de trabajo, los docentes universitarios tienden a diferenciarse del resto de profesionales en cuanto a los conocimientos legales y éticos, mientras que el perfil de autónomo y empleado muestra un nivel de conocimientos inferior que el resto de la muestra.Conclusioneslas conclusiones obtenidas de este estudio ponen de relieve la necesidad que tienen los dentistas de completar y actualizar sus conocimientos en materia legislativa sanitaria actual y en odontología legal, pues un buen conocimiento significa una garantía para evitar posibles problemas legales, lo que no solo supone una mejor defensa del odontólogo, sino una garantía para la adecuada atención al paciente. (AU)

Objectives: The main objective of this study is to analyze the knowledge of the dentists and stomatologists registered in Alicante, Castellón and Valencia, the three provinces that make up the Valencian Community, in Spain, with regard to legal dentistry and the current health legislation and regulations, and to quantify the relationship between level of training, specialization, work experience, position and workplace with the degree of knowledge.Material and methodsAn anonymous survey was designed and validated, consisting of thirty-three multiple-choice questions. The survey included questions regarding their professional profile and key questions regarding current dental law, forensic science and expertise.ResultsThe level of knowledge is moderate, with only 63% of the questions answered correctly. Neither their level of training nor work experience correlates significantly with their degree of knowledge. Dentists specialized in General and Aesthetics Dentistry were found to be more knowledgeable about ethical issues. Regarding the workplace, those working as university teachers tended to stand out from the other professionals in terms of legal and ethical knowledge whilst self-employed and employee dentists show a lower level of knowledge than the rest of the sample.ConclusionsThe conclusions obtained from this study highlight the need for dentists to complete and update their knowledge of legal dentistry and current health legislation, as having proper knowledge is a means of avoiding possible legal problems, which not only means better legal protection for the dentist but is also a means of guaranteeing adequate patient care. (AU)

Pilot Investigation of SARS-CoV-2 Variants in the Island of Sicily Prior to and in the Second Wave of the COVID-19 Pandemic.

After 2 years of the COVID-19 pandemic, we continue to face vital challenges stemming from SARS-CoV-2 variation, causing changes in disease transmission and severity, viral adaptation to animal hosts, and antibody/vaccine evasion. Since the monitoring, characterization, and cataloging of viral variants are important and the existing information on this was scant for Sicily, this pilot study explored viral variants circulation on this island before and in the growth phase of the second wave of COVID-19 (September and October 2020), and in the downslope of that wave (early December 2020) through sequence analysis of 54 SARS-CoV-2-positive samples. The samples were nasopharyngeal swabs collected from Sicilian residents by a state-run one-health surveillance laboratory in Palermo. Variant characterization was based on RT-PCR amplification and sequencing of four regions of the viral genome. The B.1.177 variant was the most prevalent one, strongly predominating before the second wave and also as the wave downsized, although its relative prevalence decreased as other viral variants, particularly B.1.160, contributed to virus circulation. The occurrence of the B.1.160 variant may have been driven by the spread of that variant in continental Europe and by the relaxation of travel restrictions in the summer of 2020. No novel variants were identified. As sequencing of the entire viral genome in Sicily for the period covered here was restricted to seven deposited viral genome sequences, our results shed some light on SARS-CoV-2 variant circulation during that wave in this insular region of Italy which combines its partial insular isolation with being a major entry point for the African immigration.

Icariin in Combination with Amoxycillin-Clavulanate and Ampicillin, but Not Vancomycin, Increases Antibiotic Sensitivity and Growth Inhibition against Methicillin-Resistant Staphylococcus aureus.

The widespread irrational use of antibiotics in recent years has resulted in an increase in the detection of multi-resistant bacterial strains, particularly methicillin-resistant Staphylococcus aureus (MRSA). The use of natural derivatives such as flavonoids is postulated as one of the most promising avenues to solve this emerging public health problem. The objective of the present study is to characterize the antimicrobial activity of icariin, a flavonoid compound isolated from a variety of plants of the Epimedium genus, against human and animal clinical MRSA isolates. Our study found that icariin alone did not have any antimicrobial effect on S. aureus or MRSA clinical isolates. However, icariin enhanced the effect of amoxycillin-clavulanate or ampicillin, whereas no effect was seen when used in combination with vancomycin. Specifically, co-incubation of S. aureus with amoxycillin-clavulanate plus icariin resulted in an increased proportion of dead cells, suggesting that this flavonoid potentially increases antimicrobial activity when used in combination with the beta-lactam antibiotic amoxycillin-clavulanate. Furthermore, we demonstrate that co-incubation of S. aureus with AmoxyClav plus icariin resulted in increased membrane disruption and growth inhibition. This study demonstrates the potential utility of icariin in permitting lower antibiotic therapeutic doses in alignment with strategies to reduce the spread of antibiotic resistance. Further research is required to determine the optimum concentration of icariin and to define clinically relevant combinations of flavonoid and antibiotic.

Salivary biomarkers and their efficacies as diagnostic tools for Oral Squamous Cell Carcinoma: Systematic review and meta-analysis.

BACKGROUND: More than 90% of malignant tumors of the head and neck are oral squamous cell carcinomas. Early detection of oral squamous cell carcinoma using salivary biomarkers could prevent malignant transformations and enhance patient survival. METHODS: A systematic search in MEDLINE and the Central Register of Controlled Trials and meta-analysis were undertaken to identify the screening potential of six salivary biomarkers for early oral squamous cell carcinoma detection: interleukins IL-8 and IL1-ß, DUSP-1 and S100P messenger RNAs, and miR125a and miR200a microRNAs. RESULTS: The sensitivities of IL-8 (0.41; 95%CI 0.19-0.99), IL1-ß (0.26; 95%CI 0.19-0.99), DUSP-1 (0.61; 95%CI 0.01-0.98), and S100P (0.67; 95%CI 0.32-0.99) were calculated. Specificities of the biomarkers analyzed were found to be IL-8 (0.69; 95%CI 0.66-0.99), IL1-ß (0.47; 95%CI 0.46-0.90), DUSP-1 (0.75; 95%CI 0.33-1), and S100P (0.73; 95%CI 0.18-0.99). CONCLUSIONS: Early detection of oral squamous cell carcinoma was best achieved by screening for salivary messenger RNA DUSP-1 and S100P. Further investigation is required into miRNAs as novel biomarkers.

Herbal Extracts with Antifungal Activity against Candida albicans: A Systematic Review.

In the era of antimicrobial resistance, fungal pathogens are not an exception. Several strategies, including antimicrobial stewardship programs and high throughput screening of new drugs, are being implemented. Several recent studies have demonstrated the effectiveness of plant compounds with antifungal activity. In this systematic review, we examine the use of natural compounds as a possible avenue to fight fungal infections produced by Candida albicans, the most common human fungal pathogen. Electronic literature searches were conducted through PubMed/MEDLINE, Cochrane, and Science Direct limited to the 5 years. A total of 131 articles were included, with 186 plants extracts evaluated. Although the majority of the natural extracts exhibited antifungal activities against C. albicans (both in vivo and in vitro), the strongest antifungal activity was obtained from Lawsonia inermis, Pelargonium graveolens, Camellia sinensis, Mentha piperita, and Citrus latifolia. The main components with proven antifungal activities were phenolic compounds such as gallic acid, thymol, and flavonoids (especially catechin), polyphenols such as tannins, terpenoids and saponins. The incorporation of nanotechnology greatly enhances the antifungal properties of these natural compounds. Further research is needed to fully characterize the composition of all herbal extracts with antifungal activity as well as the mechanisms of action of the active compounds.

Oral colonization by Candida species in orthodontic patients before, during and after treatment with fixed appliances: A prospective controlled trial.

BACKGROUND: Orthodontic treatment with fixed appliances is associated with changes in oral microbiota, including increased Candida colonization. The Candida fungus can cause oral lesions and infections such as candidiasis and angular cheilitis, and is harmful to both the patient and the orthodontist. Poor hygiene facilitates the colonization of these microorganisms. The key aim was to quantify the colonization of C. albicans in patients prior to beginning orthodontic treatment, and during the treatment process. MATERIAL AND METHODS: A total of 124 patients (43 males and 80 females) with a mean age of 19.5 years, who required treatment with metal or aesthetic (ceramic) braces, were studied. Microbiological samples were taken from the oral cavity using the swab technique throughout the treatment and cultured on a Sabouraud Dextrose Agar plate and, if positive, cultured on a CHROMagar® Candida plate. RESULTS: In contrast to other published studies, no statistically significant increase in C. albicans colonization was observed during the orthodontic treatment. The fixed appliances had no influence on the presence, absence or level of colonization by C. albicans and there were no significant differences between the different appliances studied. CONCLUSIONS: Our study showed that frequency of oral hygiene measures by study participants did not affect the rate of oral carriage of Candida in a statistically significant manner. This observation contrasted with published literature, which suggests that thorough brushing is important to prevent the build-up of Candida species. Key words:Orthodontics, fixed appliances, oral microflora, Candida albicans.

Dental black plaque: metagenomic characterization and comparative analysis with white-plaque.

Extrinsic black dental staining is an external dental discoloration of bacterial origin, considered a special form of dental plaque. Currently, there is no definitive therapeutic option for eliminating black stain. This study employed 16S rRNA metagenomics to analyze black stain and white-plaque samples from 27 adult volunteers. Study objectives were to: describe the microbial diversity of adult black stain samples; characterize their taxonomic profile; compare the microbiomes of black stain versus white-plaque from adult volunteers and propose a functional map of the black stain microbiome using PICRUSt2. The black stain microbiome was poorer in species diversity as compared to white-plaque. The five most abundant genera in black stain were Capnocytophaga, Leptotrichia, Fusobacterium, Corynebacterium and Streptococcus. Functional analysis of microbial species revealed conserved and consistent clustering of functional pathways within and between black stain and white-plaque microbiomes. We describe enrichment of heme biosynthetic pathways in black stain. Our results suggest that the dysbiosis in black stain resembles "orally healthy" communities. The increased abundance of heme biosynthetic pathways suggests that heme-dependent iron sequestration and subsequent metabolism are key for black stain formation. Further research should decipher the regulation of heme biosynthetic genes and characterize the temporal sequence leading to colonization and dysbiosis.

Raising awareness about microbial antibiotic resistance in undergraduate dental students: a research-based strategy for teaching non-laboratory elements of a microbiology curriculum.

BACKGROUND: Resistance to antimicrobial agents has become a problem in modern society. Antibiotic resistant bacteria undermine the prevention and treatment of infections. Undergraduate dental students in Europe are required to receive information in aspects of microbiology relevant for dental practice, including oral microbial pathogens and resistance mechanisms against antimicrobial compounds. The objective of this study was to implement a research-based strategy to aid the understanding of the increase in antimicrobial resistance in undergraduate dental student training. The primary outcome of this project is the efficacious delivery of the learning objectives. METHODS: Ten volunteer undergraduate student "ambassadors" were recruited to manage the project with assistance from lead academics. Student ambassadors were a source of peer learning for their colleagues. The project consisted of three phases: Pre-project preparation (in which the ambassadors received special instruction and training); Practical experience (in which the ambassadors worked with volunteer student colleagues to carry out the project); Public presentation of results (in which ambassadors presented study results at a scientific conference of their choosing). RESULTS: A total of 1164 students volunteered for the project, corresponding to an average participation rate of 76.4% students per year of the course. Following final debriefing, student participants and ambassadors were strongly positive in their evaluation of the achievement of 8 key student learning objectives. The results demonstrate that most volunteers improved their knowledge related to antimicrobial resistance mechanisms in microbiology. Additional benefits of participation in this project included an improvement in dental knowledge and ethics in biomedical research for the student volunteers, whilst the student ambassadors reported improved knowledge about critical thinking and study design, as well as a deeper understanding about microbiological analysis methods. CONCLUSIONS: To the best of our knowledge, this the first instance of the application of project-based methodologies to the teaching of a traditionally non-laboratory component of a subject taught in the dentistry curriculum. Results from both students and ambassadors highlighted the increase in dental knowledge and an increased awareness of antimicrobial resistance as the key outcomes of this project.

Modulation of salivary cytokines in response to alcohol, tobacco and caffeine consumption: a pilot study.

It has been shown that long-term stimulant consumption alters the biological and microbiological status of the oral cavity. We present a pilot study describing stimulant-specific oral immunomodulation in the oral cavity. Changes in salivary cytokine levels in response to long-term alcohol, tobacco and caffeine were identified. Volunteers were recruited from amongst the patients visiting University Dental Clinic of CEU Cardenal Herrera University (Alfara del Patriarca, Spain). Participants were grouped according to their self-reported levels of consumption of either caffeine, alcohol or tobacco (control group volunteers were non-consumers of all three). Informed consent was provided and stimulated saliva samples were obtained and assayed for interleukin-1α IL-1α), Tumor Necrosis Factor α (TNF-α) and Interferon γ IFN-γ). Long-term, high-level consumers of alcohol or tobacco exhibited elevated salivary concentrations of the three inflammatory cytokines with respect to control values. Specifically, IL-1α was found to be elevated in alcohol users whilst IFN-γ concentration higher in tobacco users versus controls. Long-term caffeine consumers displayed elevated levels of IFN-γ and TNF-α, whereas IL-1α levels were reduced with respect to control volunteers. This pilot study demonstrates that salivary cytokines can be modulated in response to quantity and duration of alcohol, caffeine or tobacco consumption.

Prevalence of Premolars with Dens Evaginatus in a Taiwanese and Spanish Population and Related Complications of the Fracture of its Tubercle.

OBJECTIVE: Dens evaginatus (DE) is an odontogenic developmental anomaly that can be defined as a supernumerary tubercle structure that extends from the occlusal surface of the affected tooth. Tubercular fracture or attrition of the tubercle, invaded by pulp tissue, may cause various pulpal diseases, such as pulpitis, pulp necrosis, and periapical periodontitis. This study aimed to determine the prevalence of premolars with DE in Taiwanese and Spanish study populations and to report associated dental complications. METHODS: This study was based on the clinical examination of volunteers of Spanish and Taiwanese origin, recruited among the undergraduate dentistry students at CEU-Cardenal Herrera University, Valencia, Spain. Informed consents were obtained from students identified with DE. Additional examinations were performed, including vitality test, percussion, palpation, and radiographs, to diagnosis the status of the pulp and periapical tissue. RESULTS: The prevalence of DE among Taiwanese students was 4.08%, compared to 0% in the Spanish Caucasian group. In the Taiwanese group, the occurrence of DE in premolars was more common in the mandibular (78.9%) compared to the maxillar region (21.1%). In 84% of the cases, DE-affected teeth were found bilaterally. The mandibular second premolar was the most commonly involved tooth (57.8%). When investigating the complications related to DE among six affected Taiwanese students, it was found that two teeth (10.5%) had received pulp treatment, one of them prior to complete root formation. CONCLUSION: DE appears to be more prevalent in people of Mongoloid origin and rare in those of Caucasian origin. The wear or fracture of DE may affect pulpal tissue, leading to incomplete root growth.

Integration of antimicrobial pectin-based edible coating and active modified atmosphere packaging to preserve the quality and microbial safety of fresh-cut persimmon (Diospyros kaki Thunb. cv. Rojo Brillante).

BACKGROUND: The greatest hurdle to the commercial marketing of fresh-cut fruits is related to their higher susceptibility to enzymatic browning, tissue softening, and microbial growth. The aim of this study was to test the efficacy of a pectin-based edible coating and low oxygen modified atmosphere packaging (MAP) to control enzymatic browning and reduce microbial growth of fresh-cut 'Rojo Brillante' persimmon. The survival of Escherichia coli, Salmonella enteritidis and Listeria monocytogenes artificially inoculated on fresh-cut fruit was also assessed. The pectin coating was amended with 500 IU mL-1 nisin (NI) as antimicrobial agent and 10 g kg-1 citric acid and 10 g kg-1 calcium chloride as anti-browning and firming agents, respectively. Persimmon slices were dipped in the coating or in water (control) and packed under 5 kPa O2 (MAP) or in ambient atmosphere for up to 9 days at 5 °C. Microbial growth, package gas composition, colour, firmness, polyphenol oxidase activity, visual quality and overall sensory flavour of persimmon slices were measured during storage. RESULTS: Coating application combined with active MAP significantly reduced the CO2 emission and O2 consumption in the package. The coating was effective in reducing browning and also inhibited the growth of mesophilic aerobic bacteria. Coating also reduced the populations of E. coli, S. enteritidis and L. monocytogenes. CONCLUSION: The combination of the pectin-based edible coating and active MAP proved to be the most effective treatment to maintain the sensory and microbiological quality of persimmon slices for more than 9 days of storage. © 2016 Society of Chemical Industry.

Platelet-Rich Plasma: A Study of the Variables that May Influence Its Effect on Bone Regeneration.

BACKGROUND: Currently, the use of platelet-rich plasma in bone regeneration is a real option, although more than one opinion has alerted us to the absence of clinical benefits. PURPOSE: Analysis of the factors able to modify the characteristics of the platelet preparation obtained by Curasan, Plasma Rich in Growth Factors (PRGF), Platelet Concentrate Collection System (PCCS) and SmartPrep systems, relating them to the type of clinical application and the final bone regeneration achieved. MATERIALS AND METHODS: A search was conducted in PubMed using the keywords "platelet-rich plasma," "PRP," "platelet rich growth factors," and "oral bone regeneration." Four widely accepted protocols for the obtention of PRP (above) were analyzed. Any clinical studies with controls, using the four preparation protocols and with a 4 to 6 weeks follow-up period were compared. The protocols were also grouped according to the type of PRP application: PRP-alone, with bone, or with bone substitutes. RESULTS: Bone regeneration was not achieved in any of the cases using PRP obtained by Curasan and PCCS systems, whereas PRP obtained by SmartPrep achieved it only in one in three published cases and PRGF in one in six. CONCLUSION: Based on the poor results observed in current literature, the use of PRP in oral surgery cannot be recommended.

Is revascularization of immature permanent teeth an effective and reproducible technique?

BACKGROUND: Revascularization has been proposed as an improved alternative treatment for irreversibly damaged pulp of immature teeth as it has been shown to preserve the potential for continued root growth in treated teeth. AIM: To review clinical cases of revascularization in humans to evaluate their utility and reproducibility. MATERIAL AND METHODS: A structured electronic search of scientific articles published between 2001 and 2014 was carried out using the following keywords: 'pulp revascularization', 'pulp revitalization' and/or 'immature tooth'. Clinical revascularization cases conducted on human subjects were selected, reviewed and organized into two charts including patient information, diagnostic information, treatment and results in follow-up visits. RESULTS: Ninety-seven of 101 teeth (96.0%) were successfully treated with the revascularization technique. The range of technique variations available for irrigation, disinfection and blood clot induction have a negligible impact on the clinical outcome variables tested in our analysis. During the follow-up visits, apical closure was detected in fewer cases (55.4%) as compared to the other apexogenesis phenomena (increased root length, 76.2%; increased root width, 79.2%). CONCLUSIONS: The review shows that the revascularization technique showed marked increase in the root length, width and apical closure in the cases that were reported independently of clinical variables such as operator and material selection and individual differences in protocols.

Reporters for the analysis of N-glycosylation in Candida albicans.

A large proportion of Candida albicans cell surface proteins are decorated post-translationally by glycosylation. Indeed N-glycosylation is critical for cell wall biogenesis in this major fungal pathogen and for its interactions with host cells. A detailed understanding of N-glycosylation will yield deeper insights into host-pathogen interactions. However, the analysis of N-glycosylation is extremely challenging because of the complexity and heterogeneity of these structures. Therefore, in an attempt to reduce this complexity and facilitate the analysis of N-glycosylation, we have developed new synthetic C. albicans reporters that carry a single N-linked glycosylation site derived from Saccharomyces cerevisiae Suc2. These glycosylation reporters, which carry C.albicans Hex1 or Sap2 signal sequences plus carboxy-terminal FLAG3 and His6 tags, were expressed in C.albicans from the ACT1 promoter. The reporter proteins were successfully secreted and hyperglycosylated by C.albicans cells, and their outer chain glycosylation was dependent on Och1 and Pmr1, which are required for N-mannan synthesis, but not on Mnt1 and Mnt2 which are only required for O-mannosylation. These reporters are useful tools for the experimental dissection of N-glycosylation and other related processes in C.albicans, such as secretion.

Bisphosphonates-related osteonecrosis of the jaws: a preliminary study of salivary interleukins.

OBJECTIVE: The aim of this preliminary study was analyze the possible alterations in some salivary interleukins, usually associated with the inflammatory processes. MATERIAL AND METHODS: The study comprised three groups: group 1, with 26 cases with bisphosphonates-related osteonecrosis of the jaws (BRONJ). Group 2, with 29 patients who had received iBF but without BRONJ. Group 3, with 26 control patients not treated with BF and without oral lesions. We collected unstimulated whole saliva in all groups. A semiquantitative study was performed based on a cytokine array panel. We used the proteome profiler array for the study. We analyzed: Interleukin 1 alpha (IL-1α), interleukin-1 receptor antagonist (IL-1RA), and interleukin 1 beta (IL-1ß). RESULTS: We found higher salivary values for all the cytokines studied in group 1 than in group 2 and 3. IL-1ß showed the major differences compared with control group. (P < 0.05) CONCLUSIONS: This preliminary study confirms that there are alterations in these interleukins in patients with BRONJ. These results give support to further additional salivary studies on these biomarkers by quantitative measures.

Recognition and blocking of innate immunity cells by Candida albicans chitin.

Chitin is a skeletal cell wall polysaccharide of the inner cell wall of fungal pathogens. As yet, little about its role during fungus-host immune cell interactions is known. We show here that ultrapurified chitin from Candida albicans cell walls did not stimulate cytokine production directly but blocked the recognition of C. albicans by human peripheral blood mononuclear cells (PBMCs) and murine macrophages, leading to significant reductions in cytokine production. Chitin did not affect the induction of cytokines stimulated by bacterial cells or lipopolysaccharide (LPS), indicating that blocking was not due to steric masking of specific receptors. Toll-like receptor 2 (TLR2), TLR4, and Mincle (the macrophage-inducible C-type lectin) were not required for interactions with chitin. Dectin-1 was required for immune blocking but did not bind chitin directly. Cytokine stimulation was significantly reduced upon stimulation of PBMCs with heat-killed chitin-deficient C. albicans cells but not with live cells. Therefore, chitin is normally not exposed to cells of the innate immune system but is capable of influencing immune recognition by blocking dectin-1-mediated engagement with fungal cell walls.

Glycosylation status of the C. albicans cell wall affects the efficiency of neutrophil phagocytosis and killing but not cytokine signaling.

The cell wall of the opportunistic human fungal pathogen, Candida albicans is a complex, layered network of rigid structural polysaccharides composed of ß-glucans and chitin that is covered with a fibrillar matrix of highly glycosylated mannoproteins. Polymorphonuclear cells (PMNs, neutrophils) are the most prevalent circulating phagocytic leukocyte in peripheral blood and they are pivotal in the clearance of invading fungal cells from tissues. The importance of cell-wall mannans for the recognition and uptake of C. albicans by human PMNs was therefore investigated. N- and O-glycosylation-deficient mutants were attenuated in binding and phagocytosis by PMNs and this was associated with reduced killing of C. albicans yeast cells. No differences were found in the production of the respiratory burst enzyme myeloperoxidase (MPO) and the neutrophil chemokine IL-8 in PMNs exposed to control and glycosylation-deficient C. albicans strains. Thus, the significant decrease in killing of glycan-deficient C. albicans strains by PMNs is a consequence of a marked reduction in phagocytosis rather than changes in the release of inflammatory mediators by PMNs.

Candida albicans HSP12 is co-regulated by physiological CO2 and pH.

Global transcriptional analysis of Candida albicans exposed to elevated ambient CO(2) revealed a statistically significant differential regulation of 14 genes. Subsequent RNA hybridisation analysis of one gene, HSP12, confirmed CO(2)-regulation via a cAMP-dependent mechanism. Additionally, Northern analyses and gel mobility shift assays demonstrate the co-regulation of HSP12 by environmental pH via a Rim101-dependent mechanism.

Phenotypic identification of Candida albicans by growth on chocolate agar.

In this study, we describe a simple method for the identification of Candida albicans in clinical samples. A total of 383 clinical isolates of Candida species were streaked onto chocolate agar and incubated for 48 h at 37 degrees C in the presence of an atmosphere of 6% CO2. All 208 of the C. albicans isolates tested, developed an easy to identify filamentous colony morphology. Of 175 other Candida species tested, 172 (98.3%) were distinguishable from C. albicans by their smooth colony morphology. Three isolates (1.7%) exhibited weak filamentation after prolonged incubation. Although not a routine medium in medical mycology a significant advantage of using chocolate agar lies in its use in clinical bacteriology laboratories for the isolation of fastidious bacteria. Implementation of the proposed method is applicable across a range of specimen types, thus allowing the direct identification of C. albicans in clinical samples. This simple method may allow a quicker entry into directed treatment.