Project description:PurposeTo determine the incidence and density of Demodex species on the eyelashes of subjects with normal eyelids, anterior blepharitis (AB), meibomian-gland dysfunction (MGD), and mixed blepharitis (MB).Materials and methodsOne hundred and fifty consecutive patients diagnosed with AB, MGD, and MB were recruited. An additional 50 individuals were recruited who were free of lid and margin disease to serve as a control group. All patients underwent a standard eye examination. Data on ocular symptomatology were gathered. Digital photography was performed of the lid margins. Lash sampling was performed by epilating the lashes and the lashes were checked for Demodex based on morphology using light microscopy. The total number of mites were tabulated for each eye. Comparison between the affected eyes and the control eyes was performed. Data were analyzed using the Chi-square test.ResultsA total of 200 patients were included. Twenty percenth had AB, 40% had MGD, and 40% had MB. The incidence of Demodex infestation was 90% in cases of AB, 60% in MGD cases, and 90% in MB cases. The incidence of Demodex in control subjects was 18%.ConclusionsThe incidence and density of Demodex infestation was highest in patients with AB and MB. Lid irritation and presence of cylindrical dandruff were indicative of a high-density infestation. These signs should alert the clinician to treat concomitant Demodex infestation.
Project description:Purpose: The aim of this study was to compare the ocular microbial communities in humans with and without demodex blepharitis in order to elucidate the relationship between ocular microorganisms and demodex infestation. Methods: Bacterial 16S rRNA genes of conjunctival sac samples from 30 demodex blepharitis patients and 14 healthy controls were sequenced using a pyrosequencing method, and their bacterial community structures were compared by bioinformatics. Results: Bacterial community clustering of conjunctival sac in the demodex blepharitis group were significantly distinct from the healthy control group, with significantly higher relative abundances of Firmicutes and Corynebacterium at the phyla level, as well as higher abundances of Lactobacillus and Bifidobacterium at the genus level. The relative abundance of Staphylococcus epidermidis (0.07-2.27%) was positively correlated with the demodex amount and modified OSDI. The major potential factors contribute to demodex blepharitis were Bacilli, Firmicutes, Cyanobacteria, Lactobacillus and Streptophyta. Conclusions: Patients with demodex blepharitis have varying degrees of bacterial microbiota imbalance in the conjunctival sac. Demodex serving as vectors to transfer both skin and environmental flora might be the potential mechanism. In addition, the number and type of demodex affect the specific ocular surface bacteria, presenting as ocular discomfort and obvious signs of blepharitis.
Project description:S. aureus and MRSA are susceptible to TTO and are therefore targeted in nascent TTO clinical trails. We now report the alterations in the transcriptome of a S. aureus exposed to a growth inhibitory concentration of TTO. These efforts have uncovered additional mechanisms by which this membrane active antimicrobial substance inhibits bacterial growth.
Project description:S. aureus and MRSA are susceptible to TTO and are therefore targeted in nascent TTO clinical trails. We now report the alterations in the transcriptome of a S. aureus exposed to a growth inhibitory concentration of TTO. These efforts have uncovered additional mechanisms by which this membrane active antimicrobial substance inhibits bacterial growth. Four hybridizations, including a biological replicate and a dye-swap experiment for each replicate was carried out. LOWESS normalization was performed on the raw data to correct for dye-bias. Statistical analysis was performed using a significance analysis of microarrays (SAM, MultiExperiment Viewer, ver. 4.0) unpaired contrast. A d-statistic was calculated for each gene based on repeated permutations, and a false discovery rate FDR of 0.01 was used to assign a critical cutoff for significance.
Project description:BackgroundDemodex blepharitis is a common chronic disease. The number of mites is associated with ocular discomfort. The accurate number derived from well-stained specimens is, hence, in favor of diagnosing, monitoring, and determining treatment responses.MethodsA cross-sectional study was conducted between April and July 2022 at the dermatology and ophthalmology clinic, Walailak University, Thailand. Adult participants with clinical suspicion of Demodex blepharitis were recruited. We examined eyelashes under light microscopy to quantify the number of Demodex mites before and after adding CSB gel. The mite counts, evaluated by an untrained investigator and an experienced investigator, were recorded and compared.ResultsA total of 30 participants were included for final analysis, among which 25 (83.3%) were female. The median age was 64.0 years (IQR, 61.0-68.0). The median Demodex counts evaluated by the experienced investigator before and after adding CSB gel were 1.0 (IQR, 0.0-1.0) and 2.5 (IQR, 2.0-3.0), respectively (p < 0.001). Moreover, the median Demodex counts evaluated by the untrained investigator before and after adding CSB gel were 1.0 (IQR, 0.0-1.0) and 2.0 (IQR, 1.0-3.0), respectively (p < 0.001). The correlation coefficient between Demodex counts after the addition of CSB counted by the experienced investigator and those counted by the untrained investigator was 0.92 (p < 0.001). CSB gel is a promising product to identify and quantify the number of Demodex mites. The findings supported the consideration of CSB gel as one of the diagnostic stains.
Project description:Tea tree oil (TTO) is a steam distillate of Melaleuca alternifolia that demonstrates broad-spectrum antibacterial activity. This study was designed to document how TTO challenge influences the Staphylococcus aureus transcriptome. Overall, bioinformatic analyses (S. aureus microarray meta-database) revealed that both ethanol and TTO induce related transcriptional alterations. TTO challenge led to the down-regulation of genes involved with energy-intensive transcription and translation, and altered the regulation of genes involved with heat shock (e.g. clpC, clpL, ctsR, dnaK, groES, groEL, grpE and hrcA) and cell wall metabolism (e.g. cwrA, isaA, sle1, vraSR and vraX). Inactivation of the heat shock gene dnaK or vraSR which encodes a two-component regulatory system that responds to peptidoglycan biosynthesis inhibition led to an increase in TTO susceptibility which demonstrates a protective role for these genes in the S. aureus TTO response. A gene (mmpL) encoding a putative resistance, nodulation and cell division efflux pump was also highly induced by TTO. The principal antimicrobial TTO terpene, terpinen-4-ol, altered ten genes in a transcriptional direction analogous to TTO. Collectively, this study provides additional insight into the response of a bacterial pathogen to the antimicrobial terpene mixture TTO.
Project description:Tea tree oil (TTO)-reduced susceptibility (TTORS) mutants of two Staphylococcus aureus laboratory strains were isolated utilizing TTO gradient plates. Attempts to isolate TTORS mutants employing agar plates containing single TTO concentrations failed. All TTORS mutants demonstrated a small colony variant (SCV) phenotype and produced cells with a smaller diameter, as determined by scanning electron microscopy. The addition of SCV auxotrophic supplements to media did not lead to an increase in TTORS mutant colony size. Revertants were also isolated from the TTORS mutants following growth in drug-free media, and all revertant strains demonstrated phenotypes similar to their respective parent strains. Transmission electron microscopy revealed that an SH1000 TTORS mutant demonstrated a thinner cell wall and novel septal invaginations compared with parent strain SH1000. In addition, comparative genomic sequencing did not reveal any mutations in an SH1000 TTORS mutant previously linked to well-characterized SCV genotypes. This study demonstrates that TTO can select for a unique SCV phenotype.
Project description:Tissue engineering is crucial, since its early adoption focused on designing biocompatible materials that stimulate cell adhesion and proliferation. In this sense, scaffolds made of biocompatible and resistant materials became the researchers' focus on biomedical applications. Humans have used essential oils for a long time to take advantage of their antifungal, insecticide, antibacterial, and antioxidant properties. However, the literature demonstrating the use of essential oils for stimulating biocompatibility in new scaffold designs is scarce. For that reason, this work describes the synthesis of four different film composites of chitosan/polyvinyl alcohol/tea tree (Melaleuca alternifolia), essential oil (CS/PVA/TTEO), and the subdermal implantations after 90 days in Wistar rats. According to the Young modulus, DSC, TGA, mechanical studies, and thermal studies, there was a reinforcement effect with the addition of TTEO. Morphology and energy-dispersive (EDX) analysis after the immersion in simulated body fluid (SBF) exhibited a light layer of calcium chloride and sodium chloride generated on the material's surface, which is generally related to a bioactive material. Finally, the biocompatibility of the films was comparable with porcine collagen, showing better signs of resorption as the amount of TTEO was increased. These results indicate the potential application of the films in long-term biomedical needs.
Project description:Purpose: High-throughput RNA sequencing has been used to examine mRNA expression profiles in fungal cells treated with essential oils. The goals of this study are to analyze the global gene expression profiles in Botrytis cinerea with or without tea tree oil and its two characteristic components treatment by RNA-Seq. Methods: The mRNA profiles of Botrytis cinerea with or without tea tree oil and its two characteristic components treatment were generated by deep sequencing, in triplicate, using Illumina HiSeq™ 2500 sequencing platform. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR was performed to verified the sensitivity of the RNA-seq method. Results: After high-throughput RNA sequencing, reads were filtered to yield 111.22 Gb of clean sequence data. The GC content for all samples exceeded 45%. The Q20 ratio (used to evaluate reads quality) was greater than 94%, and Q30 base percentage was at least 87.07%. Altered expression of 7 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Most differentially expressed genes (DEGs) from B. cinera cells treated with terpinen-4-ol participated in biosynthesis of secondary metabolites, and the metabolism of amino acid, carbohydrate and lipid. 1,8-cineole mainly affected DEGs involved in genetic information processing, and thus inducing cell death. Conclusions: Terpinen-4-ol exerts antifungal activity mainly by blocking the expression of genes related to cell integrity and mitochondrial function. 1,8-cineole primarily affects genes involved in genetic information processing including DNA replication, transcription and repair. This study provides insight into the molecular mechanism by which tea tree oil acts against Botrytis cinerea based on the data from RNA-seq.