Fungicidal activity of human antimicrobial peptides and their synergistic interaction with common antifungals against multidrug-resistant Candida auris.

Emergence of Candida auris, a multidrug-resistant yeast, demonstrates the urgent need for novel antifungal agents. Human antimicrobial peptides (AMPs) are naturally occurring molecules with wide spectrum antimicrobial activity, particularly against a variety of fungi. Therefore, this study examined the antifungal activity of seven different human AMPs against C. auris following the CLSI guidelines. The antifungal activity was further assessed using time kill curve and cell viability assays. For combination interaction, effectiveness of these peptides with three antifungals, fluconazole, amphotericin B, and caspofungin was done following standard protocols. To elucidate the antifungal mechanism, the effects of peptides on membrane permeability were investigated using propidium iodide staining method and confocal imaging. Antifungal susceptibility results showed that all the examined peptides possessed fungicidal effect against C. auris at different levels, with human ß-defensin-3 being the most potent antifungal with MIC values ranging from 3.125 to 12.5 µg/ml. Time kill curves further confirmed the killing effect of all the tested peptides. Viability assay showed a significant decrease in the percentage of viable cells exposed to different inhibitory and fungicidal concentrations of each peptide (p < 0.01). Furthermore, peptides showed mostly synergistic interaction when combined with conventional antifungal drugs, with caspofungin showing 100% synergy when combined with different AMPs. As antifungal mechanism, peptides disrupted the membrane permeability at concentrations that correlated with the inhibition of growth. Overall, the findings of this study point towards the application of the tested peptides as a monotherapy or as a combination therapy with antifungal drugs to treat multidrug-resistant C. auris infections.

Oral Cavity and Candida albicans: Colonisation to the Development of Infection.

Candida colonisation of the oral cavity increases in immunocompromised individuals which leads to the development of oral candidiasis. In addition, host factors such as xerostomia, smoking, oral prostheses, dental caries, diabetes and cancer treatment accelerate the disease process. Candida albicans is the primary causative agent of this infection, owing to its ability to form biofilm and hyphae and to produce hydrolytic enzymes and candialysin. Although mucosal immunity is activated, from the time hyphae-associated toxin is formed by the colonising C. albicans cells, an increased number and virulence of this pathogenic organism collectively leads to infection. Prevention of the development of infection can be achieved by addressing the host physiological factors and habits. For maintenance of oral health, conventional oral hygiene products containing antimicrobial compounds, essential oils and phytochemicals can be considered, these products can maintain the low number of Candida in the oral cavity and reduce their virulence. Vulnerable patients should be educated in order to increase compliance.

Detection of cfxA2, cfxA3, and cfxA6 genes in beta-lactamase producing oral anaerobes

ABSTRACT Purpose The aim of this study was to identify β-lactamase-producing oral anaerobic bacteria and screen them for the presence of cfxA and BlaTEM genes that are responsible for β-lactamase production and resistance to β-lactam antibiotics. Material and Methods Periodontal pocket debris samples were collected from 48 patients with chronic periodontitis and anaerobically cultured on blood agar plates with and without β-lactam antibiotics. Presumptive β-lactamase-producing isolates were evaluated for definite β-lactamase production using the nitrocefin slide method and identified using the API Rapid 32A system. Antimicrobial susceptibility was performed using disc diffusion and microbroth dilution tests as described by CLSI Methods. Isolates were screened for the presence of the β-lactamase-TEM (BlaTEM) and β-lactamase-cfxA genes using Polymerase Chain Reaction (PCR). Amplified PCR products were sequenced and the cfxA gene was characterized using Genbank databases. Results Seventy five percent of patients carried two species of β-lactamase-producing anaerobic bacteria that comprised 9.4% of the total number of cultivable bacteria. Fifty one percent of β-lactamase-producing strains mainly Prevotella, Porphyromonas, and Bacteroides carried the cfxA gene, whereas none of them carried blaTEM. Further characterization of the cfxA gene showed that 76.7% of these strains carried the cfxA2 gene, 14% carried cfxA3, and 9.3% carried cfxA6. The cfxA6 gene was present in three Prevotella spp. and in one Porphyromonas spp. Strains containing cfxA genes (56%) were resistant to the β-lactam antibiotics. Conclusion This study indicates that there is a high prevalence of the cfxA gene in β-lactamase-producing anaerobic oral bacteria, which may lead to drug resistance and treatment failure.

Influence of cancer treatment on the Candida albicans isolated from the oral cavities of cancer patients.

PURPOSE: Cancer treatment causes mucositis and the manifestation of oral candidiasis. This study investigated the virulence properties and antifungal susceptibilities of Candida albicans isolated from cancer patients undergoing therapy. METHODS: C. albicans were isolated from 49 patients on cancer treatment and 21 healthy individuals and their virulence attributes measured. A correlation was determined between the length of treatment and the fungal counts and their virulence factors. RESULTS: Although Candida carriage was similar in all the study groups, high quantities of C. albicans and variety of Candida were found in cancer patients. Germ tubes were produced by all the strains. Significantly high number of yeast isolated from radiotherapy and chemotherapy produced large quantities of phospholipase compared to healthy individuals (p < 0.01). The length of chemotherapy was associated with an increase in the phospholipase production (p = 0.03) by the C. albicans. Proteinase production was seen in a significant number of isolates from the radiotherapy group (p < 0.01). Type of cancer treatment had no effect. Resistance to antifungal agents was low. CONCLUSIONS: High quantities of phospholipase were produced by C. albicans in cancer patients on therapy which also increased with the length of chemotherapy suggesting enhanced risk of oral and systemic infection. Therefore, during treatment, prophylactic topical antifungal therapy may be considered.

Virulence of oral Candida isolated from HIV-positive women with oral candidiasis and asymptomatic carriers.

OBJECTIVE: This study compared the virulence of oral Candida species isolated from human immunodeficiency virus (HIV)-positive women with and without oral candidiasis. STUDY DESIGN: Candida species were isolated from 197 women, and their virulence attributes were measured. RESULTS: Of the 197 women, 117 (59.4%) carried Candida. Of these, 15 (12.8%) had symptoms of oral candidiasis. Among highly active antiretroviral therapy (HAART)-naive patients, 33% were diagnosed with oral candidiasis, whereas 5.9% were asymptomatic carriers (P < .01). C. albicans was the predominant species, with higher virulence attributes than non-albicans Candida. Women diagnosed with oral candidiasis had higher levels of Candida (P = .02) than asymptomatic carriers. There was no difference in the CD4 counts and the virulence attributes of Candida from both the groups. CONCLUSIONS: This study indicates that oral candidiasis is mainly caused by high counts of C. albicans and suggests the importance of therapies targeting Candida counts in the oral cavity even in patients on HAART to reduce the development of infections.

Inhibitory activity of Dodonaea viscosa var. angustifolia extract against Streptococcus mutans and its biofilm.

ETHNOPHARMACOLOGICAL RELEVANCE: The leaves of Dodonaea viscosa var. angustifolia (DVA) are traditionally used for the treatment of fever, colds, oral thrush, toothaches and related problems. Streptococcus mutans is implicated in many oral infections. This study investigated the inhibitory activity of DVA extract against Streptococcus mutans and its biofilm. MATERIALS AND METHODS: Crude extract of the leaves was prepared using methanol. The time-kill curve for Streptococcus mutans at different concentrations of methanol extract after 6 and 24 h was determined. Biofilms of Streptococcus mutans were grown in the presence of subinhibitory concentration of extract (0.78 mg/ml) for 30 h and the bacterial counts were obtained after 6, 24 and 30 h. The chemical profile of the crude extract was obtained using gas chromatography-mass spectrometry (GC-MS). RESULTS: The reduction of Streptococcus mutans was concentration and exposure time dependent. The crude extract killed 48% of S. mutans at a lowest concentration of 0.1 mg/ml and 100% at 25 mg/ml after 6h. Biofilm formation was reduced by 95, 97 and 99% after 6, 24 and 30 h of exposure to the subinhibitory concentration of crude extract respectively. GC-MS analyses revealed the presence of polyphenols such as catechin or chromene groups, chalcones with trimethoxyphenyl group and tannin with 4-O-ß-D-xylopyranoside. At high concentration the crude extract was bactericidal to Streptococcus mutans but subinhibitory concentration significantly reduced the planktonic cells and biofilm formation. CONCLUSIONS: These results suggest that this plant has the potential to be used to control S. mutans and its biofilm which are responsible for oral infections.

Isolation of Staphylococcus aureus and black-pigmented bacteroides indicate a high risk for the development of Ludwig's angina.

OBJECTIVE: The aim of this study was to compare clinical, microbiological, enzymatic, and host immune response variables between subjects hospitalized with facial cellulitis, with Ludwig's angina (LA) and without Ludwig's angina (WOLA). STUDY DESIGN: Microbiological and enzymatic tests on pus, and hematological and immunological assessments on blood samples of 15 patients with LA and 42 patients with WOLA were performed. Laboratory findings of both groups were compared using the Student t test. Multiple logistic regression analysis was performed and significant differences identified by univariate analysis. RESULTS: Patients with LA demonstrated increased levels of white blood cell counts, urea, and CRP levels, and decreased levels of CIC compared with patients WOLA. However, only CRP and urea were found to be significantly raised in the LA group. A greater population of Staphylococcus aureus and black-pigmented bacteroides were isolated from patients with LA. CONCLUSION: Elevated levels of CRP and urea could indicate the severity of infection in patients with LA. This could be because of the highly virulent and fast-spreading organisms, S. aureus and black-pigmented bacteroides, which may be a factor indicative of LA.

The effect of Dodonaea viscosa var. angustifolia on Candida albicans proteinase and phospholipase production and adherence to oral epithelial cells.

AIM OF THE STUDY: This study investigated effect of a crude extract of Dodonaea viscosa on the proteinase and phospholipase production and adherence to epithelial cells by Candida albicans isolated from HIV positive and HIV negative patients. MATERIALS AND METHODS: Twenty Candida albicans strains isolated from HIV positive and 20 from HIV negative patients were investigated. The isolates were exposed to subinhibitory concentration of crude plant extract and adherence, proteinase and phospholipase production were assessed. The results were analysed using Student's t-test and a two-way ANOVA. RESULTS: Dodonaea viscosa var. angustifolia inhibited the adherence of Candida albicans to oral epithelial cells (p=<0.01) but no significant effect of the plant extract on proteinase and phospholipase production was observed. Results from Candida albicans strains isolated from HIV positive and HIV negative patients were similar. CONCLUSIONS: Dodonaea viscosa var. angustifolia inhibited the adherence of Candida albicans to oral epithelial cells, which is the initial step of colonization in the infection process. This plant has a therapeutic potential at subinhibitory concentration.