Nondermatophyte Molds Research Articles

Study of Clinico-mycological profile of onychomycosis at a tertiary care hospital

Abstract Introduction: Onychomycosis (OM) is a common nail disorder caused by dermatophytes, non-dermatophyte moulds and yeasts. The increased prevalence of OM along with the wide range of associated fungi necessitates the accurate laboratory isolation and identification of the causative fungus for optimal management strategies. The aim of the study was to evaluate clinical, epidemiological and mycological characteristics of OM at a tertiary care hospital in Karimnagar. Materials and Methods: The study was conducted at Department of Microbiology for a period of one year. 109 patients clinically diagnosed as OM were subjected to detailed history, clinical examination and investigations. Nail clippings and subungual debris of OM patients were subjected to KOH preparation and culture. Isolates were identified by colony characters, pigment production, microscopic morphology. Results: The maximum number of OM cases belonged to the age group 21-30 years with a male preponderance. Higher incidence was noted in males (71.5%) than females (28.4%). Direct microscopy of the nail clippings in 40% KOH solution was positive in 56.8% and culture was positive in 75.2% cases. The common etiological agent was dermatophytes (45.12% cases) followed by yeasts (34.14% and non dermatophyte molds (20.7% cases). T.rubrum was the commonest among dermatophytes and Candida albicans among the yeasts. Conclusions: Although onychomycosis is not life threatening, it can cause a significant negative impact on the quality of life of infected patients. Early diagnosis and treatment is important to prevent distortion of the nails which can prevent loss of working efficiency of the commonly affected population.

Profile of onychomycosis in dermatology outpatient department at Sanglah General Hospital Denpasar, Bali-Indonesia periods 2016-2017

Introduction: Onychomycosis is a fungal infection of the nails that causes discoloration, thickening, and separation from the nail bed. Onychomycosis is caused by various organisms, most often dermatophytes of the genus Trichophyton. Other organisms include Candida, which is more common in fingernail infections and patients with chronic mucocutaneous candidiasis. Nondermatophyte molds are a less common cause in the general population. This study aimed to identify sex, age, classification, fungal culture, treatment, and incident of secondary infection in onychomycosis at Dermatology Outpatient Department at Sanglah General Hospital Denpasar Periods January 2016 – December 2017.
 Methods: Study design using descriptive retrospective model done by taking data from medical record in Dermatology Outpatient Department at Sanglah General Hospital Denpasar Periods 2016-2017.
 Result: There were 268 patients in Dermatovenereology Outpatient Department at Sanglah General Hospital Denpasar, 29 (10,82%) of whom were diagnosed with onychomycosis in Periods 2016-2017. According to age group, the most cases occur in 26-65 year age groups. Females were more frequently affected than males. The most classification finding is distal and lateral subungual onychomycosis and the most commonly isolated fungi were yeasts from the Candida species. The most commonly given therapy is a combination therapy of systemic antifungal with topical. Fluconazole is the most common drug that uses orally and ciclopirox as topical agent.
 Conclusions: Onychomycosis is one of the common fungal infection observed in patient attending dermatology outpatient department with the most classification finding is distal and lateral subungual onychomycosis. The most commonly isolated fungi were Candida sp.

Metagenomics of Toenail Onychomycosis in Three Victorian Regions of Australia.

Onychomycosis is a fungal disease of the nail that is found worldwide and is difficult to diagnose accurately. This study used metagenomics to investigate the microbiology of 18 clinically diagnosed mycotic nails and two normal nails for fungi and bacteria using the ITS2 and 16S loci. Four mycotic nails were from Bass Coast, six from Melbourne Metropolitan and eight from Shepparton, Victoria, Australia. The mycotic nails were photographed and metagenomically analysed. The ITS2 sequences for T. rubrum and T. interdigitale/mentagrophytes averaged over 90% of hits in 14/18 nails. The high abundance of sequences of a single dermatophyte, compared to all other fungi in a single nail, made it the most likely infecting agents (MLIA). Trichophyton rubrum and T. interdigitale/mentagrophytes were found in Bass Coast and Shepparton while only T. interdigitale/mentagrophytes was found in Melbourne. Two nails with T. interdigitale/mentagrophytes mixed with high abundance non-dermatophyte moulds (NDMs) (Aspergillus versicolor, Acremonium sclerotigenum) were also observed. The two control nails contained chiefly Fusarium oxysporum and Malassezia slooffiae. For bacteria, Staphylococcus epidermidis was in every nail and was the most abundant, including the control nails, with an overall mean rate of 66.01%. Rothia koreensis, Corynebacterium tuberculostearicum, and Brevibacterium sediminis also featured.

Novel and Investigational Treatments for Onychomycosis.

Onychomycosis is a common nail disease caused by fungi. The primary pathogens are dermatophytes; however, yeasts, non-dermatophyte moulds, and mixed fungal populations may also contribute to the development of a recalcitrant condition, usually accompanied by difficulties in everyday life and severe emotional stress. Treatment failure and relapse of the infection are the most frequent problems, though new issues have become the new challenges in the therapeutic approach to onychomycosis. Resistance to antifungals, an increasing number of comorbidities, and polydrug use among the ageing population are imperatives that impose a shift to safer drugs. Topical antifungals are considered less toxic and minimally interact with other drugs. The development of new topical drugs for onychomycosis is driven by the unmet need for effective agents with prolonged post-treatment disease-free time and a lack of systemic impact on the patients’ health. Efinaconazole, Tavaborole, and Luliconazole have been added to physicians’ weaponry during the last decade, though launched on the market of a limited number of countries. The pipeline is either developing new products (e.g., ME-1111 and NP213) with an appealing combination of pharmacokinetic, efficacy, and safety properties or reformulating old, well-known drugs (Terbinafine and Amphotericin B) by using new excipients as penetration enhancers.

Prevalence and Risk Factors of Superficial Fungal Infection among Patients Attending a Tertiary Care Hospital in Central Nepal.

Fungal infections of hair, nail, and skin are common worldwide and tend to increase. The present study was conducted to determine the prevalence of dermatomycoses, estimate the efficiency of rapid potassium hydroxide (KOH) wet-mount, and observe the hygienic status and the predisposing risk factors. Altogether 115 samples (nail = 77, skin = 30, and hair = 8) were obtained in a duration of December 2019 to June 2020 at Grande International Hospital, Nepal. The samples were examined by KOH wet-mount microscopy and further processed for culture. The dermatophyte test medium (DTM) was used to isolate dermatophytes separately. The fungal colonies obtained in SDA, SDA with cycloheximide/chloramphenicol and dermatophyte medium were subjected to lactophenol cotton blue (LPCB) reagent to study fungal morphology. The yeast colonies grown on SDA were subjected to Gram staining, germ-tube tests, and biochemical tests for identification. CHROMagar was used to distinguish different Candida species based on its pigment production in the medium. Various factors (age, sex, occupation, and hygiene condition) were analyzed which were associated with mycological infection. Out of 115 samples, the presence of fungal elements was detected in 20 samples by KOH. Nondermatophyte molds were the most isolated fungus in nails, skin, and hair, followed by yeast and dermatophytes, respectively. Dermatomycosis molds were the most common causative agents with 22 (14.7%) cases, followed by yeasts with 6 (5.21%) cases. Candida albicans was isolated from 5 (4.3%) cases, whereas Rhodotorula species accounted for a single (0.8%) case. Dermatophytes were isolated from 5 (4.3%) cases. Among them, n = 4(3.4%) cases revealed Trichophyton rubrum and Trichophyton mentagrophytes was isolated from single (0.8%) case. The most isolated nondermatophyte mold that follows criteria as a pathogen in our study was Cladosporium species 6 (25%) out of 27 total fungal isolates. Poor hygiene and sweating were found to be statistically significant (P < 0.05) in fungal cases detected by both KOH and culture. Dermatophytes and nondermatophyte fungi were emerging as important causes of fungal infection. Both direct microscopy and culture followed by LPCB together were vital tools for the diagnosis of fungal infections.

Identification of Dermatophyte and Non-Dermatophyte Agents in Onychomycosis by PCR and DNA Sequencing-A Retrospective Comparison of Diagnostic Tools.

Rapid and reliable fungal identification is crucial to delineate infectious diseases, and to establish appropriate treatment for onychomycosis. Compared to conventional diagnostic methods, molecular techniques are faster and feature higher accuracy in fungal identification. However, in current clinical practice, molecular mycology is not widely available, and its practical applicability is still under discussion. This study summarizes the results of 16,094 consecutive nail specimens with clinical suspicion of onychomycosis. We performed PCR/sequencing on all primary nail specimens for which conventional mycological diagnostics remained inconclusive. In specimens with a positive direct microscopy but negative or contaminated culture, molecular mycology proved superior and specified a fungal agent in 65% (587/898). In 75% (443/587), the identified pathogen was a dermatophyte. Positive cultures for dermatophytes, yeasts and non-dermatophyte molds (NDMs) were concordant with primary-specimen-DNA PCR/sequencing in 83% (10/12), 34% (22/65) and 45% (76/169), respectively. Among NDMs, agreement was high for Fusarium spp. (32/40; 80%), but low for Penicillium spp. (5/25; 20%) and Alternaria spp. (1/20; 5%). This study underlines the improvement in diagnostic yield by fungal primary-specimen-DNA PCR/sequencing in the event of a negative or contaminated culture, as well as its significance for the diagnosis of dermatophyte and non-dermatophyte onychomycosis. Molecular mycology methods like PCR and DNA sequencing should complement conventional diagnostics in cases of equivocal findings, suspected NDM onychomycosis or treatment-resistant nail pathologies.

Onychomycosis in Two Populations with Different Socioeconomic Resources in an Urban Nucleus: A Cross-Sectional Study.

Onychomycosis is one of the most common foot conditions. Mixed onychomycosis and onychomycosis caused by non-dermatophyte moulds are increasing in incidence, especially in vulnerable populations, hence the importance of this study, which presents the prevalence of onychomycosis in a population of homeless people, comparing the findings with a sample of a well-resourced population. The total sample consisted of 70 participants, divided into two separate groups, a homeless population and a second group in which we included people attending a private clinic. The average age of the sample is [49.19 ± 28.81] with an age range of 18 to 78 years. In the homeless group, the most prevalent infectious agents were non-dermatophyte fungi, with a total of 48%, compared to 28% in the group housed. The most common site of infection in both groups was the nail of the first finger. We, therefore, conclude that there is a difference in the infecting agent in the homeless population and the population with homes.

P116 Cutaneous fungal infections in elderly population from Bhopal

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM ObjectivesTo know the distribution of fungal infections based on various demographic characteristics.To study the clinical presentation of the lesions and their distribution.To isolate and identify the fungal pathogens.MethodsHospital-based cross-sectional study for 5-year duration January 2017-December 2021. Study location is the department of Microbiology AIIMS, Bhopal. Elderly patients (60 years and above) visiting as Outpatient and Inpatient of dermatology and venereology, General Medicine, and other departments with cutaneous infections suspected to be of fungal etiology and fulfilling the inclusion criteria comprised the study population. Patients already on antifungals topical or systemic were excluded. Data extraction was based on predesigned proforma for a detailed history and clinical examination entries. Necessary ethical approval and patient consent were obtained. Samples of skin, hair, nail, and exudates were processed for direct microscopy, culture isolation in suitable media, and identification phenotypically.ResultsA total of 480 elderly patients clinically suspected of cutaneous fungal infections were included in the study. Majority of the suspected cases were in 60–70 years age group 360/480 (75%). Males were 349/480 (72.71%) and females were 131/480 (27.29%) of the total suspected cases. Male to female ratio in study population was 2.66: 1. Tinea corporis 140/480 cases (29.17%) was most common clinical type followed by onychomycosis 64/480 cases (13.33%), ulcerations 62/480 (12.92%), T. cruris 56/480 (11.67%), T. pedis 35/480 (7.29%), T. mannum 25/480 (5.21%), and T. facei 12/480 (2.50%) in suspected cases of cutaneous fungal infection. Majority of cases were found in non-dependent 253/480 (52.70%) population. Out of 480 clinically suspected cases of cutaneous fungal infection was demonstrated in 193 cases (40.20%) either by direct microscopy and/or culture.A total of 176/480 cases (36.67%) were KOH positive and 113(23.54%) cases were culture positive.Taking culture as a gold standard sensitivity and specificity of KOH in diagnosing fungal infection was 84.96% and 78.2% respectively. Among 113 culture isolates dermatophytes 53.10% (60/113) were most common mold isolates followed by non-dermatophyte molds 28.3% (32/113), and yeasts 18.59% (21/113). Trichophyton mentagrophytes most common 21.24% followed by T. tonsurans 9.73%, and T. violaceum 7.96% are the common dermatophyte isolates. Aspergillus species is the most common non-dermatophyte mold isolated.Diabetes was the most common comorbid condition in culture-confirmed cases followed by hypertension and thyroid disorder.ConclusionThis study showed the prevalence of cutaneous fungal infection among elderly visiting AIIMS, Bhopal as 23.54% (113/480). With increase in elderly population, changing environmental conditions, and association with non-communicable diseases it becomes important that all elderly patients visiting hospital OPD and those hospitalized for long should be evaluated for fungal infections especially cutaneous.

P304 Epidemiological and clinical characteristics of patients with onychomycosis caused due to Neoscytalidium spp. in two reference centers in Bogotá D.C.

Poster session 2, September 22, 2022, 12:30 PM - 1:30 PMObjective: To describe the sociodemographic and clinical characteristics, diagnostic methods, and therapies received in patients with onychomycosis confirmed due to Neoscytalidium spp, who attended the San José Hospital and the Medical Mycology Laboratory for 1 year, in order to get a better perspective of the disease in our field, enhance its detection and increase early prevention.MethodsA descriptive cross-sectional observational study was carried out in two centers: San José Hospital and Medical Mycology Laboratory from November 2019 to November 2020; patients of all ages with a confirmed diagnosis of onychomycosis due to Neoscytalidium spp. were included. The cases were identified based on ICD-10 codes and the information was collected from the clinical history, questionnaires, clinical photographs, and additional documents.ResultsA total of 465 cases of onychomycosis were confirmed in the chosen period of time, of which 60 cases were caused due to Neoscytalidium spp. Affecting 31 women and 29 men, with an average age of 48.5 years in both sexes; 85% of these patients lived in Bogota D. C. The mean duration between the onset of the disease and its diagnosis was 3 years, 13% of the patients had a history of chronic disease including thyroid disorders, arterial hypertension, and diabetes mellitus. A total of 98% of the patients had toenail involvement, 88% had 1 to 5 nails involved, 33% of the cases showed mild nail involvement, 20% moderate, and 6% severe. Likewise, 35% of the cases presented dermatomycosis. 43% of the patients reported having received topic and systemic treatment, where there was no improvement in the major of the cases (78%).Conclusions: Our results support what is reported in the literature regarding risk factors, sociodemographic findings, and difficulty in therapeutic response to the pharmacological measures available to treat this infection; as well as the opinion of certain authors, onychomycosis caused due to non-dermatophyte mold is clinically indistinguishable from that caused by dermatophytes. Therefore, a complete mycological study (direct examination/KOH and culture) should always be performed to identify the causative agent before starting any treatment, since as our study showed, infections due to Neoscytalidium spp. are, in general frequency, the third most common cause of infection in onychomycosis and the first by non-dermatophyte molds.

Onychomycosis by Syncephalastrum racemosum a rare non-dermatophyte mould: Case Report from Karnataka, India

Onychomycosis is a common nail infection caused by two groups of pathogenic fungi, dermatophyte and yeast. However, in a small portion of the cases, the etiological agents are non-dermatophyte moulds, belonging to different genera and species. Among the non-dermatophytes, the yeast like Candida albicans, Candida tropicalis, and other moulds like Scopulariopsis spp., Scytalidium spp, Fusarium spp, and Aspergillus spp may be responsible. We are reporting here, a case of onychomycosis in a 45-year-old female who is non-diabetic, immunocompetent patient, due to Syncephalastrum racemosum which is a rare non dermatophyte mould and its complete improvement with oral Itraconazole and topical Nystatin combination. The affected nails were clipped after disinfecting the surface and collected in sterile container. The nails were subjected for potassium hydroxide wet mount and fungal culture was done on Sabouraud Dextrose Agar with cycloheximide. The culture slants were incubated at 37°C. The growth observed on the slants was identified using Lacto phenol cotton blue (LPCB) by morphological characteristics. Keywords: Onychomycosis, Syncephalastrum.

First report of onychomycosis caused by Phoma glomerata (Didymella glomerata): case report and literature review

Onychomycosis is the fungal infection of nails caused by yeasts, dermatophytes, and nondermatophyte molds. As a divergent set of fungi can cause onychomycosis, reliable and accurate laboratory identification of the etiological agent is crucial for choosing the proper treatment. Herein, we present a rare case of nondermatophytic onychomycosis of the toenail caused by Phoma glomerata in a 27-year-old Iranian female patient. In addition, a review of the literature on human infections caused by Phoma species is provided.

32994 Pezadeftide is a potent antifungal peptide with rapid fungicidial activity via a unique mechanism of action

Background: Fungal infections of skin, hair, and nails are extremely common and can be caused by a broad range of fungal pathogens including dermatophytes, yeasts, and nondermatophytic molds. Pezadeftide is a plant defensin peptide with potent fungicidal activity against a range of fungal pathogens that cause skin and nail infections, including Trichophyton spp., Candida spp, and a range of nondermatophytic molds. Pezadeftide is currently in development as a new topical treatment for onychomycosis. Methods: The activity of pezadeftide against a range of fungal pathogens was tested using the microbroth dilution method to determine the minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC). Fluorescent dyes were employed to assess the effect of pezadeftide on various cellular components. To identify potential targets for pezadeftide, a yeast deletion library was screened for fitness in the presence of low concentrations of pezadeftide. Results: Pezadeftide has potent antifungal activity across a broad range of fungal pathogens that cause dermatophytic infections. Pezadeftide inhibits growth at low micromolar concentrations and is fungicidal at similar concentrations, suggesting that its primary mode-of-action is fungicidal. Pezadeftide rapidly enters fungal cells and causes a rapid mitochondrial response that results in hyperpolarization of the mitochondrial membrane. Fusion of the vacuoles and production of reactive oxygen species are observed prior to disruption of the plasma membrane and cell death. Conclusions: Pezadeftide is a potent, broad-spectrum antifungal with a fungicidal mode-of-action. It rapidly kills fungal cells via interaction with the mitochondria, hyperpolarisation of the mitochondrial membrane and production of reactive oxygen species.

Prevalence of Onychomycosis in Diabetic Patients: A Case-Control Study Performed at University Hospital Policlinico in Catania.

Diabetes is characterized by an increased rate of serum glucose due to defects in insulin secretion, insulin action or both conditions. Glucose excesses can lead to extended cellular damage, with the consequence of several infectious and non-infectious skin disorders. The aim of the present study was to evaluate the toenail onychomycosis incidence in diabetic patients and healthy ones. The non-interventional, retrospective study was performed at the mycology laboratory of the University hospital “Policlinico-San Marco” in Catania, Italy, for over one year. Nail clippings were collected to perform microscopic and cultural exams, which allowed for the identification of fungal aetiological agents. A total of 715 patients (47 diabetic and 668 non-diabetic patients) were enrolled. In diabetic patients, dermatophytes were the most common cultural isolates (50%), followed by yeasts and moulds in 30.8% and 19.2%, respectively. In non-diabetic patients, the distribution of dermatophytes, yeasts and non-dermatophytic moulds was 67.4%, 5.3% and 27.3%, respectively. According to our results, diabetic patients are more predisposed to nail fungal infection. Our data suggest that dermatological follow-ups should always be performed for diabetic patients. All skin and nail disorders should be carefully monitored to perform a diagnostic confirmation and correct management of diabetic patients.

Insights on the etiopathogenesis of onychomycosis by dermatophyte, yeast and non-dermatophyte mould in ex vivo model.

Fungal biofilms have been involved in the pathogenesis of onychomycosis, but the aspects contributing to this association need to be enlightened. This study aimed to investigate the ability of three different fungi to form biofilm on the nail. All evaluated fungi were able to grow on the nails, using them as the only nutritional source and formed a structure strongly suggestive of biofilms. However, their architecture and morphology were highly contrasting: Candida albicans showed dense growth, exhibited a well-structured community and a large amount of extracellular matrix (ECM), and FTIR-ATR spectroscopy reinforced these findings revealing components suggestive of the biofilm. For Fusarium oxysporum, these events were also observed, but in lower intensity. Furthermore, while Trichophyton rubrum presented a well-organized architecture, the ECM was not visualized. We hypothesize that these findings are related to the symptomatology of onychomycosis. When it is caused by a non-dermatophyte, it usually is accompanied with paronychia, pain, oedema, inflammation and few signals of keratolysis, while dermatophytes are more associated with intense onycholysis and absence of the inflammatory signals. Biofilm seems to be crucial for non-dermatophytes to cause onychomycosis, but not for T.rubrum.

Prevalence, genetic diversity and antifungal susceptibility profiles of F.fujikuroi, F.solani and Fusarium incarnatum-equiseti species complexes from onychomycosis in north of Iran.

Onychomycosis, a nail fungal infection, is normally caused by dermatophytes. However, yeasts and non-dermatophyte moulds (NDM) are among pathogens that cause nail disease. Regarding, this study aimed to describe the molecular epidemiology of Fusarium onychomycosis in the North of Iran. Two hundred and fifty seven nail samples collected from the patients clinically suspected of onychomycosis were subjected to direct microscopy, calcofluor white staining and culture. Fusarium isolates were identified at a species level through determination of multi-locus sequences for internal transcribed spacer and translation elongation factor 1 alpha. Based on the findings, Fusarium species were isolated from onychomycosis patients (n=27). According to a previous partial genes analysis, the species in the recent study belonged to the members of F.fujikuroi species complex (n=14), Fusarium incarnatum-equiseti species complex (n=1) and F.solani species complex (n=12). In this study, F.proliferatum was the dominant Fusarium species collected from the samples. The correct identification of Fusarium species is essential regarding the increased prevalence of Fusarium onychomycosis and the inherent resistance of these agents to a wide spectrum of antifungals. The obtained results indicated variation in the epidemiology of Fusarium species isolated from onychomycosis. Moreover, the minimum inhibitory concentration (MIC) of luliconazole and lanoconazole was in the range of 0.001-1μg/ml, with the geometric mean of MICs obtained at 0.0103 and 0.0343 μg/ml against Fusarium species, respectively. These findings can increase researchers' knowledge regarding diversity of species, distribution of onychomycosis and the choice of a proper treatment.

High Trichophyton violaceum-Induced Tinea Capitis with Isolation of Many Non-Dermatophyte Molds in Scalp Scrapings in Patients Referred to a Dermatology Clinic in Addis Ababa, Ethiopia.

ObjectiveThis work aimed to determine the magnitude of tinea capitis, the diversity and species composition of fungi, and the predominant dermatophytes implicated in causing tinea capitis.MethodsA prospective, cross-sectional study was conducted at a dermatology clinic. Scalp scrapings were collected and cultured, and dermatophyte and non-dermatophyte molds were identified.ResultsOf 364 scalp scrapings, fungi were recovered from 301 (82.7%) clinical samples. About 60.7% of the samples were collected from women, while 39.3% were collected from male study subjects. The association between the magnitude of scalp ringworm and gender was not statistically significant (P = 0.105). Two hundred eighty study subjects were less than 15 years of age, of which 254 were culture positive. The association of tinea capitis and the age of patients was statistically significant (P = 0). Three hundred forty-nine fungal isolates were isolated, of which 54.2% were dermatophytes, while 45.8% were non-dermatophyte molds. The occurrence of dermatophytes in their decreasing order was T. violaceum (138; 73%), Trichophyton mentagrophytes (18; 9.5%), Trichophyton tonsurans (16; 8.5%), Trichophyton verrucosum (8; 4.5%), Microsporum audouinii (7; 3.7%), Trichophyton schoenleinii (1; 0.5%), and Trichophyton soudanense (1; 0.5%).ConclusionA high prevalence rate of T. violaceum-induced tinea capitis was reported. The magnitude of scalp ringworm in adults was remarkably high. Therefore, conducting a nationwide epidemiological survey on tinea capitis regardless of age is suggested. The isolation of many non-dermatophyte molds in the current study may shade questions about the perception that tinea capitis is caused by dermatophytes only. Therefore, studies on their potential pathogenic role on skin and skin-related (nail and the scalp) infections appear to be an active field of research.

Comparison of Fungal Fluorescent Staining and ITS rDNA PCR-based Sequencing with Conventional Methods for Diagnosis of Onychomycosis

Accurate identification of fungal causes for onychomycosis is essential for proper treatment. Presently available laboratory methods show unreliable sensitivity; so there is a requirement for innovative detection techniques. The aim for this work was to assess the efficiencies of fluorescent staining and internal transcribed spacer (ITS) ribosomal DNA (rDNA) polymerase chain reaction (PCR)-based sequencing in comparison to conventional techniques for diagnosis of onychomycosis. Nail specimens obtained from 100 patients with clinically- diagnosed onychomycosis were analyzed. Nail scrapings or clippings were subjected to direct microscopic examination by KOH mount, culture by using Sabouraud’s dextrose agar and histopathological examination with periodic-acid Schiff (PAS). Collected specimens were subsequently examined by fluorescent staining and PCR-based sequencing (30 specimens only) to compare the feasibility, sensitivity and diagnostic accuracy for these two methods. The most frequently isolated fungi were yeasts (39/76: 51.3%), dermatophytes (24/76; 31.6%) and non-dermatophyte molds (NDMs) (13/76; 17.1%). Mixed mycotic infections were recovered from 6% of the collected nail specimens. The positive detection rates were significantly different between KOH examinations (52%), nail plate histology (55%), fungal culture (70%) and fluorescent staining (80%). Considering fungal culture as the gold standard, the most sensitive technique was PCR (100%) followed by fluorescent staining (89%), PAS staining (69%) while the least sensitive technique was KOH mount (53%). Fluorescence staining can be used as a rapid and high-yield technique for identification of fungi in the specimens. PCR-based sequencing was highly sensitive and faster compared to culture. Whenever possible, it enables species identification with higher adequacy.

Competitiveness during Dual-Species Biofilm Formation of Fusarium oxysporum and Candida albicans and a Novel Treatment Strategy.

During an infection, a single or multispecies biofilm can develop. Infections caused by non-dermatophyte molds, such as Fusarium spp. and yeasts, such as Candida spp., are particularly difficult to treat due to the formation of a mixed biofilm of the two species. Fusarium oxysporum is responsible for approximately 20% of human fusariosis, while Candida albicans is responsible for superficial mucosal and dermal infections and for disseminated bloodstream infections with a mortality rate above 40%. This study aims to investigate the interactions between C. albicans and F. oxysporum dual-species biofilm, considering variable formation conditions. Further, the ability of the WMR peptide, a modified version of myxinidin, to eradicate the mixed biofilm when used alone or in combination with fluconazole (FLC) was tested, and the efficacy of the combination of WMR and FLC at low doses was assessed, as well as its effect on the expression of some biofilm-related adhesin and hyphal regulatory genes. Finally, in order to confirm our findings in vivo and explore the synergistic effect of the two drugs, we utilized the Galleria mellonella infection model. We concluded that C. albicans negatively affects F. oxysporum growth in mixed biofilms. Combinatorial treatment by WMR and FLC significantly reduced the biomass and viability of both species in mature mixed biofilms, and these effects coincided with the reduced expression of biofilm-related genes in both fungi. Our results were confirmed in vivo since the synergistic antifungal activity of WMR and FLC increased the survival of infected larvae and reduced tissue invasion. These findings highlight the importance of drug combinations as an alternative treatment for C. albicans and F. oxysporum mixed biofilms.

Studies on the Efficacy of Various Antimycotic Drugs on Emerging and Reemerging, Superficial, Cutaneous and Subcutaneous Mycotic Infections

Introduction: The efficacy of five systemic and topical antifungal medications, Voriconazole, clotrimazol, beclometasone, Itraconazole, and Fluconazole, on dermatomycosis, which affects the superficial layers of the skin, nails, foot, and hair, was tested with 180 patients.&#x0D; Methods: Included were specimen collection, processing, microscopy, and culture, as well as antifungal susceptibility testing using the E-test method. The Candida species were confirmed and their susceptibility to Voriconazole and Fluconazole was tested using the automated Vitek 2.&#x0D; Results: The final strain identification indicated 41 dermatophytes (69.49%), 11 non-dermatophytic molds (NDM) (18.64%), and 7 yeasts (11.87%). (candida). Candida was the most prevalent nondermatophyte species found. Trichophyton rubrum was the most prevalent species isolated in Tinea corporis, T. cruris, T. capitis, and T. faciei. When tested with the E strips, all dermatophyte strains showed the greatest vulnerability to beclometasone and clotrimazole (MIC range of 0.04–0.64), but homogeneous resistance to Fluconazole (i.e. MIC 32 g/ml).&#x0D; Conclusion: Variation in species distribution was shown to be statistically significant (p = 0.001) in terms of clinical presentation.

Utility of polymerase chain reaction for assessment of onychomycosis during topical therapy.

Increased detection of fungi including non-dermatophyte molds (NDMs) using polymerase chain reaction (PCR) methods is well-established. However, the use of PCR to evaluate ongoing onychomycosis treatment outcome has not been investigated. Nail samples from 28 patients receiving topical efinaconazole were evaluated by both KOH/culture and PCR methods across the study period. Detection of microorganisms by PCR was compared to the culture at baseline and end of study at month 24 (M24). Fungal detection by both methods was evaluated with respect to clinical cure observed as 100% visual clearance of the target toenail. By culture, all 28 subjects were dermatophyte-positive at pre-treatment; only 4/28 also exhibited an NDM microorganism. According to PCR, 24/28 subjects were dermatophyte-positive pre-treatment, with 25/28 also exhibiting NDMs. At M24, 18/28 (64.3%) participants had negative KOH/culture results, in contrast to 4/28 (14.3%) negative PCR results. PCR showed higher rates of NDM detection than the culture at baseline as well as M24. Calculations to compare the diagnostic utility of KOH/culture versus PCR found that positive tests with both methods reliably indicate the presence of onychomycosis, but negative PCR correlated better with onychomycosis cure than did KOH/culture. PCR confirmed a high presence of NDMs pre-treatment, and continued presence of NDMs to M24, with unknown significance requiring further investigation. Though both KOH/culture and PCR have diagnostic limitations, PCR showed better overall utility than culture in predicting onychomycosis topical treatment outcome and should be more strongly considered for evaluation of topical therapies.