Deep Mycoses Research Articles

High-throughput simultaneous quantification of five azole anti-fungal agents and one active metabolite in human plasma using ultra-high-performance liquid chromatography coupled to tandem mass spectrometry

ObjectivesFor patients with hematological malignancy, triazole antifungal agents such as fluconazole (FLCZ), itraconazole (ITCZ), voriconazole (VRCZ), posaconazole (PSCZ) and isavuconazole (ISCZ) are often used for prophylaxis of deep mycosis. Since these azoles exhibit large pharmacokinetic variability, dose adjustment by therapeutic drug monitoring is recommended for some azoles. This study aimed to develop and validate a novel method for simultaneous determination of plasma concentrations of FLCZ, ITCZ, VRCZ, PSCZ, ISCZ and ITCZ-OH, an active metabolite of ITCZ, using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). Design & methodsA high-throughput solid-phase extraction method using 96-well MCX µElution Plate was selected as the pretreatment procedure. ResultsThe calibration curves for FLCZ, ITCZ, ITCZ-OH, VRCZ, PSCZ and ISCZ showed good linearity (back-calculation of calibrators: relative error ≤ 15% [LLOQ: ≤ 20%]) over wide ranges of 100–100000, 20–20000, 40–40000, 20–20000, 5–5000 and 50–50000 ng/mL, respectively. The validation results of all six drugs fulfilled the criteria of the guidance for bioanalytical method validation of the US Food and Drug Administration for within-batch and batch-to-batch precision and accuracy. The extraction recovery rates were good at ≥ 74.9%, and almost no matrix effects were found for all the drugs. The trough (10 h post-dose in 1 patient on PSCZ) drug concentrations in patients with hematologic malignancy who received oral FLCZ, ITCZ, VRCZ or PSCZ were quantified using the method developed. The measurements for all samples were within the ranges of the calibration curves, demonstrating the feasibility of clinical application of the novel method. ConclusionsWe have succeeded in developing a novel high-throughput method using UHPLC-MS/MS for simultaneous quantification of plasma concentrations of FLCZ, ITCZ, ITCZ-OH, VRCZ, PSCZ and ISCZ.

Paracoccidioidomycosis in an Elderly Simulating Cholangiocarcinoma

Paracoccidioidomycosis, the most prevalent deep mycosis in Latin America, is caused by dimorphic species of the genus Paracoccidioides, affects predominantly male rural workers between 30-50 years of age and is an important cause of morbidity and mortality. It can present in two clinical forms: acute/subacute or chronic. In rare cases paracoccidioidomycosis may simulate neoplasia and must be considered as a differential diagnosis of intra-abdominal tumors in endemic areas. We report an exuberant case of subacute paracoccidioidomycosis in an elderly lady, although it is more frequent in the juvenile form. Multiple facial skin lesions were associated with a wasting syndrome and extensive adenopathy including in the hepatic area, causing bile duct dilatation simulating a cholangiocarcinoma. Skin histopathology was diagnostic and antifungal treatment allowed complete resolution of cutaneous and systemic lesions, thus highlighting the merit of the dermatologist, who was fundamental for the conclusive diagnosis and institution of specific therapy, resulting in a favorable outcome.

Neutrophil-suppressive activity over T-cell proliferation and fungal clearance in a murine model of Fonsecaea pedrosoi infection

Neutrophils are essential to control several fungal infections. These cells are commonly known for their pro-inflammatory activities. However, some studies have demonstrated the anti-inflammatory properties of neutrophils during certain infectious diseases, culminating in the inhibition of T cell proliferation. Chromoblastomycosis (CBM) is a deep and progressive mycosis that affects thousands of people worldwide. Although neutrophil infiltrates are observed in the lesion histopathology, the fungus can overtake the immune system response and destroy the host-infected tissue. The present study demonstrated that neutropenic animals had an increase in the IL-6 production in the spleen and liver, followed by a lower fungal burden in these organs up to 14 days of infection. Neutropenic animals also showed a lower F. pedrosoi-specific antibody production 14-days post infection and higher T-cell proliferation in the in vitro experiments after stimulation with F. pedrosoi-purified proteins. Taken together, our results suggest that the presence of regulatory neutrophils in the mouse model of F. pedrosoi infection could act favoring the spread of the fungus and the chronicity of the infection. These findings shed light on the CBM treatment, which might target neutrophil polarization as a new therapy approach to treat CBM lesions.

RHINO-ORBITAL-CEREBRAL MUCORMYCOSIS(ROCM) – AS A POST COVID COMPLICATION: CASE REPORT, SYSTEMIC REVIEW AND ANALYSIS.

The most alarming health complication of modern times include Coronavirus Disease 2019 (COVID - 19) Assosciated Mucormycosis [CAM]. Mucormycosis is a deep fungal (mostly opportunistic and secondary) infection that occurs due to extremely poor hygiene and markedly reduced immunity(due to pharmacotherapy or underlying pathology)

Subcutaneous Mycosis in Hospitals: Study of 3 Cases at The National University Hospital Centre Hubert Koutoukou Maga in Cotonou (Benin)

Introduction: Deep subcutaneous mycosis results from the involvement of subcutaneous tissue by microscopic fungi. Few studies have been conducted on these diseases in Benin. Patients and methods: This was a retrospective descriptive study that included all cases of subcutaneous mycosis recorded in the Dermatology-Venerology Department of the CNHU-HKM from 2009 to 2020. Results: 3 cases of deep subcutaneous mycosis were recorded in 12 years, including two cases of basidiobolomycosis and one case of fungal mycetoma. The cases of basidiobolomycosis were observed in two girls aged 7 and 8 years whose lesions were located on the right buttock, in the form of indurated placards, adherent to the superficial plane, measuring respectively 10cm by 5cm and 14cm by 7cm. Mycological examination of the biopsy specimens isolated Basidiobolus ranarum. The only case of fungal mycetoma was observed in a 39-year-old man, in the form of a nodular placard measuring 14cm by 12cm, polyfistulised, inflammatory and squamous-crusty with an irregular border that had been evolving for 15 years. Mycological examination of the pus revealed the presence of black grains and the identification of Madurella mycetomatis. Treatment with Ketoconazole was initiated in all three patients but the evolution could not be appreciated. Conclusion : Subcutaneous mycosis is rare in Cotonou, but probably underestimated. Keywords: Subcutaneous Mycosis; Mycetoma; Basidiobolomycosis; Benin

Aspergillus Section Fumigati Pneumonia and Oxalate Nephrosis in a Foal

Equine pulmonary aspergillosis is a rare deep mycosis often due to the hematogenous spread of hyphae after gastrointestinal tract disease. We describe herein the main clinic-pathological findings observed in a foal, which spontaneously died after showing diarrhea and respiratory distress. Necropsy and histopathological investigations allowed to diagnose pulmonary aspergillosis, which likely developed after necrotic typhlitis-colitis. Biomolecular studies identified Aspergillus section Fumigati strain as the causative agent. Notably, severe oxalate nephrosis was concurrently observed. Occasionally, oxalate nephropathy can be a sequela of pulmonary aspergillosis in humans. The present case report suggests that the renal precipitation of oxalates can occur also in horses affected by pulmonary aspergillosis and could likely contribute to the fatal outcome of the disease.

Presternal Scrofuloderma: Tip of the Iceberg Unveiling Multifocal Extrapulmonary Tuberculosis - A Rare Case Report

Introduction: Scrofuloderma, a type of secondary cutaneous tuberculosis (TB) commonly arising from the underlying cervical or axillary lymph node foci via contiguity, is frequent in the pediatric population. The origin of scrofuloderma from the underlying sternal tubercular osteomyelitic focus is exceptionally unusual. It can rarely be a part of the convoluted riddle of multifocal TB, particularly in the setting of human immunodeficiency virus (HIV) infection. Case Presentation: We hereby report a 16-year-old immunocompetent female who presented with chronic, sinus-like lesions with bridging scars on the upper chest, clinically confounded with infected keloid and deep fungal infection. She was later diagnosed with scrofuloderma arising from sternal osteomyelitis compounded with multifocal tuberculosis (sternal, mediastinal, cutaneous, lumbar vertebral) and received antitubercular treatment. Conclusions: This case highlights the importance of harboring clinical scepticism supported by thorough systemic investigations while diagnosing these innocuous mimickers occurring at rare sites, even in immunocompetent individuals.

In vivo Function of Differential Subsets of Cutaneous Dendritic Cells to Induce Th17 Immunity in Intradermal Candida albicans Infection.

The skin is the outermost barrier organ in the body, which contains several types of dendritic cells (DCs), a group of professional antigen-presenting cells. When the skin encounters invading pathogens, different cutaneous DCs initiate a distinct T cell immune response to protect the body. Among the invading pathogens, fungal infection specifically drives a protective interleukin-17-producing Th17 immune response. A protocol was developed to efficiently differentiate Th17 cells by intradermal Candida albicans infection to investigate a subset of cutaneous DCs responsible for inducing Th17 immunity. Flow cytometry and gene expression analyses revealed a prominent induction of Th17 immune response in skin-draining lymph nodes and infected skin. Using diphtheria toxin-induced DC subset-depleting mouse strains, CD301b+ dermal DCs were found to be responsible for mounting optimal Th17 differentiation in this model. Thus, this protocol provides a valuable method to study in vivo function of differential subsets of cutaneous DCs to determine Th17 immunity against deep skin fungal infection.

Nitric Oxide-Releasing Nanoparticles Are Similar to Efinaconazole in Their Capacity to Eradicate Trichophyton rubrum Biofilms.

Filamentous fungi such as Trichophyton rubrum and T. mentagrophytes, the main causative agents of onychomycosis, have been recognized as biofilm-forming microorganisms. Nitric oxide-releasing nanoparticles (NO-np) are currently in development for the management of superficial and deep bacterial and fungal infections, with documented activity against biofilms. In this context, this work aimed to evaluate, for the first time, the in vitro anti-T. rubrum biofilm potential of NO-np using standard ATCC MYA-4438 and clinical BR1A strains and compare it to commonly used antifungal drugs including fluconazole, terbinafine and efinaconazole. The biofilms formed by the standard strain produced more biomass than those from the clinical strain. NO-np, fluconazole, terbinafine, and efinaconazole inhibited the in vitro growth of planktonic T. rubrum cells. Similarly, NO-np reduced the metabolic activities of clinical strain BR1A preformed biofilms at the highest concentration tested (SMIC50 = 40 mg/mL). Scanning electron and confocal microscopy revealed that NO-np and efinaconazole severely damaged established biofilms for both strains, resulting in collapse of hyphal cell walls and reduced the density, extracellular matrix and thickness of the biofilms. These findings suggest that biofilms should be considered when developing and testing new drugs for the treatment of dermatophytosis. Development of a biofilm phenotype by these fungi may explain the resistance of dermatophytes to some antifungals and why prolonged treatment is usually required for onychomycosis.

Long term follow-up of Maduromycosis treatment: A case review.

Maduromycosis is a rare deep fungal infection characterized by painless progressive destruction of limb caused by either fungal or filamentous bacteria. Its presentation is usually initiated by trivial penetration injury in farmers or laborers, worsen by immunocompromised status. Due to its painless course, this infection will severely destruct and deform hence leading to high morbidity of patient. We report a rare presentation of maduromycosis in 49-year-old housewife with no history of penetrating injury and no comorbid. Multi-disciplinary team was involved to establish the accurate diagnosis. The patient underwent surgical debridement and was given prolonged anti-fungal therapy. Combination of the treatments with patient's adherence lead to recovery without further recurrence and the patient was able to perform daily living activity.

JAAD Game Changers∗: Diagnosis of deep cutaneous fungal infections: Correlation between skin tissue culture and histopathology

JAAD Game Changers∗: Diagnosis of deep cutaneous fungal infections: Correlation between skin tissue culture and histopathology

Characteristic CD86 stain for organisms in cutaneous lesions of Talaromyces marneffei infection

To the Editor: Talaromyces marneffei (previously Penicillium marneffei) infection is endemic to Southeast Asia.1 The change in name reflects the recent taxonomic work in which many members of a subgenus of Penicillium were reassigned to the genus Taloromyces.2 A common sign of the T marneffei infection is the presence of cutaneous lesions, and the diagnosis is often made by skin biopsy. We studied macrophage polarization by performing immunohistochemical staining of cutaneous sections of deep mycosis.

English

Patients who undergo surgical resection of the maxillo-mandibular structures as a result of trauma, infection or malignancy, suffer from psycho-social setbacks which has a profound impact on their over-all quality of life. 1,2 These defects, especially those following maxillectomy, result in oroantral communication, facial deformation, impaired speech and difficulty in deglutition. For the rehabilitation of patients with such defects, surgical and prosthetic treatment options are available. As, not all patients can be successfully rehabilitated with reconstructive surgeries due to postoperative complications like graft rejection, the extent of the surgical defect and high psychological impact factor associated with repeated surgeries, prosthetic rehabilitation proves to be an alternative treatment option. The prosthetic rehabilitation of such patients is challenging as it requires restoration of the lost form, function and aesthetics, under constantly changing state of post-surgical intraoral tissues, with limited mouth opening. The maxillofacial prosthesis designed to close congenital or an acquired tissue opening, primarily of the hard palate, is known as an obturator. 3 The obturator has two functional components, one seals the surgical defect and the other replaces the lost dentoalveolarstructures.4-7 The design of an obturator may vary depending on the extent of the defect, remnant dentoalveolar complex, soft tissue undercuts and existent muscle physiology.8,9 Among the two designs, solid and hollow, hollow obturators are widely used. The bulb portion of the hollow obturator, which accommodates the surgical defect, can be open or closed9,10and its selection depends on the prosthodontist’s clinical decision-making skills and the ease of fabrication. In this article we have discussed the rehabilitated patients with single-piece, openhollow bulb definitive obturator. Patients undergo extensive maxillary surgical resections due to aggressive lesions like malignancies and deep fungal infections. Prosthetic rehabilitation of such patients with an obturator becomes of paramount importance as it separates the oropharynx from the nasopharynx, reduces the risk of recurrent infections, replaces lost dentoalveolar structures, permits intelligible speech, reinstates mastication and deglutition, restores facial contour and patient’s self-esteem. The bulb portion of the obturator extends into the defect and accommodates it, forming a hermetic seal. In this clinical report, we highlight the success of prosthetic rehabilitation of maxillectomy patients using single-piece, open-hollow bulb definitive obturator. The meticulous follow-up carried out reveals the success of the prosthesis and adds practice-based evidence to the maxillectomy rehabilitation outcome.

Challenges in the morphological diagnostics of infectious lung diseases among HIV-infected patients: a case-series study

HIV-infection is a long-term infectious disease, characterized by deterioration of the immune system and it is often complicated by the development of opportunistic diseases, mainly of infectious origin. HIV-associated infections become generalized in the late stages of the disease and often may combine, that makes challenges in the diagnosis of these diseases. Differential morphological diagnosis of HIV-associated lung infections is quite complex. Most frequently, there is the need to differentiate between tuberculosis, bacterial or septic pneumonia, cytomegalovirus lung lesions, and deep mycoses. The aim. To demonstrate the expediency and necessity to apply special complex morphological methods in HIV-infected patients with secondary lung lesions using the series of clinical cases as an example. Materials and methods. The case series consisted of 7 cases with discordance between the clinical-radiologic, laboratory data and histological conclusions. Of these, there were 5 cases of life-time and 2 autopsy diagnoses. All the clinical and laboratory data of these cases were retrospectively analyzed, and a number of additional special histological examinations were performed. The Ziehl–Neelsen and fluorescent methods, Gomori–Grocot and alcian blue-PAS staining as well as immunohistochemical examinations were used for the detection of mycobacterial antigens with different antibodies to M. tuberculosis. Results. The case series of initial morphological misdiagnosis or indeterminate histological diagnosis was revealed among hospitalized HIV-infected patients. Clinical data did not provide an accurate diagnosis of pulmonary pathology. According to the traditional histological examination, a tuberculous process was diagnosed in 3 cases, a chronic granulomatous process – in 2 cases, but without specifying the etiology. In two cases of autopsy, pneumocystis pneumonia was suspected in combination with other infections. Additional morphological examination with using 2–6 special methods made it possible to determine the correct diagnosis. In 2 cases of provisional diagnosis of tuberculosis, invasive cryptococcosis and “pneumonia in a drug user” were revealed. Cryptococcosis and pneumocystis pneumonia were detected in 2 cases with an unspecified process. In the autopsy cases, the diagnosis of pneumocystis pneumonia was ruled out, while viral-bacterial pneumonia and tuberculosis associated with cytomegalovirus infection were detected. Conclusions. The presented factual material demonstrates the principal importance of using a comprehensive morphological examination in cases of lung inflammatory processes in HIV-infected patients. Clinical patient data, namely HIV-status at the time of diagnostic procedures as well as radiological and microbiological findings serve as a substantial aid in the diagnosis.

Disseminated Histoplasmosis Infection in HIV-Negative Patients: Series Case and Literature Review

Histoplasmosis is a deep mycosis distributed worldwide and caused by Histoplasma capsulatum. Development countries and underdeveloped countries already considered an endemic disease, and its etiology comes from fungi located in soil contaminated with bird and bat droppings. Habits such as tobacco and alcohol consumption can be factors that can help to the development of disseminated histoplasmosis mainly in immunocompetent patients. Clinical manifestations can include fever, weakness, weight loss, hepatosplenomegaly, mucocutaneous lesions, and ulcer oral lesions. Oral lesions represents a commonly manifestation in the disseminated form of the disease, and it is more frequently observed in immunocompromised patients, mainly on HIV-positive patients. In this study, we presented five cases of disseminated histoplasmosis on HIVnegative patients diagnosed with lesions located in oral cavity and we performed a literature review about cases of disseminated histoplasmosis lesions in head and neck region on HIV-negative patients associating possible factors that may contribute to lesions formatting.

Possibility of Japanese Cedar Pollen Causing False Positives in the Deep Mycosis Test

Because Japanese cedar pollen (JCP) contains beta-1,3-d-glucan (BG), there is concern that its lingering presence in the atmosphere, especially during its scattering period, may cause false positives in the factor-G-based Limulus amebocyte lysate (LAL) assay used to test for deep mycosis (i.e., G-test). Hence, we examined whether the LAL assay would react positively with substances contained in JCP by using the G-test to measure JCP particles and extracts. BG was purified from the JCP extract on a BG-specific affinity column, and the percentage extractability was measured using three different BG-specific quantitative methods. The G-test detected 0.4 pg BG in a single JCP particle and 10 fg from a single particle in the extract. The percentage extractability of JCP-derived BG was not significantly different among the three quantitative methods. As the JCP particles should technically have been removed during serum separation, they should be less likely to be a direct false-positive factor. However, given that the LAL-assay-positive substances in the JCP extract were not distinguishable by the three BG-specific quantitative methods, we conclude that they may cause the background to rise. Therefore, in Japan false positives arising from JCP contamination should be considered when testing patients for deep mycosis.

DIVERGENT VERRUCOUS PRESENTATIONS – UNANTICIPATED HISTOPATHOLOGICAL DISCOVERY

Verrucous presentation of common dermatological disorders is a rare entity. Herein we report three such presentations. One is a verrucous variant of disseminated DLE, the second is verrucous psoriasis, and the other is prurigo nodularis, rare clinico-histopathological variants. They can be misdiagnosed for other closely mimicking conditions like Keratoacanthoma, Tuberculosis verrucosa cutis, and deep fungal infections.

Investigating the presence of neutrophil extracellular traps in septal and lobular cutaneous panniculitides.

Panniculitides are a heterogeneous group of inflammatory dermatoses involving the subcutaneous fatty tissue. Histologically, they are classified into septal and lobular panniculitis, according to the predominant localization of the inflammatory infiltrate. Neutrophils are frequently found in panniculitis, mainly at the early stages. Here, we investigated whether neutrophils contribute to various types of cutaneous panniculitis by releasing neutrophil extracellular traps (NETs). Formalin-fixed paraffin-embedded skin biopsies from 25 patients with panniculitis were included in the study. Our cohort was divided into n=10 erythema nodosum (septal panniculitis) and n=15 lobular panniculitis, including n=7 lupus panniculitis, n=1 pancreatic panniculitis, n=1 Weber-Christian disease, n=1 deep fungal infection, n=2 lipodermatosclerosis, and three cases did not have an identified etiology. The presence of neutrophils and NETs was assessed by double immunofluorescence using antibodies against elastase, a neutrophilic marker, and citrullinated histone 3, a marker of NETs. The mean percentages (±SEM) of elastase-positive neutrophils showing NETs were 44%±3% in erythema nodosum and 43%±7% in lobular panniculitis. The difference was not statistically significant and reflects the implication of NETs not only in severe scarring lobular panniculitis but also in benign non-scarring self-remitting reactive inflammation such as erythema nodosum. In tissues, NETs were located in the interlobular septa in erythema nodosum and in the inflamed fat lobules in lobular panniculitis. NETs are massively present in septal and lobular subtypes of panniculitides, suggesting their involvement in tissue damage.

Chasing the culprit in shadow - Sinonasal mucormycosis in an immunocompetent host.

Rhino-orbital mucormycosis is an uncommon, rapidly progressive, fulminant, angio-invasive deep mycosis usually affecting individuals with underlying immunosuppression, the most common being diabetes mellitus. In such, the course may be fatal with dissemination. Early detection with control of underlying predisposing conditions is critical with respect to the prognosis. Rarely, mucormycosis may develop in an immunocompetent host, when its non-specific symptomatology and inconspicuous radiological findings may obscure its diagnosis. Any delay in treatment predisposes to multi-organ morbidity and potential mortality. We present an immunocompetent adult with no underlying predisposing illness who was harbouring invasive sinonasal mucormycosis causing orbital cellulitis. A high index of suspicion is mandatory.

Paracoccidiomicose juvenil: um relato de caso

Objetivo: Descrever um caso de paracoccidioidomicose em um paciente previamente hígido, pois se trata de uma patologia ainda subnotificada. Detalhamento do caso: Paciente jovem do sexo masculino iniciou com quadro de lesões de pele e linfonodomegalia generalizada além de febre diária e perda de peso durante duas semanas. Ao exame físico foi evidenciado além das lesões de pele e aumento dos linfonodos uma hepatoesplenomegalia. Iniciado tratamento para PBmicose e paciente entrou em remissão das lesões recebendo alta para acompanhamento ambulatorial. Considerações finais: PBmicose é uma micose profunda de grande importância epidemiológica ainda nos dias de hoje. No entanto a mesma por não ser de notificação compulsória é ainda subdiagnosticada. É uma patologia que deve ser sempre lembrada em casos de linfonodomegalia generalizada e hepatoesplenomegalia. Seu diagnóstico envolve fatores clínicos, laboratoriais e biopsia. Seu tratamento envolve drogas de acesso amplo como o sulfametoxazol + trimetoprim e pode ser realizado em ambiente ambulatorial.