Comparative genomics of opportunistic Phialophora species involved in divergent disease types.

BACKGROUND: Black opportunists Phialophora verrucosa complex species can cause different disease types in competent and in immunocompromised individuals, but are remarkably overrepresented in CARD9-related infections. OBJECTIVES: To better understand the ecology and potential pathogenicity of opportunistic Phialophora species and reveal eventual genetic parameters associated with the behaviour in vivo and genetic profiles in patients with CARD9 immunodeficiency. METHODS: Genomes of 26 strains belonging to six species of the Phialophora verrucosa complex were sequenced. Using multilocus analysis, all environmental and clinical strains were identified correctly. We compared the genomes of agents from different disease types among each other including CARD9 immunodeficiency. RESULTS: We obtained genome sizes of the 26 Phialophora strains ranged between 32 and 37 MB. Some species showed considerable intraspecific genomic variation. P americana showed the highest degree of variability. P verrucosa was variable in CAZy enzymes, whereas P americana varied in PKS-related genes. Phialophora species, particularly P verrucosa, are relatively frequent in patients with CARD9-related immunodeficiency. Different mutations in the CARD9 gene seem to increase susceptibility for infection by different groups of species, that is either Candida, dermatophytes or black fungi. A number of patients with chromoblastomycosis revealed an as yet unknown CARD9 mutation. TNFα impairment was prevalent in patients with CARD9 infections, while CBM patients were invariably IFNγ. CONCLUSIONS: From genomic investigations, the known virulence factors between clinical and environmental strains did not reveal any significant difference. Phialophora complex has an equal chance to cause infection in humans, either healthy or CARD9-impaired.

Melanin: Quantification and protection against oxidative stress in chromoblastomycosis agents.

Chromoblastomycosis (CBM) is a chronic cutaneous and subcutaneous infection caused by melanized fungal species. We quantified the extractable melanin of 77 strains of CBM agents distributed within five genera. Moreover, resistance to oxidative stress was evaluated in strains exposed or not to the melanin inhibitor tricyclazole. The median percentage of melanin mass extracted from dry fungal mass varied from 0.69 (Rhinocladiella similis) to 3.81 (Phialophora americana). Inhibition of melanin synthesis decreased survival rates to hydrogen peroxide. Together, these data highlight the importance of melanin in CBM agents.

Case report: Fever- pneumonia- lymphadenectasis- osteolytic- subcutaneous nodule: Disseminated chromoblastomycosis caused by phialophora.

Chromoblastomycosis (CBM) is a chronic cutaneous and subcutaneous fungal infection caused by certain dematiaceous fungi (usually Fonsecaea, Phialophora, or Cladophialophora). Histologically, CBM is characterized by the presence of medlar bodies. However, the diagnosis is difficult because of the rarity of these pathognomonic presentations and the wide variety of presentations. Treatment of these infections is challenging as it lacks standardization. Herein, we report a case of chromoblastomycosis caused by Phialophora, in a 42-year-old immunocompetent male agriculturist from the humid and subtropical region of southern China. He had a 3-month history of pneumonia with intermittent fever, coughing, and expectoration. The infection subsequently spread to the bone and lymph nodes forming deep lesions and eventually resulting in osteolysis and lymphadenectasis. These subcutaneous nodules were observed after 9 months. Antifungal treatment was administered for 20 months leading to clinical improvement before the patient was lost to follow-up. This case is unique because such deep lesions are rare in immunocompetent individuals and because the initial onset was associated with pneumonia.

Phaeohyphomycosis caused by Phialophora americana with CARD9 mutation and 20-year literature review in China.

BACKGROUND: Phaeohyphomycosis is a chronic cutaneous, subcutaneous or systemic mycotic infection caused by various dematiaceous fungi. The diverse clinical manifestations and poor prognosis of phaeohyphomycosis necessitate studies on it to better recognise the disease and improve its management. OBJECTIVES: To investigate the epidemiology, aetiology, diagnosis, treatment and prognosis of phaeohyphomycosis in China over the past 20 years, and to study the first case of phaeohyphomycosis caused by Phialophora americana and the genetic and immunological mechanisms. PATIENTS/METHODS: Clinical and laboratory findings of the case were studied, and the patient's DNA was sequenced for CARD9, followed by immunological studies using patient's PBMCs. Cases of phaeohyphomycosis in China from 1998 to 2018 in both the Chinese and English literature were collected and analysed, including 45 articles and 46 patients. RESULTS: We confirmed the patient holding a homozygous frameshift mutation of CARD9, which led to impairment of pro-inflammatory cytokine production, and lower Th17- and Th22-associated responses upon fungus-specific stimulation. From the literature review, we revealed that the clinical presentations of phaeohyphomycosis were diverse. Diagnoses were established mainly on the basis of histopathology and fungal culture. Oral itraconazole, voriconazole, and posaconazole are the first choices for treatment, and a combination with surgical excision is also recommended. CONCLUSIONS: Our study establishes that obtaining detailed histories is vital for understanding the immune state and that patients with recurrent or chronic phaeohyphomycosis in the absence of known immunodeficiencies should be tested for CARD9 mutations. We hope our findings will aid clinicians in the diagnoses and treatment of such infections.

Phaeoacremonium Tenosynovitis of the Wrist.

A 79-year-old man presented with a painless, soft, subcutaneous mass lesion of the right volar wrist that had been slowly growing for 3 years. A cloudy, yellow serous effusion was aspirated from the punctured mass, from which Phaeoacremonium spp., an extremely rare cause of tenosynovitis, was isolated in culture. Total synovectomy was performed without the use of antifungal agents. No recurrence or complications occurred as of 6 months after surgery. Fungal infection is rare but should be considered in the differentiation of chronic tenosynovitis.

CARD9 deficiencies linked to impaired neutrophil functions against Phialophora verrucosa.

Caspase recruitment domain-containing protein 9 (CARD9) is an adaptor molecule that is critical for NF-κB activation and forms a complex with B cell lymphoma 10 and mucosa-associated lymphoid tissue lymphoma translocation gene 1 that mediates C-type lectin receptors (CLRs)-triggered intracellular signaling during antifungal immunity. However, the role of CARD9 in the host defense against Phialophora verrucosa (P. verrucosa) infection remains to be elucidated. In the present study, we investigated the functions of polymorphonuclear neutrophils (PMNs) from patients with CARD9 deficiencies against P. verrucosa. By isolating PMNs from patients and healthy blood donors and subsequently challenging the cells with P. verrucosa, we demonstrated that, compared with healthy donors, CARD9-deficient PMNs exhibited defects in P. verrucosa killing and pro-inflammatory cytokine productions, which can be rescued in the presence of serum; however, the CARD9-deficient PMNs exhibited normal reactive oxygen species generation and phagocytotic ability. In conclusion, our results indicate that CARD9 is indispensable for P. verrucosa killing by PMNs, and serum opsonization acts as a CARD9-independent way, which could be a promising immunotherapy in the future.

Identification and typing of isolates of Cyphellophora and relatives by use of amplified fragment length polymorphism and rolling circle amplification.

The species diversity and identification of black fungi belonging to Cyphellophora and Phialophora, which colonize and infect human skin and nails, were studied using amplified fragment length polymorphism (AFLP). A total of 76 Cyphellophora and Phialophora isolates were evaluated, and their delimitation was compared to earlier studies using multilocus sequencing. The results of the AFLP analysis and sequencing were in complete agreement with each other. Seven species-specific padlock probes for the most prevalent species were designed on the basis of the ribosomal DNA internal transcribed spacer region, and identification of the respective species could easily be achieved with the aid of rolling circle amplification.

Probable Phaeoacremonium parasiticum as a cause of cavitary native lung nodules after single lung transplantation.

Lung nodules after lung transplantation most often represent infection or post-transplant lymphoproliferative disorder in the allograft. Conversely, native lung nodules in single lung transplant recipients are more likely to be bronchogenic carcinoma. We present a patient who developed native lung cavitary nodules. Although malignancy was anticipated, evaluation revealed probable Phaeoacremonium parasiticum infection. Phaeoacremonium parasiticum is a dematiaceous fungus first described as a cause of soft tissue infection in a renal transplant patient. Lung nodules have not been previously described and this is the first case, to our knowledge, of P. parasiticum identified after lung transplantation.

Generalized subcutaneous phaeohyphomycosis caused by Phialophora verrucosa: report of a case and review of literature.

We report a case of subcutaneous phaeohyphomycosis due to Phialophora verrucosa in a 64-year-old Chinese farmer suffering from CD4+ lymphopenia. He presented with diffuse and infiltrated plaques involving the entire face including the eyes, neck, occiput, and extending to the dorsal regions of his torso. The patient is notable for the discrete multifocal nature of the illness in the absence of disseminated infection and rarity of P. verrucosa as a cause of subcutaneous phaeohyphomycosis.

Recalcitrant primary subcutaneous phaeohyphomycosis due to Phialophora verrucosa.

Phialophora verrucosa has rarely been reported for causing phaeohyphomycosis, which tends to occur in immunocompromised individuals. The case of primary subcutaneous phaeohyphomycosis due to P. verrucosa in an otherwise healthy Chinese female is presented. The girl presented with asymptomatic skin lesions when she was only 16 year old. Histological examinations revealed multiple dematiceous hyphael elements in the dermis and subcutaneous tissues. Fungal cultures were identified as P. verrucosa repeatedly based on the morphological features and confirmed by the internal transcribed spacer region nucleotide sequencing. The infection was so extremely recalcitrant that prolonged systemic antifungal regimens for 12 years revealed limited effect. The skin lesions slowly progressed and caused marked disfigurement despite the encouraging results of in vitro susceptibility. However, no relevant side effects have been reported in the course, and the patient gave birth to a healthy baby while under the maintenance treatment of itraconazole and terbinafine. This case is special in terms of the early onset, the rare clinical aspect of the pathogen, the discrepancy between in vitro and in vivo antifungal activities and especially the prolonged and recalcitrant course in such an otherwise healthy host.

[Generalized chromomycosis caused by Phialophora verrucosa]. / Chromomycose cutanée diffuse àPhialophora verrucosa.

BACKGROUND: Chromomycosis is a chronic fungal skin infection that generally presents in the form of verrucous or vegetative lesions on uncovered areas of skin. We report an unusual case of generalised chromomycosis due to Phialophora verrucosa. PATIENTS AND METHODS: A 42-year-old town-dwelling housewife was hospitalised for erythematous keratotic nodules on the arm showing sporotrichoid distribution, associated with a crusted ulcerative lesion on the homolateral index finger, as well as subcutaneous papulonodular lesions. In places, the lesions on the patient's back presented an umbilical and molluscoid appearance. The patient had suffered no previous injuries and had not visited any areas in which leishmaniasis is endemic. Her history included insulin-dependent diabetes. Screening for Leishman bodies was negative. Histopathological analysis of the skin biopsy revealed an epithelioid giant-cell granuloma with no caseous necrosis. The mycological study demonstrated the presence of fumagoid bodies and P. verrucosa was isolated. Treatment with terbinafine was initially given, followed by clarithromycin, but in the absence of any improvement, the patient was readmitted to hospital and is currently on itraconazole and amphotericin B. DISCUSSION: The novel features of our case comprise the clinical aspect of chromomycosis, the extent of the lesions, their unusual site on the back and upper limbs, and the isolation of a rare species, P. verrucosa (only the second observation in Morocco). It also highlights the therapeutic difficulties posed by this type of chromomycosis.

Occurrence and characteristics of group 1 introns found at three different positions within the 28S ribosomal RNA gene of the dematiaceous Phialophora verrucosa: phylogenetic and secondary structural implications.

BACKGROUND: Group 1 introns (ribozymes) are among the most ancient and have the broadest phylogenetic distribution among the known self-splicing ribozymes. Fungi are known to be rich in rDNA group 1 introns. In the present study, five sequences of the 28S ribosomal RNA gene (rDNA) regions of pathogenic dematiaceous Phialophora verrucosa were analyzed using PCR by site-specific primers and were found to have three insertions, termed intron-F, G and H, at three positions of the gene. We investigated the distribution of group 1 introns in this fungus by surveying 34 strains of P. verrucosa and seven strains of Phialophora americana as the allied species. RESULTS: Intron-F's (inserted at L798 position) were found in 88% of P. verrucosa strains, while intron-G's (inserted at L1921) at 12% and intron-H's (inserted at L2563) at 18%. There was some correlation between intron distribution and geographic location. In addition, we confirmed that the three kinds of introns are group 1 introns from results of BLAST search, alignment analysis and Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR). Prediction of secondary structures and phylogenetic analysis of intron sequences identified introns-F and G as belonging to subgroup IC1. In addition, intron-H was identified as IE. CONCLUSION: The three intron insertions and their insertion position in the 28S rDNA allowed the characterization of the clinical and environmental isolates of P. verrucosa and P. americana into five genotypes. All subgroups of introns-F and G and intron-H were characterized and observed for the first time in both species.

The global burden of chromoblastomycosis.

BACKGROUND: Chromoblastomycosis (CBM), represents one of the primary implantation mycoses caused by melanized fungi widely found in nature. It is characterized as a Neglected Tropical Disease (NTD) and mainly affects populations living in poverty with significant morbidity, including stigma and discrimination. METHODS AND FINDINGS: In order to estimate the global burden of CBM, we retrospectively reviewed the published literature from 1914 to 2020. Over the 106-year period, a total of 7,740 patients with CBM were identified on all continents except Antarctica. Most of the cases were reported from South America (2,619 cases), followed by Africa (1,875 cases), Central America and Mexico (1,628 cases), Asia (1,390 cases), Oceania (168 cases), Europe (35 cases), and USA and Canada (25 cases). We described 4,022 (81.7%) male and 896 (18.3%) female patients, with the median age of 52.5 years. The average time between the onset of the first lesion and CBM diagnosis was 9.2 years (range between 1 month to 50 years). The main sites involved were the lower limbs (56.7%), followed by the upper limbs (19.9%), head and neck (2.9%), and trunk (2.4%). Itching and pain were reported by 21.5% and 11%, respectively. Malignant transformation was described in 22 cases. A total of 3,817 fungal isolates were cultured, being 3,089 (80.9%) Fonsecaea spp., 552 (14.5%) Cladophialophora spp., and 56 Phialophora spp. (1.5%). CONCLUSIONS AND SIGNIFICANCE: This review represents our current knowledge on the burden of CBM world-wide. The global incidence remains unclear and local epidemiological studies are required to improve these data, especially in Africa, Asia, and Latin America. The recognition of CBM as NTD emphasizes the need for public health efforts to promote support for all local governments interested in developing specific policies and actions for preventing, diagnosing and assisting patients.

Endophthalmitis caused by Phialophora verrucosa and Streptococcus intermedius: a case report.

The most common pathogenic germs of mycotic endophthalmitis are Candida, Fusarium and Aspergillus fumigatus. As a dematiaceous fungus, Phialophora verrucosa (Medlar, 1915) has not been reported to cause endophthalmitis. Herein, we report a case of endophthalmitis induced by P. verrucosa and Streptococcus intermedius.

Isolation, characterization, and sensitivity to 2,4-diacetylphloroglucinol of isolates of Phialophora spp. from Washington wheat fields.

Dark pigmented fungi of the Gaeumannomyces-Phialophora complex were isolated from the roots of wheat grown in fields in eastern Washington State. These fungi were identified as Phialophora spp. on the basis of morphological and genetic characteristics. The isolates produced lobed hyphopodia on wheat coleoptiles, phialides, and hyaline phialospores. Sequence comparison of internal transcribed spacer regions indicated that the Phialophora isolates were clearly separated from other Gaeumannomyces spp. Primers AV1 and AV3 amplified 1.3-kb portions of an avenacinase-like gene in the Phialophora isolates. Phylogenetic trees of the avenacinase-like gene in the Phialophora spp. also clearly separated them from other Gaeumannomyces spp. The Phialophora isolates were moderately virulent on wheat and barley and produced confined black lesions on the roots of wild oat and two oat cultivars. Among isolates tested for their sensitivity to 2,4-diacetylphloroglucinol (2,4-DAPG), the 90% effective dose values were 11.9 to 48.2 microg ml(-1). A representative Phialophora isolate reduced the severity of take-all on wheat caused by two different isolates of Gaeumannomyces graminis var. tritici. To our knowledge, this study provides the first report of an avenacinase-like gene in Phialophora spp. and demonstrated that the fungus is significantly less sensitive to 2,4-DAPG than G. graminis var. tritici.

[Melanonychia: The purpose of a case of ungual mycotization mimicking melanoma]. / Melanoníquia: A propósito de um caso de micotização ungueal simulando melanoma.

INTRODUCTION: Phialophora sp. dematiaceous fungus frequently isolated from soil and organic debris from animals and plants. There have been no reports of nail involvement caused by this fungus. We report the case of a 77-year-old male with blackish-brown pigmentation involving a single finger for one year. The identification of the fungus was determined by macromorphological and micromorphological study of the colony and we discuss the clinical, microbiological, differential diagnosis, and treatment. COMMENT: We describe the first case of nail infection caused by Phialophora sp.

[Chronic fistula of the forefoot in a Moroccan man: bone chromomycosis due to Phialophora verrucosa]. / Fistule chronique de l'avant pied chez un marocain: chromomycose osseuse à Phialophora verrucosa.

Chromomycosis is a chronic infection caused by various fungi. Bone involvement is exceptional. The purpose of this report is to describe a case of chromoblastomycosis localized on the tarsal bone in a 27-year-old man from southern Morocco. The patient was a former barefoot long-distance runner. He presented with thickening of the skin with a cutaneous fistula on the left forefoot. These manifestations had been ongoing for 2 years. Imaging demonstrated scattered lytic lesions involving tarsal bone and the base of the third metatarsus. Definitive diagnosis was based on direct mycological examination and anatomopathology. Medical and surgical treatment led to stabilization of the lesions. No recurrence was observed during follow-up examination two years later.

Phaeohyphomycosis: Cytomorphologic Evaluation in Eleven Cases.

OBJECTIVE: Phaeohyphomycosis caused by phaeoid fungi is a type of mycosis emerging worldwide which causes a wide variety of clinical manifestations. STUDY DESIGN: A retrospective analysis of 11 cases diagnosed with fungal inflammation on cytology over a period of 6 years (2013-2018) was done along with culture/histopathologic confirmation. RESULTS: Of the total of 11 cases, 9 cases presented with subcutaneous swellings and 1 case each with brain and lung lesions. The age range was 30-83 years (mean: 53.6); 8 patients were male and 3 were female. Cytologic smears showed fungal profiles with septate tortuous hyphae, as well as swollen and narrow, yeast-like swellings with an irregular breadth of the hyphae in all cases. The fungal profiles were visualized on a Masson-Fontana stain. The background showed inflammatory cells, giant cells, and necrosis in variable proportions. Five cases were diagnosed as phaeohyphomycosis on cytology, whereas 3 cases were misdiagnosed as aspergillus and 2 as candida. In 1 case, typing of the fungus was not done. Histopathology was available in 5 cases, and in all these a diagnosis of phaeohyphomycosis was reached. Ten of the 11 cases had confirmation on fungal culture. CONCLUSIONS: Phaeoid fungi are rarely seen in routine cytologic practice. Careful evaluation of cytologic smears and an awareness of the characteristic morphologic features of phaeohyphomycosis are helpful in arriving at a correct diagnosis. Fine needle aspiration cytology provides a rapid diagnosis, enabling prompt therapy.

Immigration Brings New Pathology with No Standardized Treatment Protocol Madura Foot Case Studies Phialemonium and Phaeoacremonium.

Madura foot is an uncommon invasive soft-tissue infection that foot and ankle specialists encounter. We present two rare cases of Phialemonium and Phaeoacremonium fungi infections of the foot diagnosed in northern California to inform physicians on the presentation and current treatment options for this unique pathology. The two cases presented outline the clinical presentations, diagnostic data, and surgical and antimicrobial interventions. There is a concentration on the antimicrobial options depending on which of the over 20 species is encountered. The pertinent literature and supporting data are reviewed to create an outline for discussion of treatment protocols when faced with these emerging opportunistic infections.