Potential anti-Pythium insidiosum therapeutics identified through screening of agricultural fungicides.

Pythiosis is a life-threatening infectious disease caused by the oomycete Pythium insidiosum. Clinical manifestations of pythiosis include an eye, blood vessel, skin, or gastrointestinal tract infection. Pythiosis has been increasingly reported worldwide, with an overall mortality rate of 28%. Radical surgery is required to save patients' lives due to the limited efficacy of antimicrobial drugs. Effective medical treatments are urgently needed for pythiosis. This study aims to find anti-P. insidiosum agents by screening 17 agricultural fungicides that inhibit plant-pathogenic oomycetes and validating their efficacy and safety. Cyazofamid outperformed other fungicides as it can potently inhibit genetically diverse P. insidiosum isolates while exhibiting minimal cellular toxicities. The calculated therapeutic scores determined that the concentration of cyazofamid causing significant cellular toxicities was eight times greater than the concentration of the drug effectively inhibiting P. insidiosum. Furthermore, other studies showed that cyazofamid exhibits low-to-moderate toxicities in animals. The mechanism of cyazofamid action is likely the inhibition of cytochrome b, an essential component in ATP synthesis. Molecular docking and dynamic analyses depicted a stable binding of cyazofamid to the Qi site of the P. insidiosum's cytochrome b orthologous protein. In conclusion, our search for an effective anti-P. insidiosum drug indicated that cyazofamid is a promising candidate for treating pythiosis. With its high efficacy and low toxicity, cyazofamid is a potential chemical for treating pythiosis, reducing the need for radical surgeries, and improving recovery rates. Our findings could pave the way for the development of new and effective treatments for pythiosis.IMPORTANCEPythiosis is a severe infection caused by Pythium insidiosum. The disease is prevalent in tropical/subtropical regions. This infectious condition is challenging to treat with antifungal drugs and often requires surgical removal of the infected tissue. Pythiosis can be fatal if not treated promptly. There is a need for a new treatment that effectively inhibits P. insidiosum. This study screened 17 agricultural fungicides that target plant-pathogenic oomycetes and found that cyazofamid was the most potent in inhibiting P. insidiosum. Cyazofamid showed low toxicity to mammalian cells and high affinity to the P. insidiosum's cytochrome b, which is involved in energy production. Cyazofamid could be a promising candidate for the treatment of pythiosis, as it could reduce the need for surgery and improve the survival rate of patients. This study provides valuable insights into the biology and drug susceptibility of P. insidiosum and opens new avenues for developing effective therapies for pythiosis.

Invasive fungal infections and oomycoses in cats 2. Antifungal therapy.

CLINICAL RELEVANCE: Invasive fungal infections (IFIs) and oomycoses (hereafter termed invasive fungal-like infections [IFLIs]) are characterised by penetration of tissues by fungal elements. The environment is the most common reservoir of infection. IFIs and IFLIs can be frustrating to treat because long treatment times are usually required and, even after attaining clinical cure, there may be a risk of relapse. Owner compliance with medication administration and recheck examinations can also decline over time. In addition, some antifungal drugs are expensive, have variable interpatient pharmacokinetic properties, can only be administered parenterally and/or have common adverse effects (AEs). Despite these limitations, treatment can be very rewarding, especially when an otherwise progressive and fatal disease is cured. AIM: In the second of a two-part article series, the spectrum of activity, mechanisms of action, pharmacokinetic and pharmacodynamic properties, and AEs of antifungal drugs are reviewed, and the treatment and prognosis of specific IFIs/IFLIs - dermatophytic pseudomycetoma, cryptococcosis, sino-orbital aspergillosis, coccidioidomycosis, histoplasmosis, sporotrichosis, phaeohyphomycosis, mucormycosis and oomycosis - are discussed. Part 1 reviewed the diagnostic approach to IFIs and IFLIs. EVIDENCE BASE: Information on antifungal drugs is drawn from pharmacokinetic studies in cats. Where such studies have not been performed, data from 'preclinical' animals (non-human studies) and human studies are reviewed. The review also draws on the wider published evidence and the authors' combined expertise in feline medicine, mycology, dermatology, clinical pathology and anatomical pathology. ABBREVIATIONS FOR ANTIFUNGAL DRUGS: AMB (amphotericin B); FC (flucytosine); FCZ (fluconazole); ISA (isavuconazole); ITZ (itraconazole); KCZ (ketoconazole); PCZ (posaconazole); TRB (terbinafine); VCZ (voriconazole).

Invasive Fungal Infections and Oomycoses in Cats: 1. Diagnostic approach.

CLINICAL RELEVANCE: In contrast to superficial fungal infections, such as dermatophytosis, invasive fungal infections (IFIs) are characterised by penetration of tissues by fungal elements. Disease can spread locally within a region or can disseminate haematogenously or via the lymphatics. The environment is the most common reservoir of infection. Since fungal spores are airborne, indoor cats are also susceptible to IFIs. Some environmental fungi are ubiquitous and present globally, while others are endemic or hyperendemic within specific geographic regions. Zoonotic pathogens include Microsporum canis, Sporothrix schenckii and Sporothrix brasiliensis. AIM: In the first of a two-part article series, the approach to the investigation of feline IFIs and oomycoses is reviewed. As well as tips for diagnosis, and information on the ecological niche and distribution of fungal pathogens, the review covers clinical presentation of the most common IFIs, including cryptococcosis, histoplasmosis, blastomycosis, coccidioidomycosis, sporotrichosis, phaeohyphomycosis, aspergillosis and dermatophytic pseudomycetoma, as well as the oomycoses pythiosis, lagenidiosis and paralagenidiosis. In Part 2, the spectrum of activity, mechanisms of action, pharmacokinetic and pharmacodynamic properties and adverse effects of antifungal drugs are reviewed, and the treatment and prognosis for specific IFIs and oomycoses are discussed. EVIDENCE BASE: The review draws on published evidence and the authors' combined expertise in feline medicine, mycology, dermatology, clinical pathology and anatomical pathology.

Generation of protoplasts provides a powerful experimental research tool for biological and pathogenicity studies of Pythium insidiosum.

INTRODUCTION: Pythiosis is a high-mortality infectious condition in humans and animals. The etiologic agent is Pythium insidiosum. Patients present with an ocular, vascular, cutaneous/subcutaneous, or gastrointestinal infection. Antifungal medication often fails to fight against P. insidiosum. The effective treatment is limited to radical surgery, resulting in organ loss. Fatal outcomes are observed in advanced cases. Pythiosis needs to be studied to discover novel methods for disease control. Genome data of P. insidiosum is publicly available. However, information on P. insidiosum biology and pathogenicity is still limited due to the lack of a cost-effective animal model and molecular tools. MATERIALS AND METHODS: We aimed to develop a high-efficiency protocol for generating P. insidiosum protoplast, and used it to set up an animal model, in vitro drug susceptibility assay, and DNA transformation for this pathogen. RESULTS: P. insidiosum protoplast was successfully generated to establish a feasible pythiosis model in embryonic chicken eggs and an efficient in vitro drug susceptibility assay. DNA transformation is a critical method for gene manipulation necessary for functional genetic studies in pathogens. Attempts to establish a DNA transformation method for P. insidiosum using protoplast were partly successful. Significant work needs to be done for genetically engineering a more robust selection marker to generate stable transformants at increased efficiency. CONCLUSION: This study is the first to report an efficient P. insidiosum protoplast production for clinical and research applications. Such advances are crucial to speeding up the pathogen's biology and pathogenicity exploration.

Oomicidal activity of polypyrrole nanoparticles against Pythium insidiosum.

This study evaluated in-vitro action of a new molecule, the polypyrrole nanoparticles (Ppy-NP), against Pythium insidiosum isolates using M38-A2/CLSI; the minimal inhibitory (MIC) and minimal oomicidal (MOC) concentrations were also determined. Additionally, changes in the hyphae wall of P. insidiosum CBS 575.85 treated with Ppy-NP were evaluated by scanning electron microscopy (SEM). The MIC100 and MOC for all isolates ranged from 8 to 32 µg mL-1, and the MIC90 and MIC50 were 16 µg mL-1. The SEM showed structural damage to the hyphae of P. insidisoum treated with Ppy-NP, as hyphae surfaces with less turgidity were found, thereby showing scaling and ruptures compared to the control (untreated hyphae). Our findings highlighted the anti-P. insidiosum properties of Ppy-NP proved to be a promising candidate for research using pythiosis experimental models.

Selection of an Appropriate In Vitro Susceptibility Test for Assessing Anti-Pythium insidiosum Activity of Potassium Iodide, Triamcinolone Acetonide, Dimethyl Sulfoxide, and Ethanol.

The orphan but highly virulent pathogen Pythium insidiosum causes pythiosis in humans and animals. Surgery is a primary treatment aiming to cure but trading off losing affected organs. Antimicrobial drugs show limited efficacy in treating pythiosis. Alternative drugs effective against the pathogen are needed. In-house drug susceptibility tests (i.e., broth dilution, disc diffusion, and radial growth assays) have been established, some of which adapted the standard protocols (i.e., CLSI M38-A2 and CLSI M51) designed for fungi. Hyphal plug, hyphal suspension, and zoospore are inocula commonly used in the drug susceptibility assessment for P. insidiosum. A side-by-side comparison demonstrated that each method had advantages and limitations. Minimum inhibitory and cidal concentrations of a drug varied depending on the selected method. Material availability, user experience, and organism and drug quantities determined which susceptibility assay should be used. We employed the hyphal plug and a combination of broth dilution and radial growth methods to screen and validate the anti-P. insidiosum activities of several previously reported chemicals, including potassium iodide, triamcinolone acetonide, dimethyl sulfoxide, and ethanol, in which data on their anti-P. insidiosum efficacy are limited. We tested each chemical against 29 genetically diverse isolates of P. insidiosum. These chemicals possessed direct antimicrobial effects on the growth of the pathogen in a dose- and time-dependent manner, suggesting their potential application in pythiosis treatment. Future attempts should focus on standardizing these drug susceptibility methods, such as determining susceptibility/resistant breakpoints, so healthcare workers can confidently interpret a result and select an effective drug against P. insidiosum.

Pythiosis in a patient with Acute Myeloid Leukemia: Diagnosis and clinical course.

Pythiosis, caused by Pythium insidiosum (a fungal-like stramenipila, a group of eukaryotes away from the true fungi). Pythium insidiosum causes rare human and animal infections. Transmission from animals to human is yet to be reported. Wet soil and plants near watery environments are the source of infection. We report here a fatal case of human pythiosis in a 9-year old child with acute myeloid leukemia. Organism was identified by DNA sequencing.

A Review: Antimicrobial Therapy for Human Pythiosis.

Human pythiosis is associated with poor prognosis with significant mortality caused by Pythium insidiosum. Antimicrobials' in vitro and in vivo results against P. insidiosum are inconsistent. Although antimicrobials are clinically useful, they are not likely to achieve therapeutic success alone without surgery and immunotherapy. New therapeutic options are therefore needed. This non-exhaustive review discusses the rationale antimicrobial therapy, minimum inhibitory concentrations, and efficacy of antibacterial and antifungal agents against P. insidiosum. This review further provides insight into the immunomodulating effects of antimicrobials that can enhance the immune response to infections. Current data support using antimicrobial combination therapy for the pharmacotherapeutic management of human pythiosis. Also, the success or failure of antimicrobial treatment in human pythiosis might depend on the immunomodulatory effects of drugs. The repurposing of existing drugs is a safe strategy for anti-P. insidiosum drug discovery. To improve patient outcomes in pythiosis, we suggest further research and a deeper understanding of P. insidiosum virulence factors, host immune response, and host immune system modification by antimicrobials.

Successful potassium iodide treatment for rhinofacial pythiosis in sheep.

BACKGROUND: Pythiosis in sheep is an important disease in Brazil, which could cause rhinitis, dermatitis and alimentary tract inflammation. It is caused by the aquatic oomycete, Pythium insidiosum. The rhinofacial pythiosis causes facial deformity and upper respiratory tract clinical signs associated with necroproliferative masses occupying the rostral nasal cavity and hard palate. Little is known regarding the therapy, prophylaxis and pathogenesis of this disease. METHODOLOGY: During the 6-year study, we examined 13 sheep presenting rhinofacial pythiosis. The diagnosis was performed through biopsy of the rhinofacial lesions followed by histopathology and immunohistochemistry using specific antibodies against P insidiosum, polymerase chain reaction and an indirect enzyme-linked immunosorbent assay. RESULTS: This study presents the clinical findings of a potassium iodide treatment of rhinofacial pythiosis in sheep. All sheep were treated with 10 ml of 10% potassium iodide solution, administered orally every day during 63-120 (mean 85) days. Among treated sheep, 84.6% demonstrated complete recovery. CONCLUSION: Potassium iodide therapy may treat rhinofacial pythiosis in sheep.

Vascular pythiosis caused by Pythium aphanidermatum: the first case report in Asia.

BACKGROUND: Pythium, soil-borne plant pathogens, are in the class Oomycetes. They are not true fungi, but are related to diatom and algae. There are two human pathogens including P. insidiosum and P. aphanidermatum. To date, only one case of pythiosis caused by P. aphanidermatum has been reported. We present herein the first case of P. aphanidermatum vascular pythiosis in Asia. CASE PRESENTATION: A 47-year-old Thai woman, living in North Thailand, with ß thalassemia/hemoglobin E presented with acute recurrent arterial insufficiency of both legs. Emergent embolectomy with clot removal was performed. The pathology of the clot exhibited noncaseous granulomatous inflammation with many fungal hyphal elements. PCR identified P. aphanidermatum with 100% identity. Final diagnosis is vascular pythiosis. Unfortunately, the patient eventually expired after treatment with itraconazole, terbinafine, azithromycin, and doxycycline. CONCLUSIONS: To date, only one case of pythiosis caused by P. aphanidermatum has been reported. We present herein the first case of P. aphanidermatum vascular pythiosis in Asia.

History and Perspective of Immunotherapy for Pythiosis.

The fungus-like microorganism Pythium insidiosum causes pythiosis, a life-threatening infectious disease increasingly reported worldwide. Antimicrobial drugs are ineffective. Radical surgery is an essential treatment. Pythiosis can resume post-surgically. Immunotherapy using P. insidiosum antigens (PIA) has emerged as an alternative treatment. This review aims at providing up-to-date information of the immunotherapeutic PIA, with the focus on its history, preparation, clinical application, outcome, mechanism, and recent advances, in order to promote the proper use and future development of this treatment modality. P. insidiosum crude extract is the primary source of immunotherapeutic antigens. Based on 967 documented human and animal (mainly horses) pythiosis cases, PIA immunotherapy reduced disease morbidity and mortality. Concerning clinical outcomes, 19.4% of PIA-immunized human patients succumbed to vascular pythiosis instead of 41.0% in unimmunized cases. PIA immunotherapy may not provide an advantage in a local P. insidiosum infection of the eye. Both PIA-immunized and unimmunized horses with pythiosis showed a similar survival rate of ~70%; however, demands for surgical intervention were much lesser in the immunized cases (22.8% vs. 75.2%). The proposed PIA action involves switching the non-protective T-helper-2 to protective T-helper-1 mediated immunity. By exploring the available P. insidiosum genome data, synthetic peptides, recombinant proteins, and nucleic acids are potential sources of the immunotherapeutic antigens worth investigating. The PIA therapeutic property needs improvement for a better prognosis of pythiosis patients.

Clinical Outcomes of Radical Surgery and Antimicrobial Agents in Vascular Pythiosis: A Multicenter Prospective Study.

Vascular pythiosis is a rare, neglected, life-threatening disease with mortality of 100% in patients with incomplete surgical resection or patients with persistently elevated serum ß-d-glucan (BDG). The study was conducted to understand the clinical outcomes of new treatment protocols and potential use of erythrocyte sedimentation rate (ESR) and c-reactive protein (CRP) as alternative monitoring tools, given recent favorable minimum inhibitory concentrations (MICs) of antibacterial agents and prohibitive cost of serum BDG in Thailand. A prospective cohort study of patients with vascular pythiosis was conducted between February 2019 and August 2020. After diagnosis, patients were followed at 0.5, 1, 1.5, 3, and 6 months. Descriptive statistics, Spearman's correlation coefficient, and general linear model for longitudinal data were used. Amongst the cohort of ten vascular pythiosis patients, four had residual disease after surgery. Among four with residual disease, one developed disseminated disease and died, one developed relapse disease requiring surgery, and two were successfully managed with antimicrobial agents. The spearman's correlation coefficients between BDG and ESR, and between BDG and CRP in patients without relapse or disseminated disease were 0.65 and 0.60, respectively. Tetracyclines and macrolides had most favorable minimum inhibitory concentrations and synergistic effects were observed in combinations of these two antibiotic classes. Adjunctive use of azithromycin and doxycycline preliminarily improved survival in vascular pythiosis patients with residual disease. Further studies are needed to understand the trends of ESR and CRP in this population.

Granulomatous pneumonia due to pythiosis in captive South American coatis (Nasua nasua) / Pneumonia granulomatosa decorrente de pitiose em quatis sul-americanos (Nasua nasua) de cativeiro

Pythiosis is an emerging infectious disease affecting captive and free-ranging wild animals. We report granulomatous pneumonia due to Pythium insidiosum in two South American coatis (Nasua nasua), who were found dead without any clinical records. Severe granulomatous pneumonia associated with pleural effusion was revealed in the necropsy. Microscopically, variably sized granulomas and pyogranulomas presented negative hyphae profiles at the periphery of their necrotic cores. Grocott methenamine silver stain highlighted these structures, and immunostain (anti- P. insidiosum) was strongly positive. Molecular analysis by polymerase chain reaction amplified P. insidiosum specific DNA. These findings characterized P. insidiosum as a cause of granulomatous pneumonia in coatis and proved that pythiosis needs to be considered in the differential diagnosis of respiratory diseases affecting this species in endemic areas.(AU)

A pitiose é uma doença infecciosa emergente que afeta animais silvestres de cativeiro e em vida livre. Reportamos dois casos de pneumonia granulomatosa decorrentes da infecção por Pythium Insidiosum em quatis sul-americanos (Nasua nasua), que foram encontrados mortos sem apresentar nenhum quadro clínico prévio. Pneumonia granulomatosa severa associada a efusão pleural foi observada durante a necropsia. Na microscopia, foram observados múltiplos granulomas e piogranulomas de tamanhos variados que continham imagens negativas de hifas na periferia de seus centros necróticos. A coloração de metenamina de prata (Grocott) evidenciou estas estruturas, e a imunomarcação (anti-P. insidiosum) foi fortemente positiva. A análise molecular pela reação de polimerase em cadeia amplificou o DNA específico do P. insidiousum. Estes achados caracterizaram o P. insidiosum como a causa da pneumonia granulomatosa nos quatis e provou que a pitiose deve ser considerada um diagnostico diferencial para outras doenças respiratórias que afetam esta espécie.(AU)

Colorectal basidiobolomycosis in a dog.

A 7-year-old castrated male French Bulldog was examined for chronic large intestinal enteropathy. A colonic mass and thickened rectal mucosa were identified, and histopathologic examination of endoscopic biopsy specimens disclosed eosinophilic proctitis with large (5-20 µm), irregularly shaped, pauciseptate hyphae that were Gomori methenamine silver and periodic acid-Schiff positive. Amplification and sequencing of ribosomal DNA extracted from paraffin-embedded tissues yielded a sequence with 97% identity to GenBank sequences for Basidiobolus ranarum. After itraconazole, terbinafine, and prednisone administration, clinical signs resolved rapidly, and sonographic lesions were largely absent after 6 weeks. Treatment was discontinued by the owner 15 weeks after diagnosis. Three weeks later, the dog collapsed acutely and was euthanized. Necropsy identified metastatic islet cell carcinoma and grossly unremarkable colorectal tissues. However, histopathology of the rectum disclosed multifocal submucosal granulomas with intralesional hyphae morphologically similar to those previously observed. This report is the first to describe medical treatment of gastrointestinal basidiobolomycosis in a dog.

Pythiosis in cattle in Northeastern Brazil / Pitiose em bovinos no Nordeste do Brasil

The epidemiological, clinical and pathological aspects of cutaneous pythiosis occurring in cattle from three farms in the Northeastern of Brazil are described. A biopsy of the lesions of one bovine from each farm was performed. In two cases, the affected cattle had contact with water accumulated in dams during the dry season in the semiarid region. Another case occurred in the coastal tropical region in cattle grazing around irrigation channels. Clinically, lesions were observed mainly on the skin of the thoracic and/or pelvic limbs, characterized by flat and irregular ulcerated areas or nodules of varying sizes, some with fistulous tracts penetrating deep into the subcutaneous tissue. In one case the regional lymph nodes were affected. Histologically, in all cases, pyogranulomatous dermatitis associated with negative hyphae images, in hematoxylin-eosin stained sections, were observed. In sections stained by Grocott methenamine silver, the hyphae measured 2-8μm and had irregular ramifications and rare septations. Immunohistochemistry technique demonstrated strong immunolabeling for Pythium insidiosum. Pythiosis should be included in the differential diagnosis of dermatopathies in cattle in the Northeastern of Brazil.(AU)

Descrevem-se os aspectos epidemiológicos, clínicos e patológicos da pitiose cutânea em bovinos de três propriedades do Nordeste do Brasil. Uma biópsia das lesões de um bovino de cada propriedade foi realizada. Em dois casos, os bovinos afetados tiveram acesso à água acumulada em açudes durante a estação seca da região semiárida. O outro bovino acometido estava a pastoreio próximo a canais de irrigação na região litorânea. Clinicamente, as lesões foram observadas principalmente na pele dos membros torácicos e/ou pélvicos e caracterizavam-se por áreas planas e irregulares de ulceração ou nódulos de tamanhos variados, alguns com trajetos fistulosos penetrando profundamente no tecido subcutâneo. Em um caso, os linfonodos regionais foram afetados. Histologicamente, em todos os casos, observou-se dermatite piogranulomatosa associada a imagens negativas de hifas, em secções corados por hematoxilina e eosina. Em seções coradas por metenamina de prata de Grocott, as hifas mediam 2-8μm e possuíam ramificações irregulares com raras septações. A imuno-histoquímica demonstrou forte imunomarcação para Pythium insidiosum. A pitiose deve ser incluída como diagnóstico diferencial de dermatopatias de bovinos no Nordeste do Brasil.(AU)

Evaluating the efficacy of surgical excision and topical dimethyl sulphoxide (DMSO) in the treatment of equine cutaneous pythiosis.

BACKGROUND: Cutaneous pythiosis in horses is a chronic ulcerative granulomatous disease caused by the oomycete Pythium insidiosum. AIMS: The objective of the present study was to evaluate the response of cutaneous pythiosis in horses to surgical excision and topical dimethyl sulphoxide (DMSO). METHODS: Thirty horses were presented clinically with pruritus, fistulae discharging serosanguineous fluid, and output kunkers in different body areas (limb, abdomen, neck, and face). The clinical diagnosis was confirmed by isolation of the causative agent and histopathology. All animals were treated by surgical excision alone, or surgical excision followed by topical DMSO. The healing process was monitored every week macroscopically to evaluate the response to treatment until complete recovery. RESULTS: The existence of Pythium insidiosum was confirmed in all cases. Histologically, affected horses were characterized by granulation tissue with abundant eosinophils. The size of wounds and the clinical features of pythiosis lesions decreased more after surgical debridement with DMSO application than surgical excision alone. The cutaneous pythiosis lesions were completely recovered at 35 ± 7 and 60 ± 5 days after the surgical excision with topical DMSO and surgical excision alone, respectively. CONCLUSION: The combination of surgical excision and topical DMSO is found an effective treatment for cutaneous pythiosis in horses.

Human Pythiosis: Emergence of Fungal-Like Organism.

Pythiosis is an emerging infectious disease caused by the aquatic oomycete Pythium insidiosum, a fungal-like organism. It is believed that P. insidiosum's zoospores, its infected form, play major role in pathogenesis. Vascular and ocular infections are the most common clinical manifestation in humans. It is difficult to establish the diagnosis given its relatively rarity and difficulty to distinguish P. insidiosum from other molds. Delay in diagnosis and treatment has been associated with poor outcomes. High index of suspicion is the key, particularly in thalassemia patients with arterial insufficiency and patients with fungal keratitis/endophthalmitis without improvement on antifungal therapy. Tissue culture and zoospore induction remain gold standard for diagnosis; however, DNA-based method should be performed simultaneously. The combination of radical surgery, antifungal agents, and immunotherapy has been recommended. It was previously believed that surgery with negative surgical margins was the essential to survive in vascular pythiosis; however, it was recently found that patients could have residual disease despite documented negative surgical margins as infected clot may be dislodged to proximal arterial sites prior to surgery. Serum ß-D-glucan (BG) has been used to monitor disease response after treatment initiation in vascular pythiosis. A significant decrease in BG levels within 2 weeks after surgery is indicative of the absence of residual infection. Unfortunately, monitoring tools for ocular pythiosis are not yet available. Itraconazole plus terbinafine have generally been used in P. insidiosum-infected patients; however, antibacterial agents, including azithromycin and linezolid, have also been used with favorable outcomes in ocular disease. Recently, azithromycin or clarithromycin plus doxycyclin were used in two relapsed vascular pythiosis patients with good outcomes.

Adjunctive antibacterial agents as a salvage therapy in relapsed vascular pythiosis patients.

Human vascular pythiosis is a life-threatening condition caused by Pythium insidiosum. Patients with unresectable intra-abdominal artery involvement have not previously survived, despite being treated with antifungal agents and immunotherapy. We report two novel cases of intra-abdominal pythiosis in patients for whom surgery could not be performed, who were successfully treated with adjunctive antibacterial agents.

Equine pythiosis in the eastern wetlands of Uruguay / Pitiose equina em áreas úmidas do leste do Uruguai

Equine pythiosis is an ulcerative and granulomatous disease of the skin, caused by the oomycete Pythium insidiosum (Pythiaceae). The objective of this study was to describe seven cases of equine pythiosis that occurred from 2012 to 2017 in the eastern region of Uruguay. Six of the seven cases occurred in the eastern wetland ecosystems of the Merin basin, and the remaining case occurred in the wetland fluvial plains of the Tacuarembó River. Lesions consisted of a large, rapidly growing ulcerated tumor with abundant granulation tissue, serosanguineous secretion, and fistulous tracts containing large concretions or kunkers. The animals presented intense pruritus, claudication and loss of body condition, with death or euthanasia in extremis in six cases. The main histological lesions consisted of an eosinophilic and pyogranulomatous inflammatory process, with numerous foci of eosinophilic necrosis (kunkers), collagenolysis, and a Splendore-Hoeppli reaction. In all cases, silver coloration (Grocott) showed intralesional hyphae compatible with P. insidiosum, which was confirmed by immunohistochemistry in three cases. A horse in the terminal phase of the disease was treated with triamcinolone acetonide (50mg IM every 15 days), and fully recovered after 1 year. It is concluded that equine pythiosis is prevalent in the wetland ecosystems of eastern Uruguay and that treatment with triamcinolone is auspicious.(AU)

Pitiose é uma doença granulomatosa e ulcerativa da pele dos equinos causada pelo oomyceto Pythium insidiosum (Pythiaceae). O objetivo deste trabalho foi descrever sete casos de pitiose equina que ocorreram de 2012 a 2017 na região leste do Uruguai. Seis dos sete casos ocorreram no ecossistema de áreas pantanosas da bacia da bacia da Lagoa Mirim Merin e o restante nas planícies fluviais pantanosas do rio Tacuarembó. As lesões se caracterizaram por tumores ulcerados de crescimento rápido com abundante tecido de granulação, secreção serossanguinolenta e presença de tratos fistulosos contendo material coraloide ou kunkers. Os equinos apresentavam prurido intenso, claudicação e perda da condição corporal e seis morreram ou foram eutanasiados in extremis. As principais lesões histológicas consistiam de um processo inflamatório piogranulomatoso com numerosos focos de necrose eosinofílicos (kunkers), colagenólise e reação de Splendori-Hoepli. Em todos os casos a impregnação pela prata (Grocott) revelou a presença de hifas intralesionais compatíveis com P. insidiosum, o que foi confirmado pela imuno-histoquímica em três casos. Um equino em fase terminal da doença foi tratado com triamcinolona acetonida (50mg, IM, a cada 15 dias), recuperando-se completamente após um ano. Conclui-se que a pitiose é uma enfermidade presente em áreas úmidas na região leste do Uruguai e o tratamento com triamcinolona pode ser uma alternativa promissora.(AU)

The Repurposed Drug Disulfiram Inhibits Urease and Aldehyde Dehydrogenase and Prevents In Vitro Growth of the Oomycete Pythium insidiosum.

Pythium insidiosum is an oomycete microorganism that causes a life-threatening infectious disease, called pythiosis, in humans and animals. The disease has been increasingly reported worldwide. Conventional antifungal drugs are ineffective against P. insidiosum Treatment of pythiosis requires the extensive removal of infected tissue (i.e., eye and leg), but inadequate surgery and recurrent infection often occur. A more effective treatment is needed for pythiosis patients. Drug repurposing is a promising strategy for the identification of a U.S. Food and Drug Administration-approved drug for the control of P. insidiosum Disulfiram has been approved to treat alcoholism, but it exhibits antimicrobial activity against various pathogens. In this study, we explored whether disulfiram possesses an anti-P. insidiosum activity. A total of 27 P. insidiosum strains, isolated from various hosts and geographic areas, were susceptible to disulfiram in a dose-dependent manner. The MIC range of disulfiram against P. insidiosum (8 to 32 mg/liter) was in line with that of other pathogens. Proteogenomic analysis indicated that several potential targets of disulfiram (i.e., aldehyde dehydrogenase and urease) were present in P. insidiosum By homology modeling and molecular docking, disulfiram can bind the putative aldehyde dehydrogenase and urease of P. insidiosum at low energies (i.e., -6.1 and -4.0 Kcal/mol, respectively). Disulfiram diminished the biochemical activities of these enzymes. In conclusion, disulfiram can inhibit the growth of many pathogenic microorganisms, including P. insidiosum The drug can bind and inactivate multiple proteins of P. insidiosum, which may contribute to its broad antimicrobial property. Drug repurposing of disulfiram could be a new treatment option for pythiosis.