Vascular Pythiosis Research Articles

Functional characterization of macrophages and change of Th1/Th2 balance in patients with pythiosis after Pythium insidiosum antigen immunotherapy

There has been limited research into the role of the Pythium insidiosum antigen (PIA) in modulating immune response in patients with pythiosis. This study investigated the balance of T helper type 2 (Th2) and T helper type 1 (Th1) responses after receiving PIA immunotherapy in patients with pythiosis. Next, the phagocytic activity and phagocytic index of IFN-γ primed PIA-treated macrophages were examined. Furthermore, the phagocytosis of infective P. insidiosum zoospores by macrophages was investigated. This work showed that the PIA vaccine induced Th1 response and M1 macrophages in patients with vascular pythiosis who survived and those with localized pythiosis. Phagocytic activity and phagocytic index were increased considerably in localized pythiosis patients compared to vascular pythiosis patients with hematological diseases. IFN-γ priming of PIA-treated macrophages against P. insidiosum zoospores enhanced the phagocytic activity and phagocytic index in vascular and localized pythiosis patients. Macrophages engulfed P. insidiosum zoospores, but the zoospores continued germination, resulting in macrophage death. Overall, our results suggest that PIA can modulate the immune responses, contributing to higher levels of Th1-type cytokine and potentially improving the survival of patients with vascular pythiosis. This study is the first to uncover that P. insidiosum zoospores can survive within macrophages.

Computed Tomography Finding of Crusty Thrombosed Arteries: An Appearance of Lower Extremity Vascular Pythiosis.

Vascular pythiosis is a dreadful vascular infection that presents as a chronic leg ulcer. To distinguish this infectious disease from atherosclerotic occlusion of the lower limb, we investigated the imaging appearance and presence of thick and irregular arterial wall enhancement, named crusty thrombosed arterial sign, on computed tomography images in cases with vascular pythiosis infection of lower extremities. In this study, 13 cases of vascular pythiosis of lower extremities with available images from 2016 to 2022 were reviewed and the presence of crusty thrombosed arterial signs, segments of vascular involvement, and radiological findings of hematologic disease, including hepatosplenomegaly and bone changes were recorded. Crusty arterial sign with long segmental arterial involvement was found in all cases. All cases had hepatomegaly and abnormal spleen, either splenomegaly or splenectomy, found in 12 cases (92.3%). Six cases (46.1%) had thalassemic bone changes. We proposed the pathognomonic radiologic sign of vascular pythiosis: the crusty thrombosed arterial sign, which manifested as diffusely thick and irregular arterial wall enhancement along long arterial thrombosis without skip lesions. Other associated findings included splenomegaly, splenectomy, and thalassemic bone changes. These radiologic findings facilitated the diagnosis of vascular pythiosis, particularly in cases of atypical presentation or unreliable clinical context.

Ex vivo observation of Pythium insidiosum-antigen treated neutrophils on three Pythium insidiosum strains isolated from vascular pythiosis patients

ABSTRACT The mechanisms of Pythium insidiosum-antigen (PIA) immunotherapy activating a patient’s immune system are unknown. We evaluated the interleukin-8 (IL-8) serum levels during P. insidiosum infection and after vaccination with PIA in vascular pythiosis cases. Furthermore, we studied the anti-P. insidiosum activity of neutrophils stimulated with various concentrations of PIA ex vivo in 3 strains of P. insidiosum isolated from vascular pythiosis patients. IL-8 serum levels were evaluated using the ELISA technique. We assessed the effect of PIA-stimulated neutrophils on the viability of zoospores using MTT assay, visualized neutrophil extracellular trap (NET) formation via microscopy, and measured the levels of double-stranded DNA (dsDNA) using PicoGreen dsDNA quantitation assay in 3 strains of P. insidiosum isolated from vascular pythiosis patients. Serum levels of IL-8 gradually lowered from the early to the end phases of vaccination with PIA among the surviving group of vascular pythiosis cases. Neutrophils stimulated with 0.01 µg/ml PIA reduced zoospore viability significantly compared to PIA-unstimulated neutrophils for strain 1 and strain 3 (p < .05). Neutrophils stimulated with 0.01, 0.1, 1, and 10 µg/ml PIA exhibited significantly lower zoospore viability than PIA-unstimulated neutrophils for strain 2 (p < .05). IL-8 can be used as a biomarker for monitoring vascular pythiosis cases treated with the PIA vaccine. Also, anti-P. insidiosum activity of PIA-stimulated neutrophils was probably due to the disruption of cellular activity in zoospores rather than the mechanisms based on the formation of NETs.

Retrospective Review of Management and Overall Survival Rate of Patients With Vascular Pythiosis of the Lower Extremity: 20 Years Experience.

Pythiosis is an infectious disease in humans and animals caused by an aquatic fungus-like micro-organism, Pythium insidiosum. Vascular pythiosis is a difficult condition to treat and can lead to loss of limb in addition to being a potentially life-threatening infection. The condition is furthermore unfamiliar among healthcare workers, which often results in delayed treatment or even misdiagnosis. In this study, we report our findings, which have been gathered over a 20-year period in caring for vascular pythiosis in Maharaj Nakorn Chiang Mai Hospital, Chiang Mai, Thailand. We made a retrospective medical review of 32 patients presented with arterial occlusion who have serum anti-Pythium insidiosum antibodies. All patients underwent computed tomography angiography to confirm the level of arterial occlusion and decided on a treatment plan. Twelve out of 22 patients with infrainguinal disease, femoropopliteal or below-knee vascular occlusion, survived. The mean survival time is 6.58 years. Eight in 10 patients presented with suprainguinal disease died during the follow-up with a mean survival time of 31.6 months. The suprainguinal extension of the disease influenced the outcome, resulting in a higher mortality rate. However, patients presented with chronic limb ischemia had a much greater rate of survival compared to other clinical presentations. Extensive surgical resection combined with antifungal treatment and immunotherapy have proven to be effective in patients with vascular pythiosis in our findings.

374. A Multicenter Open-Label Single-Arm Clinical Trial of Combination Therapy of Surgery, Itraconazole, Doxycycline, and Azithromycin for Vascular Pythiosis

Abstract Background Vascular pythiosis, caused by Pythium insidiosum, is a neglected, life-threatening condition which carried a 100% mortality rate, within 3 months, among patients with residual disease after surgery, regardless of antifungal treatment. Recently, Thai P. insidiosum isolates were found to susceptible to doxycycline and azithromycin, with evidence of synergy. We report results of a clinical trial from 15 centers in Thailand (Thai Clinical Trial Registry number TCTR20191217006). Methods This study enrolled patients with vascular pythiosis who had 2 of the 4 diagnostic criteria: consistent clinical presentation, positive serum P. insidiosum-specific antibodies, positive cultures, or positive histopathology. All received surgery, itraconazole, doxycycline, and azithromycin. Patients were followed up until 6 months after diagnosis. Subsequent follow-ups after 6 months were at the clinician’s discretion if there were any concerns for residual diseases or unresectable arterial lesions. Results 40 patients were enrolled, with a median age (interquartile range [IQR]) of 51 (45-56) years. 38 (95%) had major thalassemia, 1 (2.5%) had liver cirrhosis, and 1 (2.5%) had myeloproliferative syndrome. None had known immunocompromised conditions. After surgery, 14 (35%) still had residual diseases. At 6 months after diagnosis, 3 (7.5%) patients died (disseminated pythiosis, sepsis with multi-organ failure, and ESBL Klebsiella pneumoniae bacteremia). Among patients with residual disease, 1 (7.1%) died and 13 are still alive. Median follow-up time (IQR) among patients with residual disease was 385 (209-622) days. There were no statistical differences in time from onset to surgery and time from onset to first antimicrobial between patients with and without residual disease after surgery (Table). One patient had to stop doxycycline prematurely due to photosensitivity. Patient characteristics IQR: interquartile range; N: number Conclusion The new treatment combination with surgery, itraconazole, azithromycin, and doxycycline improved survival among patients with vascular pythiosis who had residual disease. Disclosures Nitipong Permpalung, MD, MPH, Alcimed: Advisor/Consultant|CareDx: Grant/Research Support|Cidara Therapeutics: Grant/Research Support|Clarion: Advisor/Consultant|ClearView: Advisor/Consultant|IMMY Diagnostics: Grant/Research Support|Merck: Grant/Research Support|Scynexis: Grant/Research Support

M1.2 Human pythiosis

Meet the expert session, September 22, 2022, 8:00 AM - 9:00 AMHuman pythiosis is a rare, life-threatening infection, which is generally caused by Pythium insidiosum, a fungal-like oomycete. Four forms of human pythiosis are described: 1) vascular pythiosis affecting the patient's arteries causing arteritis, thrombosis, and gangrene; 2) ocular pythiosis, mainly causing infections of the cornea; 3) skin and soft tissue infections (cutaneous and subcutaneous pythiosis); and 4) disseminated pythiosis. Vascular pythiosis is associated with high morbidity and has a mortality rate of 10%-40%, which attributes to difficulties in diagnosis of the infection and the lack of effective standard treatment. Rice farmers and patients with thalassemia are at risk for vascular infection. Early diagnosis of Pythium infection requires a high index of clinical suspicion and is essential for good treatment results. Microscopic morphology of Pythium spp. resembles other non-septate hyphae, such as agents of mucormycosis. Definite laboratory diagnosis includes tissue cultivation with zoospore induction, polymerase chain reaction (PCR), and detection of Pythium antibodies. Radical surgery together with immunotherapy, and anti-fungal agents (e.g., terbinafine and itraconazole) have previously been used for the treatment of human pythiosis. However, patients with incomplete surgical resection had almost 100% mortality. A novel therapeutic approach and reliable biomarkers are needed to improve patient outcomes. There are several studies demonstrating excellent in vitro activity of antibacterial agents, especially macrolides, tetracyclines, and oxazolidinone against P. insidiosum. There are also evidences of synergy among antibacterial classes such as tetracyclines and macrolides. In 2019, Susaengrat, et al. reported successful treatment of two patients, who had inoperable intra-abdominal vascular pythiosis, with adjunctive antibacterial agents as salvage therapy. The medications included itraconazole in combination with doxycycline and azithromycin or clarithromycin. This case report also confirmed the result of a previous study by Worasilchai, et al. (2018) that serum 1,3-beta-D-glucan (BDG) is useful for monitoring disease activity. The ongoing multicenter prospective study is conducted to evaluate the use of antibacterial agents (azithromycin and doxycycline) with itraconazole and surgery in vascular pythiosis and to evaluate the alternative markers such as erythrocyte sedimentation rate (ESR) and c-reactive protein (CRP). Torvorapanit, et al. (2021) reported preliminary results of 10 patients, which demonstrated favorable outcomes. It was shown that 2/4 patients with the residual disease could successfully be treated with mediations. ESR and CRP declined over time after treatment and Spearman's correlation between ESR and BDG was 0.65, and between CRP and BDG was 0.6. The study aims to recruit 50 patients, which should attest the usefulness of antibacterial agents and these markers in patients with vascular pythiosis.

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

BackgroundPythium, 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 presentationA 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.ConclusionsTo 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 β-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.

In Vitro Susceptibility of Thai Pythium insidiosum Isolates to Antibacterial Agents.

Human pythiosis is a life-threatening human disease caused by Pythium insidiosum In Thailand, vascular pythiosis is the most common form and carries a mortality rate of 10 to 40%, despite aggressive treatment with radical surgery, antifungal agents, and immunotherapy. Itraconazole and terbinafine have been the mainstay of treatment, until recently, based on case report data showing potential synergistic effects against Brazilian P. insidiosum isolates. However, the synergistic effects of itraconazole and terbinafine against Thai P. insidiosum isolates were not observed. This study tested the in vitro susceptibilities of 27 Thai human P. insidiosum isolates (clade II, n = 17; clade IV, n = 10), 12 Thai environmental P. insidiosum isolates (clade II, n = 4; clade IV, n = 8), and 11 non-Thai animal P. insidiosum isolates (clade I, n = 9; clade II, n = 2) to antibiotics in eight antibacterial classes to evaluate alternative effective treatments. Tetracycline and macrolide antibiotics demonstrated in vitro activity against Thai P. insidiosum isolates, with doxycycline MICs (1 to 16 μg/ml), minocycline MICs (1 to 4 μg/ml), tigecycline MICs (1 to 4 μg/ml), azithromycin MICs (1 to 16 μg/ml), and clarithromycin MICs (0.125 to 8 μg/ml) being the lowest, on average. Synergistic effects of tetracyclines and macrolides were also observed.

Recent update in diagnosis and treatment of human pythiosis.

Human pythiosis is an infectious condition with high morbidity and mortality. The causative agent is the oomycete microorganism Pythium insidiosum. The pathogen inhabits ubiquitously in a wet environment, and direct exposure to the pathogen initiates the infection. Most patients with pythiosis require surgical removal of the affected organ, and many patients die from the disease. Awareness of pythiosis among healthcare personnel is increasing. In this review, we summarized and updated information on the diagnosis and treatment of human pythiosis. Vascular and ocular pythiosis are common clinical manifestations. Recognition of the typical clinical features of pythiosis is essential for early diagnosis. The definitive diagnosis of the disease requires laboratory testing, such as microbiological, serological, molecular, and proteomic assays. In vascular pythiosis, surgical intervention to achieve the organism-free margin of the affected tissue, in combination with the use of antifungal drugs and P. insidiosum immunotherapy, remains the recommended treatment. Ocular pythiosis is a serious condition and earliest therapeutic penetrating keratoplasty with wide surgical margin is the mainstay treatment. Thorough clinical assessment is essential in all patients to evaluate the treatment response and detect an early sign of the disease recurrence. In conclusion, early diagnosis and proper management are the keys to an optimal outcome of the patients with pythiosis.

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 β-D-glucan (BG) has been used to monitor disease response after treatment initiation in vascular pythiosis. A significant decrease in BG levels within 2weeks 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.

280. Monitoring Serum Erythrocyte Sedimentation Rate (ESR) and C-Reactive Protein (CRP) in Correlation with (1–3)-β-d-Glucan Levels in Vascular Pythiosis: A Preliminary Study

BackgroundVascular pythiosis, caused by Pythium insidiosum, is a life-threatening disease despite aggressive treatment. Our recent study showed that serum β-d-glucan (BG) trends can be used to monitor disease activity after treatment initiation. A significant decline in BG by 0.5 month indicated complete resection without residual disease. However, BG assay is cost probihitive and available only at King Chulalongkorn Memorial Hospital in Thailand. This study was conducted to preliminarily evaluate erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) as monitoring tools.MethodsA prospective study included proven vascular pythiosis patients receiving treatment with combination therapy from November 2018 to January 2019. Clinical information, BG, ESR, and CRP were collected at 0, 0.5, 1.5, and 3 months post diagnosis. Spearman’s correlation coefficient and analysis of response profiles were used.ResultsSix patients were enrolled. All had thalassemia. Four developed disease at popliteal artery, 1 at common iliac artery, and 1 at brachial artery. All underwent amputation with negative surgical margins achievement. All received itraconazole, azithromycin, and P. insidiosum immunotherapy. One received terbinafine and one received doxycycline additionally. All had positive BG > 500 pg/mL at diagnosis. After treatment initiation, means of ESR were significantly decreased at 0.5 months (P = 0.02). Means of CRP were not significantly changed until 1 month (P = 0.02) (Figure 1a and b). Correlation coefficients between BG and ESR vs. BG and CRP were 0.74 and 0.65, respectively. All survived without relapse at 3 months.ConclusionESR and CRP are potentially valuable markers to monitor vascular pythiosis in resource-limited countries. However, ESR levels and trends seem to be correlated with BG better than CRP. Further studies are needed to enroll more patients, especially patients with incomplete resection or non-surgical candidates. Disclosures All authors: No reported disclosures.

Monitoring Anti-Pythium insidiosum IgG Antibodies and (1→3)-β-d-Glucan in Vascular Pythiosis.

Despite aggressive treatment, vascular pythiosis has a mortality rate of 40%. This is due to delays in diagnosis and a lack of effective monitoring tools. To overcome this drawback, serum beta-d-glucan (BG) and P. insidiosum-specific antibody (Pi-Ab) were examined as potential monitoring markers in vascular pythiosis. A prospective cohort study of vascular pythiosis patients was carried out from January 2010 to July 2016. Clinical information and blood samples were collected and evaluated by the BG and Pi-Ab assays. Linear mixed-effect models were used to compare BG and Pi-Ab levels. The in vitro susceptibility test was performed with all P. insidiosum isolates from culture-positive cases. A total of 50 patients were enrolled: 45 survived and 5 died during follow-up. The survivors had a significantly shorter time to medical care (P < 0.0001) and a significantly shorter waiting time to the first surgery (P < 0.0001). There were no differences in BG levels among the groups at diagnosis (P = 0.33); however, BG levels among survivors were significantly lower than those of the deceased group at 0.5 months (P < 0.0001) and became undetectable after 3 months. Survivors were able to maintain an enzyme-linked immunosorbent assay (ELISA) value (EV) of Pi-Ab above 8, whereas the EV among deceased patients was less than 4. In vitro susceptibility results revealed no synergistic effects between itraconazole and terbinafine. This study showed that BG and Pi-Ab are potentially valuable markers to monitor the disease after treatment initiation. An unchanged BG level at 2 weeks after surgery should prompt an evaluation for residual disease.

Vascular pythiosis of carotid artery with meningitis and cerebral septic emboli: A case report and literature review

Vascular Pythiosis caused by Pythium insiodiosum rarely involves carotid artery. A case of concealed ruptured pseudoaneurysm of the carotid artery with neck abscesses, and cerebral septic emboli is described. Patient presented with large pulsatile neck mass that failed to response to surgery, antifungals and immunotherapeutic vaccine. Residual unresectable disease leads to death in the patient. Pythiosis should be considered as a differential diagnosis of head and neck infection.

Clinicopathological features and outcomes of pythiosis

ObjectivesVascular pythiosis is a life-threatening infection caused by the oomycete Pythium insidiosum. This article reports the clinical presentation, serodiagnosis, pathology, and outcomes seen at the authors’ institution. MethodsThe cases of patients with proven vascular pythiosis at Ramathibodi Hospital, Mahidol University, Bangkok, Thailand from January 2006 to December 2016 were analyzed retrospectively. ResultsThirteen patients were analyzed, eight of whom had underlying thalassemias. Of the remaining five patients, one had aplastic anemia, one had myelodysplasia, one had acute leukemia, one had cirrhosis, and one had alcoholism. Neutropenic patients showed a rapid clinical deterioration. Atypical presentations including carotid arteritis, aneurysm, brain abscess, and stroke occurred in the non-thalassemic patients. Serology yielded positive results in all cases, with a rapid turnaround time. Serology has the advantage of providing a presurgical diagnosis, which allows prompt surgery and clinical cure to be achieved. Pathology revealed a neutrophilic response in the acute phase and a later shift to granuloma. Immunotherapy in combination with itraconazole and terbinafine was given. The amputation rate was 77%, and disease-free surgical margins were achieved in five cases (38%). The mortality rate was 31%. ConclusionsThis study highlights new aspects of pythiosis, such as the unusual host, clinical presentation, serology as a marker for rapid diagnosis, histopathology, and outcomes. Early recognition of the disease with prompt multimodality treatment may improve survival.

Outcomes and factors influencing prognosis in patients with vascular pythiosis

Vascular pythiosis, caused by Pythium insidiosum, is associated with a high mortality rate. We reviewed the outcomes and established the factors predicting prognosis of patients treated in our institution with surgery, antifungal therapy, or immunotherapy. We undertook a retrospective record review of patients with vascular pythiosis treated in Siriraj Hospital, Bangkok, Thailand, between January 2005 and January 2015. Patient characteristics, type of surgery, adjunctive antifungal treatment, adjunctive immunotherapy, and disease status of surgical arterial and surrounding soft tissue margins were recorded. We calculated the mortality rate and established factors predicting prognosis. The records of 11 patients were reviewed. All patients had thalassemia. Nine patients (81.8%) had a history of contact with contaminated water. The clinical presentations were chronic ulcers (45.5%), toe gangrene (27.3%), pulsatile mass (27.3%), and acute limb ischemia (27.3%). Above-knee amputation was required in 10 patients (90.9%). The mortality rate was 36.4%. Independent variables between survivors and nonsurvivors were lack of an arterial disease-free surgical margin (P= .003), lack of a surrounding soft tissue disease-free surgical margin (P< .05), a suprainguinal lesion (P< .05) and duration of symptoms (P< .05). Adjuvant itraconazole, terbinafine, and Pythium vaccine have a role to play in patients with a disease-free arterial surgical margin but in whom infected surrounding soft tissue could not be completely excised. Achieving adequate disease-free surgical margins-especially the arterial margin-at amputation or débridement is the most important prognostic factor in patients with vascular pythiosis. Early detection combined with a multidisciplinary approach to treatment, including surgery, antifungal agents, and immunotherapy, allows the best possible outcome to be obtained.

Treatment outcomes of surgery, antifungal therapy and immunotherapy in ocular and vascular human pythiosis: a retrospective study of 18 patients

Human pythiosis is a life-threatening disease for which no standard treatment protocols with proven efficacy exist. We present the results of our institutional pythiosis treatment protocol, composed of surgery, antifungal agents, iron chelator (only vascular cases) and immunotherapy. We retrospectively analysed patients with proven vascular and ocular pythiosis in King Chulalongkorn Memorial Hospital from April 2003 to May 2013. Fisher's exact test and Wilcoxon's rank-sum test were used. The MICs of seven antifungal agents and combination drugs were investigated in eight clinical Pythium insidiosum strains. Eighteen patients were evaluated. Disease-free surgical margins were obtained in all surviving patients with vascular pythiosis (P = 0.08). Patients who underwent eye enucleation were significantly older than those who did not (P < 0.05). Patients with vascular or ocular pythiosis did not differ significantly in the median time from disease onset to first surgery or in the relationship between the type of P. insidiosum antigen and treatment outcomes. In vitro susceptibility profiles of all isolates demonstrated that no single agent or combination treatment was substantially more effective than the others. The highest MIC was detected for amphotericin B, followed in order by voriconazole, fluconazole, anidulafungin, caspofungin, itraconazole and terbinafine. No synergistic effects of the combination drug treatments were found. Surgery with adequate surgical margins is a crucial determinant of survival in patients with vascular pythiosis. Itraconazole and terbinafine do not have synergistic effects on Thai P. insidiosum strains. The role of immunotherapy remains inconclusive for both vascular and ocular pythiosis.

Pythiosis: An endemic fatal infectious disease

Pythiosis is a life threatening infectious disease of humans and animals living in tropical and subtropical areas of the world. The causative agent is the fungus-like microorganism Pythium insidiosum , which is a member of the oomycetes. Most of the pathogenic oomycetes infect plants, several infect animals, and P. insidiosum is capable of infecting humans. Pythiosis in animals has been increasingly found worldwide, while the disease state in humans has been mostly reported in Thailand. Pythiosis has high rate of morbidity and mortality. Patients usually come with clinical features associated with an infection of eye (called ocular pythiosis, which presents with cornea ulcer or keratitis) or arteries (called vascular pythiosis, which presents with claudication and gangrenous ulcer at leg). Diagnosis of pythiosis can be difficult and delayed. Conventional antifungal drugs are not effective against P. insidiosum . Main treatment option for most patients with pythiosis is radical excision of infected organs. In advanced case, death is a final outcome. Information on the disease and its causative agent is limited. It is apparent that more needs to be done in the way of basic and applied research to provide insights into the disease, and thereby lead to the discovery of novel strategies for diagnosis and infection control, such as, better diagnostic assays, new antimicrobial agents, and vaccines. This talk will present update information on the disease, including biology of P. insidiosum , and diagnosis and treatment of pythiosis.

Vascular pythiosis in a thalassemic patient presenting as bilateral leg ulcers

Human pythiosis is an emerging disease caused by Pythium insidiosum, a fungus-like aquatic organism. Clinical presentations can be classified into four types: (i) cutaneous/subcutaneous, (ii) ocular, (iii) vascular, and (iv) disseminated pythiosis. Serological tests such as immunodiffusion and immunochromotographic test are useful to make rapid diagnosis in cutaneous and vascular pythiosis. We report a case of 35 year-old male with vascular pythiosis of both legs, diagnosed by serology and molecular techniques.