Subcutaneous pythiosis in human treated successfully with antimicrobial treatment, debridement and immunotherapy.

Pythiosis is caused due to a filamentous eukaryotic micro-organism called Pythium insidiosum and the disease occurs commonly in horses and cattle. Subcutaneous pythiosis infection in humans is rare with no clear clinical guidelines for treatment. We present a case of a man in his 20s with non-resolving ulcers noted over lower extremity after exposure to swamp water draining animal remains. The patient received several courses of oral antibiotics with no improvement in symptoms before getting admitted to our institution. A diagnosis of subcutaneous pythiosis was made after deep wound culture following debridement detected P. insidiosum by use of PCR. Due to the rare incidence of such infection in humans and no clear guidelines available for treatment, the case was discussed with infectious disease specialists outside our institution and with veterinary physicians. An emergent approval for use of immunotherapy in conjunction with surgical debridement and antimicrobials was obtained from Food and Drug administration. The patient underwent successful treatment of infection and skin graft following treatment.

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

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.

Needs assessment of a pythiosis continuing professional development program.

BACKGROUND: Pythiosis is a rare disease with high mortality, with over 94% of cases reported from Thailand and India. Prompt diagnosis and surgery improves patient outcomes. Therefore, continuing professional development (CPD) is essential for early recognition. However, a needs assessment related to a pythiosis CPD program has not been performed. OBJECTIVES: We conducted a needs assessment to develop a pythiosis CPD program. PATIENTS/METHODS: We conducted a survey study with 267 King Chulalongkorn Memorial Hospital residents (141 internal medicine (IM) residents and 126 surgery residents). A 30-item survey consisting of a knowledge assessment, demographic section, and an attitudes portion was distributed both electronically and via paper. The data was analyzed with descriptive and inferential statistics. RESULTS: Sixty-seven percent completed the survey (110/141 IM residents, 70/126 surgery residents). The mean score [95% confidence interval] on the knowledge assessment was 41.67% [39.64%-43.69%] across all objectives. The three domains with the highest scores were pythiosis risk factors (67.22% correct), microbiologic characteristics (50.83%), and radiographic interpretation (50.56%). The three domains with the lowest scores were laboratory investigation (15.00%), epidemiology (29.17%), and symptomatology (30.83%). Most participants noted that the program should be online with both synchronous and asynchronous sessions, with a preferred length of 60-90 minutes per session. CONCLUSION: The pythiosis CPD program should emphasize education regarding symptomatology, laboratory investigation, and epidemiology, all of which are critical for the early detection of pythiosis to decrease mortality from this devastating disease. Most respondents felt this program was necessary and should be implemented in a virtual blended format.

Case Series: Mixed Infectious Keratitis by Pythium insidiosum and Fungal Species.

SIGNIFICANCE: This case series is the first to illustrate mixed infection from Pythium sp. and fungal species in corneal ulcer. PURPOSE: This case series aimed to alert all toward the possibility of both Pythium sp. and fungal species infection in case of nonresponding corneal ulcer treated with either antifungals or antipythium drugs alone. Increased suspicion of mixed infection in case of nonresponding fungal/ Pythium keratitis may facilitate early and prompt management. CASE REPORTS: Six patients presented with signs of either fungal or Pythium keratitis. They underwent ophthalmological examinations, smear examinations, cultures, and polymerase chain reaction (PCR). Therapeutic penetrating keratoplasty was performed in cases where symptoms worsened after treatment with either antifungal or antipythium drugs. The half corneal button (HCB) was shared for histopathological and microbiological examinations. In the first case, smear examination from corneal scraping (CS) revealed Pythium -like filaments, which were confirmed with PCR; however, Aspergillus nidulans grew in culture. In the second case, iodine-potassium iodide (IKI) staining was positive for Pythium ; however, PCR was positive for both Pythium and fungus, which was further confirmed by DNA sequencing. In the third case, IKI staining and HCB were positive for Pythium ; however, PCR was positive for fungus, which was identified as Candida saitoana with DNA sequencing. In the fourth case, Pythium grew in the CS culture; however, Candida sp. grew in the HCB culture. In the fifth case, Cladosporium sp. grew in culture from CS; however, Pythium insidiosum grew from the anterior chamber exudate after therapeutic penetrating keratoplasty. In the sixth case, smear examination revealed septate fungal filaments, and Cladosporium sp. grew in culture; however, HCB on histopathological examination showed features of Pythium keratitis. CONCLUSIONS: In unresponsive cases of Pythium or fungal keratitis, diagnostic modalities such as IKI and PCR should be implemented as a routine practice, in addition to smears and cultures.

Understanding barriers, recommended solutions, and future prospects for the diagnosis and management of Pythium insidiosum keratitis.

Pythium insidiosum keratitis (PIK) is a devastating corneal infection resulting in blindness in a large number of cases. Clinically and morphologically, it closely mimics fungal keratitis, and hence is also labeled as "parafungus." Although many clinical studies have documented evidence regarding the virulence of microorganism, and anatomical and functional outcomes, it remains a clinical challenge and diagnostic dilemma for most clinicians. Till today, PIK is being diagnosed and treated with certainty at only limited centers across the globe. But the question is why this is so? Taking this as the research question, this section on current ophthalmology aims to highlight the understanding of barriers to diagnosing and treating PIK, the suggestions to improve diagnosis and treatment, and the future prospects.

Outcomes of therapeutic penetrating keratoplasty in Pythium insidiosum keratitis managed with a combination of antibiotics.

Purpose: To describe the clinical outcomes of therapeutic penetrating keratoplasty (TPK) in patients with Pythium insidiosum keratitis following treatment with anti-pythium therapy (APT) consisting of linezolid and azithromycin. Methods: A retrospective review of medical records from May 2016 to December 2019 of patients with P. insidiosum keratitis was carried out. Patients who were treated with APT for a minimum of 2 weeks and then subsequently underwent TPK were included in the study. Data on demographic characteristics, clinical features, microbiology characteristics, and intraoperative details, postoperative outcomes were documented. Results: A total of 238 cases of Pythium keratitis were seen during the study period and 50 cases that satisfied the inclusion criteria were included. The median of the geometric mean of the infiltrate was 5.6 mm (IQR 4.0-7.2 mm). The patients received topical APT for a median of 35 days (IQR 25-56) prior to surgery. The most common indication of TPK was worsening keratitis (41/50, 82%). No recurrence of infection was observed. An anatomically stable globe was noted in 49/50 eyes (98%). The median graft survival rate was 2.4 months. A clear graft was present in 10 eyes (20%) with a final median visual acuity of 20/125 after a median follow-up period of 18.4 months (IQR 11-26 months). Graft size of less than 10 mm [OR: 5.824 (CI:1.292-41.6), P = 0.02] was found to be significantly associated with a clear graft. Conclusion: Performing TPK following the administration of APT has good anatomical outcomes. A smaller graft of <10 mm was associated with a higher chance of graft survival.

Clinical differentiation of keratitis due to fungus and Pythium: A photographic survey.

Purpose: To evaluate the knowledge and skills of clinicians in differentiating cases of Pythium from fungal keratitis based on their clinical appearance. Methods: Thirty slit-lamp photographs from culture-proven cases of Pythium and fungal keratitis were selected for conducting a prospective online-based photographic survey. The participants were asked to identify the causative organism and enlist the specific clinical features which helped in reaching the diagnosis. Both cornea specialists and clinical fellows/trainees participated in the survey. Sensitivity, specificity, and predictive values were estimated, and the factors associated with these, including the identification rates of clinical signs, were evaluated using bivariate and multivariate linear regression analyses. The identification rates between the consultants and fellows were compared. Results: A total of 42 cornea specialists (28 consultants and 14 clinical fellows) participated in the survey. Clinicians were able to differentiate Pythium from fungal etiology in only 56% of cases. The differentiating ability between the consultants and fellows was statistically insignificant. Average sensitivity, specificity, and positive and negative predictive values for differentiating Pythium from fungus were 56%, 65%, 56%, and 66%, respectively. The specific clinical features enlisted by the participants to differentiate Pythium from fungus were the presence of tentacles, peripheral guttering, dot-like infiltrates, and elevated plaques. Conclusion: Clinical knowledge regarding the signs and symptoms and the microbiological identification of features for Pythium are still lacking among clinicians. As the treatment protocols are distinctly different for both, it is imperative to enhance the knowledge and diagnosing ability to tackle this emerging microorganism which causes high morbidity.

Pythium insidiosum keratitis: Review of literature of 5 years' clinical experience at a tertiary eye care center.

BACKGROUND: Pythium insidiosum is an important cause of infectious keratitis from tropical and sub-tropical countries. Due to its closely mimicking clinical and microbiological features with fungus, it remained unidentified and managed as fungal keratitis for a long time. Previously all patients had poor outcomes with antifungal therapy and needed surgical treatment with higher rates of recurrences of infection leading to loss of an eye. Thus, a novel approach was required to treat it and, in this article, we would like to elaborate on the drastic change which these 5 years have brought in the management of this condition. METHODS: In view of making a consolidated article comprising all the required information and also our clinical experience in the management of Pythium keratitis, we extensively reviewed several articles available on it over PubMed and Google scholars. Relevant literature describing details about Pythium, its clinical correlation, and recent advances from 52 articles including 12 articles from our group were finally included. RESULTS: Our group identified and highlighted the unique clinical and microbiological features of Pythium insidiosum, performed several in-vitro, in-vivo studies along with clinical trials, and proposed the strategic way of its diagnosis and treatment. The use of antifungals was replaced with antibacterial medications and this resulted in better medical and surgical outcomes. CONCLUSION: The diagnosis and management of Pythium insidiosum is constantly evolving with several recent works pointing out the possible changes in the practice patterns for the management of this challenging form of keratitis.

Role of Immunotherapy in Pythium insidiosum Keratitis.

Pythium keratitis is a potentially devastating ocular condition. Incidence of Pythium keratitis has been reported in tropical and subtropical areas. In previous reports, there were no effective or standard treatments, and combinations of medication, immunotherapy, and surgery were proposed. Pythium insidiosum antigen immunotherapy (PIAI) showed an acceptable safety profile, but its efficacy is questionable in Pythium keratitis. This retrospective review included 10 eyes from 10 patients. All cases were confirmed diagnosis of P. insidiosum keratitis by culture and/or polymerase chain reaction. Three doses of PIAI were injected at 2-week intervals in all patients. The infiltration diameter ranged from 5.2 mm to total corneal involvement, and eight cases (80%) had hypopyon. Therapeutic penetrating keratoplasty (TPK) or scleral graft were undertaken in nine cases. Enucleation was done in one case on the first visit. A second TPK was undertaken in three cases, and two globes were saved. Two cases in the globe salvage group received voriconazole via eyedrops and intracameral injection. No case received either linezolid or azithromycin. Three of nine eye globes (33.33%) were saved. PIAI did not show efficacy in the treatment of Pythium keratitis. Radical surgery including resurgery in recurrence is an approved effective treatment. The recently reported medications may offer supportive management.

Pythium insidiosum keratitis - A review.

Pythium insidiosum is an oomycete and is also called "parafungus" as it closely mimics fungal keratitis. The last decade saw an unprecedented surge in Pythium keratitis cases, especially from Asia and India, probably due to growing research on the microorganism and improved diagnostic and treatment modalities. The clinical features such as subepithelial infiltrate, cotton wool-like fluffy stromal infiltrate, satellite lesions, corneal perforation, endoexudates, and anterior chamber hypopyon closely resemble fungus. The classical clinical features of Pythium that distinguish it from other microorganisms are reticular dots, tentacular projections, peripheral furrowing, and early limbal spread, which require a high index of clinical suspicion. Pythium also exhibits morphological and microbiological resemblance to fungus on routine smearing, revealing perpendicular or obtuse septate or aseptate branching hyphae. Culture on blood agar or any other nutritional agar is the gold standard for diagnosis. It grows as cream-colored white colonies with zoospores formation, further confirmed using the leaf incarnation method. Due to limited laboratory diagnostic modalities and delayed growth on culture, there was a recent shift toward various molecular diagnostic modalities such as polymerase chain reaction, confocal microscopy, ELISA, and immunodiffusion. As corneal scraping (10% KOH, Gram) reveals fungal hyphae, antifungals are started before the culture results are available. Recent in vitro molecular studies have suggested antibacterials as the first-line drugs in the form of 0.2% linezolid and 1% azithromycin. Early therapeutic keratoplasty is warranted in nonresolving cases. This review aims to describe the epidemiology, clinical features, laboratory and molecular diagnosis, and treatment of Pythium insidiosum keratitis.

Pythium keratitis: Clinical profile, laboratory diagnosis, treatment, and histopathology features post-treatment at a tertiary eye care center in Eastern India.

Purpose: The aim of this work was to study demography, clinical profile, laboratory diagnosis, and management of Pythium keratitis at a tertiary eye care center in Eastern India. Methods: Eighteen patients with culture-positive Pythium keratitis managed at our center between January 2016 and December 2018 were included in this retrospective study. Clinical features, laboratory investigations, treatment, and outcomes were analysed. Results: Pythium keratitis commonly affects middle-aged males with low socioeconomic profile and history of trauma. Samples stained with Gomori methenamine silver showed 93.8% positivity and Iodine-potassium iodide-sulfuric acid showed 100% positivity. Periodic acid-Schiff's showed negative staining in 62.5% and weak in 37.5%. Kirby-Bauer disc diffusion method showed zone of inhibition as 30.25 ± 4.61 mm for Linezolid and 23.56 ± 6.86 mm for Azithromycin. Medical management included topical/oral linezolid and azithromycin. Therapeutic penetrating keratoplasty (TPK) was done in 15 eyes (83.3%), repeat TPK in 4 eyes, and evisceration in 3 eyes (16.7%). One patient required only medical treatment. Globe salvation was obtained in 15 (83.3%) eyes, and good visual outcome in 7 eyes (38. 9%). There was graft failure in six eyes (40%) and two (11.1%) eyes went into phthisis. Patients were divided into early and late presenters. Late presenters had more complications and worse final visual outcome. Conclusion: Pythium keratitis can be differentiated from fungal keratitis by its characteristic appearance on slit-lamp examination, smear, culture, and histopathology. Early presentation, detection, and treatment with antibacterial drugs like linezolid and azithromycin results in a better prognosis. Early full-thickness corneal transplant should be considered for Pythium keratitis not responding to treatment.

Retrospective multifactorial analysis of Pythium keratitis and review of literature.

Purpose: The aim of this work was to study the demographic profile, clinical diagnostic features, challenges in management, treatment outcomes, and ocular morbidity of microbiological culture-proven Pythium keratitis in a tertiary eye care hospital in South India. Methods: Retrospective analysis of microbiologically proven Pythium keratitis patients was performed at a tertiary eye center from October 2017 to March 2020. Demographic details, risk factors, microbiological investigations, clinical course, and visual outcomes were analyzed. Results: Thirty patients were analyzed. The mean age was 43.1±17.2 years. Most common risk factors were history of injury in 80% and exposure to dirty water in 23.3%. Visual acuity at baseline was 20/30 to perception of light (PL). The most common clinical presentation was stromal infiltrate and hypopyon in 14 (46.6%) patients each. The microbiological confirmation was based on culture on blood agar and vesicles with zoospores formation with incubated leaf carnation method. Seven (23.3%) patients improved with topical 0.2% Linezolid and topical 1% Azithromycin, 19 (63.3%) patients underwent Therapeutic keratoplasty (TPK) and 4 were lost to follow-up. Seven (23.3%) patients had graft reinfection, and 3 (10%) developed endophthalmitis. The final visual acuity was 20/20- 20/200 in 6 (20%) patients, 20/240-20/1200 in 5 (16.6%) patients, hand movement to positive perception of light in 16 patients and no perception of light (Pthisis Bulbi) in 3 (10%) patients. Conclusion: P. insidiosum keratitis is a rapidly progressive infectious keratitis with prolonged and relapsing clinical course. It usually results in irreparable vision loss in majority of the patients. Prompt diagnosis, clinical awareness, and specific treatment options are needed for successfully managing this devastating corneal disease.

Predisposing Factors, Clinical Presentations, and Outcomes of Contact Lens-Related Pythium Keratitis.

PURPOSE: To describe predisposing factors, clinical presentations, and treatment outcomes of contact lens (CL)-related Pythium keratitis. METHODS: This was an 11-year retrospective study of CL-related Pythium keratitis conducted from 2009 to 2019. Six eyes of 6 patients were identified. Demographics, predisposing factors, CL history, clinical presentation, diagnostic tests, treatments, and outcomes were reviewed. RESULTS: Mean age of the patients was 34 years (SD 16.3 years) with equal proportion between male and female patients. Five of 6 patients (83.3%) used soft CL, whereas 1 patient used rigid gas permeable lens. All patients had a history of water contamination (tap water and water from river and sea). Mean duration from the onset was 7.8 days (range 4-14 days). Mean size of the corneal lesion was 3.33 mm (SD 1.31 mm) in width. The typical feature of tentacle-like lesions radiating in a reticular pattern was observed in all patients. Feathery edge (1 eye), satellite lesions (2 eyes), and radial keratoneuritis (2 eyes) were also found. Every patient received therapeutic penetrating keratoplasty because of failed medical treatments. One patient subsequently underwent enucleation. Globe salvage was achieved in 5 patients (83.33%). CONCLUSIONS: Awareness of the history of water contamination, recognition of specific clinical features of Pythium keratitis, and performing surgical treatment are key for achieving globe salvage in patients with CL-related Pythium keratitis.

Immunotherapy based on Pythium insidiosum mycelia drives a Th1/Th17 response in mice.

Pythium insidiosum is an oomycete that affects mammals, especially humans and horses, causing a difficult-to-treat disease. Typically, surgical interventions associated with antimicrobial therapy, immunotherapy, or both are the preferred treatment choices. PitiumVac® is a therapeutic vaccine prepared from the mycelial mass of P. insidiosum and is used to treat Brazilian equine pythiosis. To better understand how PitiumVac® works, we analyzed the composition of PitiumVac® and the immune response triggered by this immunotherapy in mice. We performed an enzymatic quantification that showed a total glucan content of 21.05% ± 0.94 (α-glucan, 6.37% ± 0.77 and (1,3)(1,6)-ß-glucan, 14.68% ± 0.60) and mannose content of 1.39% ± 0.26; the protein content was 0.52 mg ml-1 ± 0.07 mg ml-1. Healthy Swiss mice (n = 3) were subcutaneously preimmunized with one, two, or three shots of PitiumVac®, and immunization promoted a relevant Th1 and Th17 responses compared to nonimmunization of mice. The highest cytokine levels were observed after the third immunization, principally for IFN-γ, IL-17A, IL-6, and IL-10 levels. Results of infected untreated (Pythiosis) and infected treated (Pythiosis + PVAC) mice (n = 3) showed that PitiumVac® reinforces the Th1/Th17 response displayed by untreated mice. The (1,3)(1,6)-ß-glucan content can be, at least in part, related to this Th1/Th17 response.

In vivo effect of minocycline alone and in combination with immunotherapy against pythium insidiosum.

This study examined the effect of minocycline alone and in combination with immunotherapy against pythiosis. Twenty rabbits, aged three months old and subcutaneously inoculated with Pythium insidiosum zoospores were divided into four groups (n = 5): treated with minocycline (10 mg/kg/day twice daily), treated with immunotherapy (34 mg subcutaneously every 14 days), treated with minocycline plus immunotherapy, and untreated (control group). The treatments were started 30 days after inoculation and continued for 70 days. The subcutaneous nodular injury areas in infected groups were measured every seven days after the beginning of treatment. Only the rabbits that developed lesions were selected for this study. When compared with the control group over 70 days, the minocycline and minocycline plus immunotherapy groups of rabbits with pythiosis showed significantly reduced injuries. The histopathology showed the presence of inflammation, macrophages and eosinophils. Grocott's staining revealed irregular hyphae-like structures that were ramified and occasionally septate. Our results suggest that minocycline has fungistatic activity and that the combination of minocycline and immunotherapy is more effective than the individual therapies tested.

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.

Clinical outcomes in ocular pythiosis patients treated with a combination therapy protocol in Thailand: A prospective study.

Ocular pythiosis is the second most common form of human pythiosis, and the rates of evisceration/enucleation in Thailand are 55-79%. This prospective study was conducted to evaluate treatment outcomes of the combination therapy protocol and the potential use of serum (1→3)-ß-glucan (BG) and Pythium insidiosum-specific antibody (Pi-Ab) as an aid to diagnosis and monitoring of ocular pythiosis. Thirty patients were enrolled in the study and 14 (non-globe salvage) required evisceration/enucleation. The globe salvage group was significantly younger, and first ocular surgeries were performed significantly sooner than in the non-globe salvage group. Serum BG and Pi-Ab levels were similar among the 2 groups over time. In vitro susceptibility testing of antifungal agents revealed relatively high minimum inhibitory concentrations and lack of synergistic effect. Serum BG and Pi-Ab would not be useful in diagnosis and monitoring of ocular pythiosis. Until effective antimicrobial agents are discovered, ocular surgeries are still the mainstay therapy in Thailand.

Clinical profile, risk factors and outcome of medical, surgical and adjunct interventions in patients with Pythiuminsidiosum keratitis.

PURPOSE: To report clinical profile and compare management options for Pythium keratitis. METHOD: Retrospective interventional study of 46 patients diagnosed as Pythium keratitis by PCR DNA sequencing from January 2014 to July 2017. Interventions were categorised into medical management (MM) (topical azithromycin and linezolid with oral azithromycin at presentation), surgery (S) (therapeutic penetrating keratoplasty, TPK), surgical adjunct (SA) (cryotherapy±alcohol with TPK) and medical adjunct (MA) (MM after TPK). RESULTS: Primary treatment included MM (1 eye), SA (3 eyes) and S (42 eyes). Recurrence occurred in 27/43 eyes (MM+S group). Second surgery (S) was required in 11 eyes (TPK-2), with additional procedures (SA) in 10 eyes and evisceration in five eyes. 8/43 eyes received MA after TPK-1. One eye required TPK-3. Recurrence occured in all eyes that received MA (100%) and in 28 of 54 TPKs (51.8%) (TPK 1+2+3) in 42 eyes. Recurrence was noted in 1/14 (7.1%) that underwent SA. CONCLUSION: The currently available and recommended treatment for Pythium keratitis is surgical by means of a TPK and in worse cases evisceration. In our study, MM/MA measures showed no benefit with recurrence or worsening of infection requiring resurgery. Almost 50% of TPKs had a recurrence requiring resurgery. However, adjunctive procedures during TPK appear to have additional benefit with low risk of recurrence and could be included as routine care.

Intradermal injection of Pythium insidiosum protein antigens for improved diagnosis and treatment of pythiosis in an experimental model.

The oomycetous pathogen Pythium insidiosum is the causative agent of pythiosis, a life-threatening disease that affects animals and humans. This infectious disease is difficult to treat, and early and accurate diagnosis is critical for effective treatment. In this sense, this study aimed to evaluate the intradermal (ID) injection of P. insidiosum protein antigens (PiPA) for the diagnosis and treatment of pythiosis using an experimental model. For diagnostic purposes, PiPA were injected by the ID route in the following groups of rabbits: (a) control; (b) previously immunized with PiPA injected by the subcutaneous (SC) route; and (c) infected with P. insidiosum zoospores. For treatment purposes, rabbits with pythiosis were also treated with PiPA by the ID or SC routes. Mean induration sizes were different at 24 h and 72 h readings when compared to the control group. Sensitivity of the protocol was 100% at 24 h and 80% at 72 h, with 100% specificity in both readings. PiPA treatment using ID or SC routes did not result in significant differences in lesion sizes and cure rates; however, serum levels of interferon-gamma were higher in SC route. This study demonstrates the applicability of PiPA ID for diagnosis and treatment of pythiosis in an experimental model.