[Show abstract][Hide abstract] ABSTRACT: Pythiosis is an invasive, ulcerative, pyogranulomatous disease caused by Pythium insidiosum, a fungus-like oomycete that has been reported to affect humans, horses, dogs, and other mammals mainly in tropical and subtropical areas of the world. The disease is characterized by an eosinophilic granulomatous and a Th2 immune response which in turn helps to protect the fungus from the host cells. Pythiosis can present clinically in subcutaneous, gastrointestinal, and vascular tissues or in a systemically disseminated form depending on the species and site of infection. Changes in iron metabolism and anemia are commonly observed. The diagnosis is accomplished through clinical and pathological features, laboratory characteristics of cultures, serological and molecular tests. Treatment includes radical surgery, antimicrobial drugs, immunotherapy or a combination of these treatments. Immunotherapy is a practical and non-invasive alternative for treating pythiosis which is believed to promote a switch from a Th2 to Th1 immune response, resulting in a favorable clinical response. This therapy has demonstrated cure rates above 70% and 55% in horses and humans but low cure rates in dogs and cats. Despite the curative properties of this type of immunotherapy, the antibodies that are produced do not prevent host reinfection. Thus, development of effective adjuvants and new diagnostic techniques for early disease diagnosis are of utmost importance. The aim of this review was to promote pythiosis awareness and to provide an update about the immunotherapy and immunobiology of this disease.
[Show abstract][Hide abstract] ABSTRACT: Photodynamic therapy has been applied successfully against cutaneous and subcutaneous mycoses. We applied methylene blue as a photosensitizing agent and light emitting diode (InGaAlP) against Sporothrix schenckii complex species in an in vitro assay. The viability of the conidia was determined by counting colony-forming units. Methylene blue in conjunction with laser irradiation was able to inhibit the growth of all tested samples. The in vitro inhibition of Sporothrix spp. isolates by laser light deserves in vivo experimental and clinical studies since it may be a promising treatment for cutaneous and subcutaneous sporotrichosis.
[Show abstract][Hide abstract] ABSTRACT: We describe the in vitro activity of the combination of azithromycin, clarithromycin, minocycline and tigecycline alone and in combination with amphotericin B, itraconazole, terbinafine, voriconazole, anidulafungin, caspofungin and micafungin against 30 isolates of the oomycete Pythium insidiosum. The assays were based on the M38-A2 technique and the checkerboard microdilution method. The main synergism observed was through the combination of minocycline with amphotericin B (73.33%), itraconazole (70%) and micafungin (70%) as well as clarithromycin with micafungin (73.33%).
Antimicrobial Agents and Chemotherapy 07/2014; · 4.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to verify the trypanocidal effectiveness of aqueous, methanolic, and ethanolic extracts of Achyrocline satureioides against Trypanosoma evansi in vitro. A. satureioides extracts, known as macela, were used on trypomastigotes at different concentrations (1, 5, 10, 50, 100, 500, and 1,000 µg/ml) and exposure times (0, 1, 3, 6, and 9 hr). A dose-dependent effect was observed when the 3 extracts were tested. The concentrations of 1, 5, and 10 µg/ml were not able to kill trypomastigotes until 3 hr after exposure, and the highest concentrations (500 and 1,000 µg/ml) were able to kill all trypomastigotes after 1 hr. When the time of exposure was increased up to 9 hr, the concentrations at 50 and 100 µg/ml were 100% effective to 3 extracts. The chemical analysis of the extracts revealed the presence of flavonoids, a trypanocidal compound already described. Based on the results, we can conclude that the A. satureioides extracts exhibit trypanocidal effects.
The Korean Journal of Parasitology 06/2014; 52(3):311-5. · 0.88 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Abstract 1. The protective effect of a natural Brazilian calcium montmorillonite (CaMont) against aflatoxins was studied in broiler chickens. 2. A total of 1056 d-old Cobb male broilers were housed in experimental pens (22 chickens per pen) for 42 d. Three levels of CaMont (0, 2.5 and 5 g/kg) and two levels of aflatoxins (0 and 3 mg/kg) were assayed. Each treatment had8 replicate pens of 22 broiler chickens each. 3. Of all the chickens tested in the experiment, the ones treated with aflatoxins were the most adversely affected. CaMont treatment at concentrations of 2.5 and 5 g/kg improved body weight of chickens at 42 d of age by 13.3 and 22.7%, increased daily feed intake by 9.7 and 24.7%, and improved the productive efficiency index of chickens by 53 and 66.5%, respectively. 4. Dietary CaMont positively affected parameters such as weight of liver, heart and gizzard; however, serum potassium concentration decreased by 15.3% compared with that of chickens given only the aflatoxin-contaminated diet. 5. CaMont did not cause adverse effects in chickens that did not receive aflatoxins. 6. CaMont at pH 8.5 partially reduced the toxic effects of aflatoxins in broilers when included at levels of 2.5 and 5 g/kg in the diet.
British Poultry Science 01/2014; · 1.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A clinical epidemiological study was conducted among 34 rural properties located within the Brazilian Pantanal region and nearby areas between 2007 and 2010. The diagnosis of equine pythiosis was based on antibody detection (by enzyme-linked immunosorbent assay), polymerase chain reaction, histopathological analysis, and cultures positive for Pythium insidiosum. The majority of the affected animals (85%) were in the Pantanal biome, which had a higher disease prevalence (0.9%-66.7%) than that of the Cerrado (2.7%-33.3%). The disease was more prevalent in the rainy season (January-March), with an abrupt fall in the number of cases during the drought period (April–September; correlation of R2 = 0.77; P < .01). Generally, the average prevalence of equine pythiosis in both regions was 5%, with mortality and lethality rates of 1.3% and 23.1%, respectively, in the Pantanal and 2.3% and 45.5%, respectively, in the Cerrado. However, the treatment with immunotherapy may have underestimated these numbers, especially in the Pantanal. Animals older than 1 year were 8.09 times more affected by the disease than younger animals in the same environment (P < .05). A correlation between the anatomical area of the lesion and the type of skin color was also observed. Approximately 73% of the lesions were found in dark-pigmented areas, and animals with a dark coat color were affected more frequently. These findings highlight the importance of hematophagous insects in the epidemiology of pythiosis because these areas are preferred for blood feeding.
[Show abstract][Hide abstract] ABSTRACT: Pythium insidiosum is an important pathogen of mammals' species, including humans. Equine is the main species affected by this oomycete. P. insidiosum requires an aquatic environment to develop its life cycle, and the susceptible hosts are contaminated when they contact the microorganism in swampy areas. The equine pythiosis is characterized by the formation of irregular masses within the cutaneous lesions, called kunkers, which easily detach from the lesion. From these structures, it is possible to isolate P. insidiosum in pure cultures. The present study aimed to reproduce in vitro the life cycle of P. insidiosum from kunkers of equine clinical lesions. Fifteen kunkers from different horses were tested. It was observed that the discharge of zoospores occurred after 24-48 h of incubation at 37 °C in, respectively, 40 and 47 % of the kunkers evaluated. Only two samples showed no development of the asexual cycle of P. insidiosum under the conditions tested. It was possible to demonstrate that kunkers are able to restart the asexual cycle of P. insidiosum. Based on our in vitro results, we highlight the importance of these structures in the epidemiology of the pythiosis, since kunkers can be a potential source of contamination of this oomycete for aquatic environments.
[Show abstract][Hide abstract] ABSTRACT: Pythium insidiosum is an oomycete, a fungal like microorganism, which infects mammals, causing pythiosis in animals and humans, especially in tropical and subtropical regions around the world. The treatment for this infection is very difficult, and therapeutic options commonly comprise surgery, immunotherapy and antimicrobial drugs. The present report describes the clinical healing of a dog with gastrointestinal pythiosis by treatment with a combination of antifungals and immunotherapy, as well as reviews the cases reported in the literature that used some type of therapy for canine pythiosis. A 2.5-year-old male beagle initially showed sporadic vomiting episodes, and this symptom became more frequent 5 months after the onset of clinical signs. Celiotomy procedure found thickness of the stomach wall extending to the pylorus and duodenum. A biopsy was performed, and the diagnosis of pythiosis was made by mycological, histopathological analyses and molecular identification. Therapy was based on an association of terbinafine plus itraconazole during 12 months and immunotherapy for 2.5 months. The healing of the dog reported here allows us to propose the use of immunotherapy associated with antifungal therapy to treat canine gastrointestinal pythiosis. However, additional studies should be performed on a larger number of patients to establish a standard treatment protocol for canine pythiosis.
[Show abstract][Hide abstract] ABSTRACT: Data regarding the susceptibility of Conidiobolus lamprauges is limited and there is no consensus about the optimal treatment for infections caused by Conidiobolus spp. In this context, the objective of this study was to evaluate the in vitro susceptibility of six C. lamprauges strains isolated from sheep conidiobolomycosis to amphotericin B, ketoconazole, fluconazole, itraconazole, posaconazole, voriconazole, anidulafungin, caspofungin, micafungin, flucytosine, and terbinafine using the CLSI M38-A2 microdilution technique. Terbinafine was the most active (MIC range <0.06-0.5μg/mL). Resistance or reduced susceptibility was observed for amphotericin B and azole and echinocandin antifungals. Additional studies are necessary to determine the therapeutic potential of terbinafine as monotherapy or in combination therapy with other antifungals.
[Show abstract][Hide abstract] ABSTRACT: There is a paucity of animal models of pythiosis, a life-threatening disease of humans and animals that has poorly understood immunopathogenesis. We developed a pythiosis model by injecting Pythium insidiosum zoospores in Toll (Tl)-deficient Drosophila melanogaster flies. The infection of Tl mutant flies resulted in significantly lower survival rates (73.7%) when compared to control flies. Our study reveals the important role of Tl pathway activation in fly immune response to pythiosis.
Microbiology and Immunology 07/2013; · 1.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Here, we evaluated combinations of diphenyl diselenide [(PhSe)2] with fluconazole and amphotericin B in a checkerboard assay against clinical Candida glabrata strains. Minimal inhibitory concentration (geometric mean) ranged from 0.25 to >64 (5.16 μg/mL) for (PhSe)2, 1 to 32 (5.04 μg/mL) for fluconazole and 0.06 to 0.5 (0.18 μg/mL) for amphotericin B. Synergistic (76.66 %) and indifferent (23.34 %) interactions were observed for (PhSe)2 + amphotericin B combination. (PhSe)2 + fluconazole combination demonstrated indifferent (50 %) and antagonistic (40 %) interactions, whereas synergistic interactions were observed in 10 % of the isolates. New experimental in vivo protocols are necessary and will promote a better understanding of the antimicrobial activity of (PhSe)2 against C. glabrata and its use as an adjuvant therapy with antifungal agents.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to investigate the susceptibility in vitro of Trypanosoma evansi to the essential oils of andiroba (Carapa guianensis) and aroeira (Schinus molle), in their conventional and nanostructured forms. For that, pure oils at concentrations of 0.5, 1.0 and 2.0% were used. A negative control (untreated) and a positive control (diminazene aceturate 0.5%) were used as comparative parameters. Later, the same tests were performed, using nanoemulsions oils at concentrations of 0.5 and 1.0%. The tests were carried out in triplicates and the numbers of parasites were quantified on 1, 3 and 6 hours from onset of the study. A dose-dependent reduction in the number of parasites to the forms of two oils tested was observed after 1 hour. The concentration of parasites was significantly reduced at low concentrations after 3 hours, as well as at 6 hours no alive parasites were observed for the essential oils tested. Ours findings indicate, for the first time, that oils of andiroba and aroeira (in their conventional and nanoemulsion forms) have high activity against T. evansi in vitro, leading to the suggestion that these oils may be applied as an alternative treatment for this disease.
[Show abstract][Hide abstract] ABSTRACT: An eight-year-old draft horse suffering from scrotal pythiosis was referred to Mansoura Veterinary Teaching Hospital, Egypt, March 2013. The horse scrotal wall showed hard thickening with skin ulcerative and granulomatous lesions. The thickened mass composed of whitish yellow tissue having numerous draining tracts that discharge serosanguineous or bloody exudates. Initial diagnosis was based on location, gross lesion, and presence of kunkers. Histopathological evaluation revealed an eosinophilic granulomatous reaction associated with intralesional hyphae characteristic of Pythium insidiosum visualized by Gomori methenamine silver stain (GMS) and immunostaining using rabbit polyclonal antibody. The lesions were located at the ventral aspect of the scrotum (Figure 1) and appeared as a cutaneous nodule that ulcerated after two weeks. External examination and palpation of scrotum showed diffuse swelling and hardening with localized ulcerations. The thickened wall was composed of whitish yellow tissue having numerous draining tracts that discharge serosanguineous or bloody exudates. The lesions were restricted to the scrotum. Initial diagnosis was based on gross lesion and presence of kunkers. Castration was indicated because of the depth of the lesion and the excessive loss of scrotal tissues. The horse was pre-medicated with intravenous injection of acepromazine (Vetranquil 1%) at a dose of 0.05 mg/kg body weight (BW). Anesthesia was induced and maintained by infusion of a freshly prepared mixture of 500 mg xylazine HCl, 40 mg midazolam, and 2 g ketamine HCl dissolved in 1 L of 5% dextrose. Before surgery, the animal received slow intravenous injection of warm physiological normal saline (0.9% NaCl) solution with 5% dextrose, penicillin G sodium (20,000 i.u/kg BW), and flunixin meglumine (1.1 mg/kg BW). The affected animal was approached in dorsal recumbency. Total surgical excision of the lesion was performed. Open covered castration was performed. Transverse sections of the excised scrotal sac revealed characteristic yellow-gray, coral-like materials (kunkers or leeches) (Figure 2) within the sinus tracts and the granulomatous tissue. Kunkers were firm and gritty in consistency, with a diameter ranging from 1 mm to 12 mm. The testicles were normal in shape and size and its cut section was free from the lesion. Representative samples were taken from the scrotum after the surgical operation then immediately fixed in neutral buffered formalin and processed according to standard technique. Five-micron-thick paraffin sections were prepared and stained with hematoxylin and eosin (H&E) and Gomori methenamine silver (GMS) stain . Selected sections of scrotal lesions were immunostained with a rabbit polyclonal antibody against P. insidiosum by using a labeled streptavidin-biotin peroxidase technique as previously described . The antibodies were produced in New Zealand rabbits from a previous study at the Universidade Federal de Santa Maria (
[Show abstract][Hide abstract] ABSTRACT: Pythiosis is a devastating infectious disease caused by an aquatic oomycete, Pythium insidioum, and affects animals and humans that inhabit wetlands. The disease is characterized mainly by granulomatous lesions in the hosts. The purpose of this study was to report the occurrence of pythiosis in sheep in the states of Pernambuco (PE) and Bahia (BA), Northeastern Brazil, as well as to evaluate the efficacy of an immunotherapic against ovine pythiosis. Blood samples were collected from 53 sheep, 49 from flocks in counties located in PE and four from BA. Seven sheep showed clinical signs of ovine pythiosis; one of them was submitted to euthanasia and its head and submandibular lymph node was collected and sent for histopathologic and mycological analyses. Other six sheep were treated with an immunotherapic. During the treatment the animals were kept in the Sheep Industry Sector facilities at Univasf/Petrolina-PE. ELISA, fungal culture and polymerase chain reaction (PCR) methods were used to confirm the diagnosis of clinical ovine pythiosis in the sheep flock. At microscopic examination of the material collected from the nasal cavity of a sheep euthanized was observed a focally extensive area of necrosis with presence of diffuse infiltration of intact and degenerated neutrophils bordering the cartilage. Only one sheep showed clinical cure, indicating efficiency in the pythiosis treatment of 16.7% (1/6). Ovine pythiosis has been increasing in several municipalities of PE and BA. In this context, the immunotherapy may be an alternative to be searched. Therefore, further studies are needed to investigate the effect of immunotherapy on ovine pythiosis.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to characterize the response of acute phase proteins (APP) in rabbits experimentally infected with Trypanosoma evansi (T. evansi), and to relate the findings with serum immunoglobulins levels, in order to verify the relation between APP and the immune response of rabbits. A total of 12 animals were used in this experiment and divided into 2 groups, control and infected, of six rabbits each. The experimental period was 118days, and blood was collected on days 0, 5, 20, 35, 65, 95 and 118 post-infection (PI). The infection with T. evansi stimulated APP and immunoglobulins production, once the infected animals showed an increase in C-reactive protein, haptoglobin, alpha 2-macroglobulin and IgM levels. The elevation in IgM levels observed in this study, when related to the increase in C-reactive protein and haptoglobin levels, suggests the involvement of these proteins in host defense against flagellated protozoa, with possible participation in the control of the parasitemia in rabbits infected with T. evansi.
Research in Veterinary Science 02/2013; · 1.77 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Growth of Pythium insidiosum mycelia around minocycline disks (30μg) did not occur within 7days of incubation at 35°C when the isolates were grown on Sabouraud, corn meal, Muller-Hinton or RPMI agar. This technique offers a simple and rapid method for the differentiation of P. insidiosum from true filamentous fungi.
Journal of microbiological methods 02/2013; · 2.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVES: Iron plays an important role in the pathogenesis of Pythium insidiosum. Human pythiosis frequently occurs in iron-overloaded thalassaemic patients and experimentally infected animals develop iron deficiency anaemia. Therefore, we sought to determine the in vitro and in vivo activities of the iron chelator deferasirox against P. insidiosum. METHODS: In vitro, the MIC and minimum fungicidal concentration (MFC) values of deferasirox for 17 strains of P. insidiosum were determined in accordance with CLSI document M38-A2. In vivo studies were carried out in 20 inoculated rabbits divided into four groups: placebo, immunotherapy obtained from vortexed P. insidiosum cultures (14 day intervals), deferasirox (15 mg/kg/day) and a combination of immunotherapy and deferasirox. Five non-infected animals were used as controls. RESULTS: The MIC and MFC values of deferasirox for P. insidiosum ranged from 12.5 to 50 mg/L and from 50 to 100 mg/L, respectively. Treatment with deferasirox alone ameliorated anaemia and normalized the serum iron levels and hepatic iron concentration in the animals. However, the mean lesion size, although decreased, did not differ significantly from that in the placebo group. The results of immunotherapy plus iron chelation therapy were worse than those of immunotherapy alone. Moreover, the disease spread to the lung tissue in 5 out of 10 deferasirox-treated animals. CONCLUSIONS: Despite its limited in vitro and in vivo activity, deferasirox improved iron deficiency anaemia in P. insidiosum-infected rabbits. Further studies are needed to investigate the immunomodulatory properties observed in this study and the benefits and drawbacks of using iron-chelating drugs as an adjuvant therapy in pythiosis.
Journal of Antimicrobial Chemotherapy 01/2013; · 5.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: : Pythium insidiosum is an oomycete classified in the Kingdom Stramenipila. P. insidiosum hyphae are not able to initiate infection without the secretion of hydrolytic enzymes, which are considered an important factor in microbial virulence. AIMS: : To evaluated the extracellular enzymatic activity of 14 Brazilian P. insidiosum isolates and a standard strain (ATCC 58637) by the API-ZYM System screening method. METHODS: : Zoospores were grown in RPMI 1640 broth, and 65μL of the liquid phase was inoculated in each cupule of the API-ZYM strips. RESULTS: : Differences in the enzymatic activities were observed among the isolates, although phosphohydrolases and ester hydrolases were conspicuous among all isolates. β-glucosidase was also present in most of the isolates. Enzymatic activities of α-glucosidase and chymotrypsin were not observed, differing from a previous study involving Australian isolates and intracellular enzymes. CONCLUSIONS: : The discrepancy in the enzymatic profile observed among Brazilian P. insidiosum isolates reflects the phenotypic variations found in susceptibility tests.