Anelise Oliveira da Silva Fonseca

In vitro susceptibility of zoospores and hyphae of Pythium insidiosum to antifungals

OBJECTIVES The purpose of this study was to compare the in vitro susceptibilities of 22 Brazilian isolates of Pythium insidiosum to antifungals using a standardized inoculum of zoospores and a proposed novel inoculum prepared from cultured mycelia (hyphae) of P. insidiosum. METHODS A zoospore suspension of P. insidiosum was obtained by the zoosporogenesis technique. The hyphal inoculum was prepared from a suspension of P. insidiosum mycelium. Susceptibility to each drug was evaluated using the CLSI M38-A2 method. RESULTS Of the 88 MIC comparisons performed, 36 (41%) showed the same MIC value for the two inocula. The agreement (differences not greater than one dilution) between MICs obtained with both types of inocula was 39.8% (35/88). In other MIC comparisons analysed, 17 (19.3%) showed differences of two or three dilutions. CONCLUSIONS We conclude that the use of hyphal inocula of P. insidiosum for in vitro susceptibility tests could be a suitable method for evaluating antimicrobial susceptibility, particularly when it is not possible to obtain a standardized zoospore inoculum.

Perfil de suscetibilidade de leveduras do gênero Candida isoladas de animais ao óleo essencial de Rosmarinus officinalis L

ABSTRACT: Susceptibility profile of Candida spp. isolated from animals to the essentialoil of Rosmarinus officinalis L. In this work the antifungal activity and chemical composition ofthe Rosmarinus officinalis (rosemary) essential oil against Candida spp. were evaluated. Theessential oil was obtained by hydrodistillation of aerial parts, and analyzed by gas chromatographywith a flame ionization detector (GC/FID) and gas chromatography coupled with mass spectrometry(GC/MS). This oil was tested against fourteen isolates of Candida sp. (ten C. albicans and fourno-albicans species) by the microdillution broth assay (CLSI-M27A2). Chromatography analysesof the essential oil showed compounds, as camphor, verbenone and 1,8-cineole as majorconstituents . MIC and MFC values for C. albicans standard strains were 1.25 - 2.5 µL mL -1 and2.5 - 5.0 µL mL -1 , respectively. MIC and MFC for the non-albicans species were 1.25 – 5.0 µL mL -1 and 2.5 – 10.0 µL mL -1 , respectively. C. albicans isolates obtained from animals exhibited MIC andMFC values of 2.5 - > 10.0 µL mL

Treatment of experimental pythiosis with essential oils of Origanum vulgare and Mentha piperita singly, in association and in combination with immunotherapy.

This study investigated the in vivo antimicrobial activity of the essential oils of Origanum vulgare and Mentha piperita both singly, associated and in combination with immunotherapy to treat experimental pythiosis. The disease was reproduced in 18 rabbits divided into six groups (n=3): group 1, control; group 2, treated with essential oil of Mentha piperita; group 3, treated with essential oil of Origanum vulgare; group 4, treated with commercial immunotherapic; group 5, treated with a association of oils of M. piperita and O. vulgare and group 6, treated with a combination of both oils plus immunotherapy. Essential oils were added in a topical cream base formula, and lesions were treated daily for 45 days. The animals in groups 4 and 6 received a dose of immunotherapeutic agent every 14 days. The results revealed that the evolution of lesions in groups 5 and 6 did not differ from one another but differed from the other groups. The lesions of group 5 increased 3.16 times every measurement, while those of group 6 increased 1.83 times, indicating that the smallest growth of the lesions occurred when the combination of therapies were used. A rabbit from group 5 showed clinical cure at day 20 of treatment. This research is the pioneer in the treatment of experimental pythiosis using essential oils from medicinal plants and a combination of therapies. This study demonstrated that the use of essential oils can be a viable alternative treatment to cutaneous pythiosis, particularly when used in association or combination with immunotherapy.

In vitro activity of antifungals in combination with essential oils against the oomycete Pythium insidiosum

The aim of this study was to investigate the in vitro susceptibility of Pythium insidiosum to combinations of the antifungal drugs terbinafine or itraconazole with Melaleuca alternifolia, Mentha piperita and Origanum vulgare essential oils (EOs).

Trichoderma virens as a biocontrol of Toxocara canis: In vivo evaluation

BACKGROUND Microorganisms have been widely studied as biological control agents of parasites of medical and veterinary importance. Coprophagous arthropods, bacteria and fungi are among the different organisms evaluated as potential biological control agents. Nematophagous fungi capture and digest the free forms of nematodes in the soil. Due to its zoonotic potential, Toxocara canis have been brought to the attention of researchers. AIMS The aim of the present study was to determine whether the administration of embryonated T. canis eggs exposed to the nematophagous fungus Trichoderma virens reduces parasite infection in experimental animals. METHODS Embryonated T. canis eggs were exposed to T. virens mycelium for 15 days at 25°C. Subsequently, 100 fungus-exposed eggs were orally administered to 20 Swiss mice. As a positive control, another 20 mice received 100 embryonated eggs that were not exposed to the fungus. After 48h, the animals were killed, and heart, lungs and liver were harvested for the recovery of larvae. RESULTS The organs of the animals that received embryonated T. canis eggs exposed to the fungus showed a lower mean larval recovery when compared with the animals that received embryonated eggs without fungus exposure (p<0.05). CONCLUSIONS The exposure of T. canis eggs to T. virens reduces the experimental infection, demonstrating the potential of this nematophagous fungus as a biocontrol agent.

Effect of Paecilomyces lilacinus, Trichoderma harzianum and Trichoderma virens fungal extracts on the hatchability of Ancylostoma eggs.

BACKGROUND Ancylostoma species have demanded attention due to their zoonotic potential. The use of anthelmintics is the usual method to prevent environmental contamination by Ancylostoma eggs and larvae. Nematophagous fungi have been widely used in their biological control due to the fungus ability to capture and digest free nematode forms. AIMS The aim of this study was to evaluate the effect of four different fungal extracts of Paecilomyces lilacinus (n=2), Trichoderma harzianum (n=1) and Trichoderma virens (n=1) isolates on the hatchability of Ancylostoma eggs. METHODS Fungal extracts consisted of fungal broth culture supernatant without filtration (crude extract) and filtered broth (filtered extract), macerated mycelium (crude macerate), and macerated mycelium submitted to filtration (filtered macerate). The Ancylostoma eggs were obtained from the feces of naturally infected dogs. In vitro assays were performed in five replicates and consisted of four treatments and one control group. RESULTS The activity of the fungal extracts of each evaluated fungus differed (p<0.05) from those of the control group, showing significant ovicidal activity. The hatching of the eggs suffered reduction percentages of 68.43% and 47.05% with P. lilacinus, and 56.43% with T. harzianum, when crude macerate extract was used. The reduction with the macerate extract of T. virens was slightly lower (52.25%) than that for the filtered macerate (53.64%). CONCLUSIONS The results showed that all extracts were effective in reducing the hatchability of Ancylostoma eggs. The ovicidal effect observed is likely to have been caused by the action of hydrolytic enzymes secreted by the fungi.