Luiza Neme Frassy

Destruição de ovos de Toxocara canis pelo fungo nematófago Pochonia chlamydosporia

INTRODUCTION: Toxocara canis is an ascarid parasite of the small intestine of dogs that causes visceral larva migrans in humans. METHODS: With the aim of demonstrating the effectiveness of the fungus Pochonia chlamydosporia on Toxocara canis eggs under laboratory conditions, a trial was set up in Petri dishes with 2% agar-water. RESULTS: There was ovicidal activity of 43.8% (p < 0.01) in the treated group in relation to the control group over the periods studied. CONCLUSIONS: The results from the present study suggest that Pochonia chlamydosporia can potentially be used as an alternative biological control for embryonated Toxocara canis eggs.

In vitro ovicidal activity of the nematophagous fungi Duddingtonia flagrans, Monacrosporium thaumasium and Pochonia chlamydosporia on Trichuris vulpis eggs

The in vitro effect of four isolates of the nematophagous fungi Duddingtonia flagrans (AC001), Monacrosporium thaumasium (NF34a) and Pochonia chlamydosporia (VC1 and VC4) on the eggs of Trichuris vulpis was evaluated. One thousand eggs of T. vulpis were plated on Petri dishes with 2% water-agar with the fungal isolates grown and without fungus as control. After 7, 14 and 21 days 100 eggs were removed from each plate and classified according to the following parameters: type 1, lytic effect without morphological damage to eggshell; type 2, lytic effect with morphological alteration of embryo and eggshell; and type 3, lytic effect with morphological alteration of embryo and eggshell, besides hyphal penetration and internal egg colonization. P. chlamydosporia demonstrated ovicidal activity (p<0.05) on the eggs of T. vulpis in the studied intervals presenting type 3 effect of 29.5% (VC1) and 36.5% (VC4), 59.5% (VC1) and 2.5% (VC4), 94.8% (VC1) and 2.95% (VC4) at 7, 14 and 21 days, respectively. The other fungi showed no type 3 effect. P. chlamydosporia should be a potential biological control agent of T. vulpis eggs.

In vitro predatory activity of the fungi Duddingtonia flagrans, Monacrosporium thaumasium, Monacrosporium sinense and Arthrobotrys robusta on Ancylostoma ceylanicum third-stage larvae

The potential role of companion animals as reservoirs for zoonotic diseases has been recognised as a significant public health problem worldwide. Ancylostoma ceylanicum is the only ancylostomatidae species known for infecting human beings. This article aimed to compare the predatory capacity of predatory fungi isolates Duddingtonia flagrans (AC001), Monacrosporium thaumasium (NF34), Monacrosporium sinense (SF53) and Arthrobotrys robusta (I31) on A. ceylanicum infectious larvae (L(3)) in a 2% water-agar plate. There was no predatory capacity variation among the fungi tested (P>0.05) over the 7-day period experimental assay. When compared to the control (without fungi), there was a significant reduction (P<0.05) of 95.6%, 85.1%, 87.4% and 90.2% on the A. ceylanicum L(3) mean recovered from treatments with isolates AC001, NF34, SF53 and I31, respectively. Regarding linear regression coefficients, negative values were noted for treatments, therefore indicating A. ceylanicum non-predated larvae reduction over 7 days. In this work, all predatory fungi isolates were efficient at capturing and destroying in vitro the A. ceylanicum L(3); therefore being able to be used as biological controllers of such nematode.

Pochonia chlamydosporia fungal activity in a solid medium and its crude extract against eggs of Ascaridia galli.

The present study aimed to evaluate the ovicidal activity (type 3 effect) of VC1 and VC4 isolates of Pochonia chlamydosporia in a solid medium and the action of a crude extract of P. chlamydosporia against eggs of Ascaridia galli. To evaluate ovicidal activity in culture medium, 1000 A. galli eggs were plated on Petri dishes containing 2% water-agar with grown fungal isolates (VC1 or VC4) and without fungus (control group) and were examined at 1, 3 and 5 days post-inoculation (assay A). Then, to test the action of crude extracts of P. chlamydosporia (VC1 or VC4), 500 eggs of A. galli were plated on Petri dishes of 4.5 cm diameter with 5 ml of fungal filtrate from each tested isolate. The control group consisted of 500 eggs of A. galli with 10 ml of distilled water on each Petri dish (assay B). Fungal isolates were effective (P < 0.01) at destroying these eggs, showing a type 3 effect at the studied intervals. On the other hand, the crude extract of isolates (VC1 or VC4) reduced the number of A. galli eggs in the treated group compared with the control group by 64.1% and 56.5%, respectively. The results of the present study show that P. chlamydosporia is effective at destroying eggs of A. galli and could therefore be used in the biological control of nematodes.

Influence of the preservation period in silica-gel on the predatory activity of the isolates of Duddingtonia flagrans on infective larvae of cyathostomins (Nematoda: Cyathostominae)

The continued maintenance of nematophagous fungi predatory activity under laboratory conditions is one of the basic requirements for a successful biological control. The purpose of this study was to evaluate the influence of time on the preservation of the fungus Duddingtonia flagrans (AC001 and CG722) stored in silica-gel for 7 years and their subsequent predatory activity on cyathostomin L(3) larvae in 2% water-agar medium (2% WA). Samples of the isolates AC001 and CG722, originating from vials containing grains of silica-gel sterilized and stored for 7 years, were used. After obtaining fungal conidia, the predation test was conducted over 7 days on the surface of 9.0 cm Petri dishes filled with 2% WA. In the treated groups each Petri dish contained 500 cyathostomin L(3) and conidia of fungal isolates in 2% WA. In the control group (without fungi) the plates contained 500 L(3) in 2% WA. The experimental results showed that isolated AC001 and CG722 were efficient in preying on cyathostomin L(3) (p<0.01) compared to control (without fungus). However, no difference was observed (p>0.01) in the predatory activity of the fungal isolates tested. Comparing the groups, there was a significant reductions of cyathostomin L(3) (p<0.01) of 88.6% and 78.4% on average recovered from the groups treated with the isolates AC001 and CG722, respectively, after 7 days. The results of this test showed that the fungus D. flagrans (AC001 and CG722) stored in silica-gel for at least 7 years maintained its predatory activity on cyathostomin L(3).

Atividade predatória dos fungos nematófagos Duddingtonia flagrans, Monacrosporium thaumasium e Artrobotrys robusta sobre larvas infectantes de Strongyloides stercoralis

INTRODUCTION: Strongyloides stercoralis is a nematode that infects much of the population worldwide. METHODS: This study aimed to compare the ability of predatory nematophagous fungi Duddingtonia flagrans (AC001), Monacrosporium thaumasium (NF34) and Arthrobotrys robusta (I-31) on infective larvae (L3) of Strongyloides stercoralis in laboratory conditions on 2% water-agar. RESULTS: At the end of the experiment, the percentage reductions in Strongyloides stercoralis L3 were 83.7% (AC001), 75.5% (NF34) and 73.2% (I-31). CONCLUSIONS: The nematophagous fungi were able to capture and destroy the L3 in vitro and may be used as biological controls of Strongyloides stercoralis.

Duddingtonia flagrans, Monacrosporium thaumasium and Pochonia chlamydosporia as possible biological control agents of Oxyuris equi and Austroxyuris finlaysoni.

The action of four fungal isolates of the species Duddingtonia flagrans (AC001), Monacrosporium thaumasium (NF34a) and Pochonia chlamydosporia (VC1 and VC4) on eggs of Oxyuris equi and Austroxyuris finlaysoni was evaluated in two assays (A and B). Eggs of O. equi (Test A) and A. finlaysoni (Test B) were plated on Petri dishes with 2% water-agar with grown fungal isolates and control without fungus. After 5, 10 and 15 days, 100 eggs were collected and classified according to the following parameters: type 1 effect, physiological and biochemical effect without morphological damage to the eggshell; type 2 effect, lytic effect with morphological alteration of the eggshell and embryo; and type 3 effect, lytic effect with morphological alteration of the eggshell and embryo, hyphal penetration and internal egg colonization. Pochonia chlamydosporia isolates VC1 and VC4 showed ovicidal activity for type 1, 2 and 3 effects on eggs of O. equi and eggs of A. finlaysoni. In vitro assays A and B showed that P. chlamydosporia had a negative influence on eggs of O. equi and A. finlaysoni and can be considered as a potential biological control agent of nematodes.

Production and partial characterization of Duddingtonia flagrans (AC001) crude extract and its in vitro larvicidal action against trichostrongylid infective larvae

Abstract The production and partial characterization of Duddingtonia flagrans (AC001) crude extract and its in vitro larvicidal action against trichostrongylid infective larvae from sheep were studied. D. flagrans was grown in liquid medium with glucose, casein, bibasic potassium phosphate (K2HPO4), magnesium sulfate (MgSO4), zinc sulfate (ZnSO4), ferrous sulfate (FeSO4), and copper sulfate (CuSO4). The proteolytic activity was measured within varied pHs and temperatures. To determine the thermostability, the crude extract was incubated at 28°C for 72 h. To study the effect of different chemical compounds on the activity of the crude extract, the samples were incubated in solutions containing (10 mM): calcium chloride (CaCl2), copper II sulfate (CuSO4), zinc sulfate (ZnSO4), magnesium sulfate (MgSO4), inhibitor phenylmethylsulfonyl fluoride (PMSF), and 0.5% SDS. Results showed that the highest activity obtained (79.23 U/mL) was at pH 9.0, while the optimum temperature was 60°C (119.6 U/mL). The thermostability analysis demonstrated that after 72 h the activity was maintained or increased. It was found that the CuSO4, ZnSO4, and PMSF strongly inhibited the proteolytic activity. Moreover, the MgSO4 and SDS, caused a weak inhibition of the proteolytic activity. There was a significant (P<0.01) reduction in number of treated L3 when compared to control (94.2%). The results suggest that the crude extract produced by D. flagrans (AC001) in liquid medium exerted larvicidal activity on trichostrongilid L3 and therefore may contribute to a large-scale industrial production.

In vitro predatory activity of conidia of fungal isolates of the Duddingtonia flagrans on Angiostrongylus vasorum first-stage larvae

INTRODUCTION Angiostrongylus vasorum is a nematode that parasitizes molluscs, dogs, and even man. METHODS The objective was to evaluate the predatory activity of the conidia of two fungal isolates of Duddingtonia flagrans (AC001 and CG722) on first-stage larvae (L1) of A. vasorum in laboratory conditions. RESULTS At the end of the experiment, there were significant reductions (p<0.01) of 74.5% and 63.2%, on average, in the A. vasorum L1 recovered in the AC001 and CG722 treatment conditions, respectively. CONCLUSIONS The two isolates of fungi were efficient in the capture and destruction of A. vasorum L1.

Activity in vitro of fungal conidia of Duddingtonia flagrans and Monacrosporium thaumasium on Haemonchus contortus infective larvae

The objective of this work was to evaluate the predatory activity of the fungi Duddingtonia flagrans (AC001) and Monacrosporium thaumasium (NF34a) on Haemonchus contortus infective larvae (L3) in two experimental assays (A and B). In assay A, two treatments and one control were formed and kept for 7 days in Petri dishes with 2% water-agar. Each treatment consisted of 1000 H. contortus L3 and 1000 conidia of only one fungal isolate, and the control group consisted of 1000 L3, without fungus, with 10 repetitions per group. In assay B, 1000 conidia of one of the fungal isolates, AC001 or NF34a, were added to coprocultures made from 20 g of faeces collected from sheep naturally infected with H. contortus. At the end of the experiment, the Baermann method was used to count the non-predated larvae of all Petri dishes from treatment and control groups. In assay A, no difference was observed (P>0.05) between the groups treated with AC001 and NF34a fungi. A difference was observed (P < 0.05) between the treated and control groups. The L3 reduction percentages at the end of the experiment were 87.75 and 85.57%, respectively, for the fungal isolates compared to the control group. In assay B, the reduction percentages for conidia of these isolates were 85.82 and 87.32%, respectively. The results obtained show that D. flagrans (AC001) and M. thaumasium (NF34a) were effective in the in vitro control of sheep H. contortus L3 and could be used in the biological control of this nematode.