Duddingtonia flagrans and its crude proteolytic extract: concomitant action in sheep coprocultures.

The presence of infective larvae (L3) of gastrointestinal nematode (GIN) parasites in pastures directly contributes to the constant recurrence of infections in ruminant herds. This study aimed to evaluate the nematophagous fungus Duddingtonia flagrans (AC001) (proteolytic crude extract and/or conidia) in the in vitro control of GIN L3 in coprocultures. To produce the proteolytic crude extract, a suspension (107 conidia/mL) of D. flagrans was inoculated into a liquid medium. After 6 days, the medium was filtered, centrifuged, and its proteolytic activity was measured. For the experimental assay, fecal samples were collected directly from the rectal ampulla of naturally infected sheep, and egg counts per gram of feces (EPG) were performed. Coprocultures were prepared using 10 g of fecal material with the groups defined as follows: control group G1 (1.0 mL of denatured proteolytic crude extract); treated group G2 (1.0 mL of active proteolytic crude extract); treated group G3 (1.0 mL of active proteolytic crude extract + 1.0 mL of AC001 conidia). The coprocultures were maintained at room temperature (25ºC), for 7 days, and then the L3 larvae were recovered. The results demonstrated that AC001 successfully produced protease (56.34 U/mL). The treatments with active proteolytic crude extract (G2) and active proteolytic crude extract + AC001 conidia (G3) were significantly different (p < 0.01) from the control group with denatured proteolytic crude extract (G1). AC001 and its proteolytic crude extract acted concomitantly on helminths directly in the fecal environment, suggesting potential future applications in the field.

Compatibility study of Duddingtonia flagrans conidia and its crude proteolytic extract.

This study aimed to evaluate the concomitant use of the nematophagous fungus Duddingtonia flagrans (AC001) and its protease-rich crude extract for the in vitro control of Panagrellus sp., Haemonchus spp., and Trichostrongylus spp. The nematicidal tests were carried out on larvae of the free-living nematode Panagrellus sp. and infective larvae of the gastrointestinal parasitic nematodes of domestic ruminants (Haemonchus spp. and Trichostrongylus spp). Five experimental groups were set: (1) one control group (G1) and (4) four treated groups -G2 - active crude extract; G3 - denatured crude extract; G4 - fungus, and G5 - fungus + active extract. Plates were incubated at 28 ºC for 24 h followed by the recovery of the larvae using the Baermann technique. The results showed a lower recovery of Panagrellus sp. larvae in the experimental groups compared to the control group, as follows: 52 % (G2), 16 % (G3), 46 % (G4), and 77 % (G5). An even greater reduction (77 ± 5 %) occurred in the group (G5). In addition, the authors observed lower averages of L3 of Haemonchus spp. and Trichostrongylus spp. in the experimental groups compared to the control group, as follows: 59 % (G2), 0 % (G3), 86 % (G4), and 76 % (G5). In turn, there was a difference (p < 0.01) between (G5) and (G2). The results this study indicate a positive effect from the compatible use of the D. flagrans fungus and its enzymatic crude extract (protease), which has been demonstrated here for the first time and with potential field applications for further designs.

Biological activity of cinnamaldehyde, citronellal, geraniol and anacardic acid on Haemonchus contortus isolates susceptible and resistant to synthetic anthelmintics.

Parasitism by gastrointestinal nematodes is a challenge for small ruminant farming worldwide. It causes productive and economic losses, especially due to parasite resistance to conventional anthelmintics. Natural compounds with antiparasitic activity are a potential alternative for controlling these parasites especially when considering the widespread occurrence of anthelmintic resistance. Our objective was to evaluate the activity of anacardic acid, geraniol, cinnamaldehyde and citronellal on Haemonchus contortus isolates with different levels of anthelmintic resistance profiles. These compounds were tested using egg hatch assays (EHAs), larval development tests (LDTs) as well as LDTs on mini-fecal cultures, on the Haemonchus contortus isolates Kokstad (KOK-resistant to all anthelmintics), Inbred-Strain-Edinburgh (ISE-susceptible to all anthelmintics) and Echevarria (ECH-susceptible to all anthelmintics). Effective concentrations to inhibit 50% (EC50) and 95% (EC95) of egg hatching and larval development were calculated. Results for EHA and LDT for all tested compounds, considering EC50 and EC95 values, showed low variation among the studied isolates with most RF values below 2x. All studied compounds showed efficacy against egg hatching and larval development of H. contortus isolates regardless of anthelmintic resistance profiles. The compounds with the smallest EC50 and EC95 values were cinnamaldehyde and anacardic acid making them promising candidates for future in vivo studies.