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The aim of the study was to examine the variation in the in vitro susceptibility of ten Haemonchus contortus isolates from different geographical origins using respective egg hatch assays (EHA) with acetone:water extracts of two tannin containing plants, chimay (Acacia pennatula) and sainfoin (Onobrychis viciifolia). Fresh eggs were incubated in PBS with different concentrations of each extract (0, 600, 1200, 2400, 3600, 5000 and 8000 μg/ml PBS). Additional concentrations were tested for O. viciifolia (75, 100, 200 and 400 μg/ml PBS). Effective concentrations 50% (EC50), with the corresponding 95% confidence interval (95% CI), were calculated for every isolate with both extracts. Moreover, a resistance ratio (RR) was calculated to compare the isolates, using the most susceptible isolate for each extract as the respective reference. A second set of incubations were made using polyvinylpolypyrrolidone (PVPP) (0, 5000 μg/ml, 5000 μg/ml+PVPP) to determine the influence of polyphenols on the AH effect. The proportion of morulated eggs, eggs with L1 larvae failing eclosion (%LFE), and emerged larvae were estimated at different extract concentrations. Data of each isolate was used to calculate the effective concentration 50% (EC50) for each extract. The EC50 of each isolate was used to determine resistance ratio (RR) for the different isolates. For the 2 extracts, a susceptibility variation in egg hatching was observed for the different H. contortus isolates. The EC50 values for A. pennatula ranged from 2203 to 14106 μg (RR from 2.01 to 6.40). The O. viciifolia extract showed higher variability with EC50 values ranging from 104 to 4783 μg (RR from 3.66 to 45.74). The main AH effects of the two extracts tested on the ten isolates consisted in blocking the emergence of L1 larvae (higher% LFE). Additional observations on emerged larvae showed that extract exposure caused alterations in the internal structure, separating the cuticle from the pharynx, bulb and intestinal cells. The use of PVPP revealed that (a) condensed tannins were not the sole plant secondary metabolites responsible for the AH effects, and (b) different H. contortus isolates showed variability in the role of tannins either on the ovicidal effect or the %LFE.

Use of plants containing bioactive compounds to control of helminths in the gastrointestinal tract, either as phytotherapeutic or nutraceutical options, has been a growing research area in recent years. We discuss strategies to identify viable candidate compounds with in vitro and in vivo anthelmintic properties. We also discuss factors which may influence in vitro and in vivo results, and difficulties of translating in vitro results to in vivo conditions are considered using experiences with small ruminants, as most published research on phytotherapeutic or nutraceutical materials has been in sheep and goats and has been reviewed recently. Therefore, we summarize results of various plant bioactive materials against helminth parasites in the gastrointestinal tract of cattle, deer, rabbits, pigs and poultry, and conclude that many plant materials have resulted in promising results in many farm animal species besides sheep and goats. These bioactive materials may be used as a part of sustainable helminth control strategies.

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This study assessed the in vitro anthelmintic (AH) activity of methanol and acetone:water leaf extracts from Annona squamosa, A. muricata and A. reticulata against Haemonchus contortus eggs. The egg hatch test was used to determine the effective concentrations required to inhibit 50% of eggs hatching (EC50). The role of polyphenols on AH activity was measured through bioassays with and without polyvinylpolypyrrolidone (PVPP). Methanolic extracts mainly caused the death of eggs at the morula stage (ovicidal activity). Meanwhile, acetone:water extracts caused egg-hatching failure of developed larvae (larvae failing eclosion (LFE) activity). The lowest EC50 values against H. contortus eggs were observed for the methanolic extracts from A. reticulata and A. muricata (274.2 and 382.9 µg/ml, respectively). From the six extracts evaluated, the methanolic extracts of A. muricata, A. reticulata and A. squamosa showed the highest ovicidal activity, resulting in 98.9%, 92.8% and 95.1% egg mortality, respectively. When the methanolic extract of A. squamosa was incubated with PVPP, its AH activity increased. Similarly, when acetone:water extracts of A. muriata and A. reticulata were incubated with PVPP, their LFE activity increased. Alkaloids were only evident in methanolic extracts, irrespective of PVPP incubation. The presence of acetogenins was not observed. In conclusion, methanolic extracts obtained from leaves of A. muricata, A. reticulata and A. squamosa showed ovicidal activity affecting the morula of H. contortus eggs, with minor LFE activity. Meanwhile, acetone:water extracts showed mostly LFE activity, with a lower proportion of ovicidal activity.

This study explored the variation in susceptibility to acetone:water plant extracts between infective larvae (L) of ten Haemonchus contortus isolates from different geographical origin. The L of 10 different isolates were exposed either to the acetone:water extract of a temperate plant (Onobrychis viciifolia) or a tropical plant (Acacia pennatula) and were evaluated with the larval exsheathment inhibition assay (LEIA). The L of each isolate were incubated with different concentrations of each extract (0, 25, 50, 100, 200, 400, 600, 800, 1000 and 1200μg/mL of phosphate buffered saline (PBS)). After incubation, the exsheathment process of L was induced using a solution with sodium hypochlorite (2%) and sodium chloride (16.5%). The proportion of exsheathed L was determined for each concentration at 0, 20, 40 and 60min. Effective concentrations 50% (EC) and the corresponding 95% confidence intervals (95% CI) were calculated for every isolate with both extracts. Moreover, a resistance ratio (RR) was calculated for each extract to compare isolates, using the most susceptible isolate as the respective reference for each extract. To determine the role of polyphenols on the reported effect, a second set of incubations was made for each isolate and each extract, using the extracts at a concentration of 1200μg/mL PBS with or without polyvinylpolypyrrolidone (PVPP), a polyphenol blocking agent, and controls without extract. The ten different H. contortus isolates showed variation in susceptibility for each of the 2 extracts tested (P<0.05). The EC values for A. pennatula extract ranged from 36 to 501μg/mL (RR: 2.11-13.68). Meanwhile, the EC values for O. viciifolia extract ranged from 128 to 1003μg/mL (RR: 1.25-7.82). The use of PVPP revealed that polyphenols were responsible for the anthelmintic activity recorded for both extracts. However, tested H. contortus isolates suggested that susceptibility to one polyphenol-rich extract did not determine the susceptibility to the other polyphenol rich extract. The latter result indicated that the different H. contortus isolates varied in their susceptibility to the polyphenols present in each extract evaluated.

The protozoan parasite Trypanosoma cruzi is the causative agent of the Chagas disease, which is endemic in southeastern Mexico and is transmitted by the vector Triatoma dimidiata (triatomide). T. cruzi infect a great variety of domestic and wild mammals; rodents are considered one of the most important reservoirs of the parasite in the transmission cycles of T. cruzi. The objective of this study was to determine the frequency of T. cruzi infection and to determine the parasitic load in synanthropic and wild rodents from the rural community of southern Mexico. A total of 41 blood samples and 68 heart tissue samples were collected from various species of synanthropic (n= 48 in 2 species) and wild rodents (n= 35 in 5 species). DNA was extracted from samples to detect the presence of T. cruzi through quantitative PCR (qPCR). T. cruzi DNA was detected in the 9.75% of the blood samples of the synanthropic species (4/41) (14.28%) for Rattus rattus samples and 25% for Ototylomys phyllotis samples, with an average of parasitic load of 4.80 ± 1.17 parasites/μL. In the case of heart tissue samples, 10.29% were positive for T. cruzi (7/68) (8.7% for Rattus rattus, 40% for Peromyscus yucatanicus, and 42.8% for Ototylomys phyllotis) with an average parasite load of 3.15 ± 1.98 eq-parasites/mg. The active and chronic infection of T. cruzi in synanthropic or wild rodents of the rural community of southern Mexico evidences the natural infection in these reservoirs which contribute to maintaining the agent in the wild and domestic environments and can represent a risk of infection for the human population when the vector is present.