Braulio Valles de la Mora

In vitro anthelmintic activity of five tropical legumes on the exsheathment and motility of Haemonchus contortus infective larvae

This study investigated the in vitro anthelmintic (AH) activity of five tropical legume plants [Arachis pintoi CIAT 22160 (A.p. 22160), Gliricidia sepium, Cratylia argentea (C.a. Yacapani), C. argentea CIAT 22386 (C.a. 22386), C. argentea Veranera (C.a. Veranera)] against Haemonchus contortus infective larvae and the role of tannins/polyphenolic compounds in the AH effect. Lyophilized leaf extracts of each plant were evaluated using the Larval Exsheathment Inhibition Assay (LEIA) and the larval migration inhibition assay (LMIA). The role of tannins/polyphenolic compounds in the AH effect was evaluated in both assays using polyethylene glycol (PEG) to remove tannins from the solutions. At the highest concentration (1200μg of extract/ml), A. pintoi 22160, C.a. Yacapani, C.a. Veranera and C.a. 22386 completely inhibited the exsheathment process of H. contortus (P<0.01). At the same concentration (1200μg of extract/ml), the inhibition of larval migration for C.a. 22386, C.a. Veranera and G. sepium was 66.0%, 35.9% and 39.2% (relative to the PBS control), respectively. In both bioassays (LEIA and LMIA), the AH effect shown by each plant was blocked after the addition of polyethylene glycol (PEG), corroborating the role of tannins/polyphenolic compounds.

Elucidation of Leucaena leucocephala anthelmintic-like phytochemicals and the ultrastructural damage generated to eggs of Cooperia spp.

Leucaena leucocephala is a tropical forage legume suggested as an alternative method to control gastrointestinal parasitism in ruminants. This study: (1) performed a bio-guided fractionation of an aqueous extract of L. leucocephala using the egg hatch assay (EHA) to identify the anthelmintic (AH)-like phytochemicals present in fresh leaves, and (2) assessed the ultrastructural damage to eggs of Cooperia spp. after incubation with the final fraction. Phytochemicals were isolated using silica gel columns and identified using high performance liquid chromatography and standards for comparison. The final fraction was evaluated using EHA at 0.06, 0.125, 0.250, 0.500 and 1.1 mg ml(-1). The lethal concentration to inhibit 50% of Cooperia spp. egg hatching (LC50) was calculated using a Probit analysis. Scanning and transmission electron microscopy revealed the ultrastructural changes present in Cooperia spp. eggs. Bio-guided isolation procedures led to the recognition of an active fraction (LlC1F3) mainly composed of quercetin (82.21%) and caffeic acid (13.42%) which inhibited 90.49 ± 2.8% of Cooperia spp. egg hatching (P<0.05), and an LC50 of 0.06 ± 0.14 mg ml(-1). Scanning electron microscopy (SEM) showed eggs exposed to the active fraction had an irregular external layer with small projections and ruptures of lateral eggshell walls. Transmission electron microscopy (TEM) showed changes to Cooperia spp. eggs in electro-density, including the thickness of the eggshell layers and fractures after incubation with the final fraction (LlC1F3). Changes in bioactivity after purification suggest synergistic interactions between quercetin and caffeic acid.

Cratylia argentea: un arbusto forrajero potencial en sistemas silvopastoriles: Rendimiento y calidad de accesiones según las edades de rebrote y estaciones climáticas

In silvopastoral systems, high nutritional quality shrub legumes represent an alternative to shortage of grasses in dry periods of the year. The effect of different regrowth ages (6, 9, 12 and 15 weeks) in three climatic seasons (rainy, winter and dry seasons, 2007-2008) of four Cratylia argentea accessions (18516, 18666, 18668 and 18676) were evaluated in Veracruz, Mexico, for dry matter yield (DMY), neutral detergent fiber (NDF), acid detergent fiber (ADF), lignin (Lig), crude protein (CP) and in situ DM degradation (ISDMD). Dry matter yield (mean ± standard error) was affected by season and regrowth age, but not by accession or interactions. The dry season showed the largest DMY (3632 ± 306 kg·ha-1), representing 45 % of total forage yield, followed by the rainy (33 %, 2615±188 kg·ha-1) and winter (22 %; 1733 ± 61 kg·ha-1) seasons. For NDF, ADF, Lig and CP, statistical effects were found for regrowth age in all three seasons. The largest ISDMD occurred in the rainy and dry seasons, reaching average values of 66 and 65 % at 72 h of incubation, respectively. The dry season was the best period for DMY and ISDMD, making Cratylia argentea a reliable feed resource for silvopastoral systems. Recibido: 22 de octubre, 2013 Aceptado: 26 de junio, 2014 doi: 10.5154/r.rchscfa.2013.11.040 http://www.chapingo.mx/revistas PALABRAS CLAVE: Rendimiento de materia seca, composición química, efectos estacionales, leguminosas forrajeras arbustivas, Veracruz, México.

Effect of Gliricidia sepium leaves intake on larval establishment of Cooperia punctata in calves and bio-guided fractionation of bioactive molecules

The aims of this study were: 1) to assess the anthelmintic effect of Gliricidia sepium on the establishment of C. punctata third-stage larvae (L3) in calves, and 2) to isolate and to elucidate an anti-exsheathment phytochemical from the plant offered during the trial. Twelve ¾ Holstein × Zebu calves were divided in two experimental groups: control (T1) and treatment (T2) (n = 6). After adaptation, each calf was infected with an oral dose of 400 C. punctata L3/Kg LW. Basal diet consisted of Digitaria decumbens hay (6.27% CP) and commercial concentrate (12% CP). In addition, during the experimental period T2 received fresh G. sepium leaves (26.88% CP) ad libitum. On day 9 post-infection, three calves per treatment were randomly selected for slaughter, and worm counts were performed. Larval establishment rates obtained were 13.44 ± 0.13% and 3.1 ± 1.42% for T1 and T2, respectively (P < .05). The reduction of larval establishment was 76.9%. The total length of worms recovered from the animals was also affected by the intake of G. sepium (P < .05). Phytochemicals present in G. sepium leaves offered to calves were isolated through silica gel columns and elucidated through Magnetic Nuclear Resonance (1H and 13C). Bio-guided isolation procedures lead to the elucidation of Oxytroside (Kaempferol 3-O-rhamnopyranosyl-(1 → 6)-β-D-glucopyranoside-7-O-rhamnopyranoside), which fully inhibited the C. punctata exsheathment process (2400 μg mL-1). Gliricidia sepium represents an alternative to prevent severe C. punctata infections by reducing larval establishment in cattle.

Reappearance of Mecistocirrus digitatus in cattle from the Mexican tropics: prevalence, molecular, and scanning electron microscopy identification.

Abstract:  Mecistocirrus digitatus is a hematophagous abomasal nematode which causes significant blood and production losses in cattle. The objectives of the present study were to: (1) report the reappearance of M. digitatus in cattle from the Mexican tropics using microscopy, scanning electron microscopy (SEM), and molecular identification, and (2) determine the prevalence of M. digitatus in slaughtered adult cattle from the Mexican tropics. Gastroinestinal nematodes (GIN) were recovered from the abomasum of an 8-yr-old cow (Holstein × Zebu) previously diagnosed with Johnes disease. Of 1,254 GIN, 98.8% were identified as M. digitatus and 1.2% as Haemonchus sp. SEM was used to identify ultrastructure features of M. digitatus (oral cavity, cervical papillae, bursa, bursa lobes papillae, male spicules, anus, and female vulva). A conventional PCR method was used to corroborate the morphological findings. The prevalence of adult cattle infected with M. digitatus and Haemonchus sp., determined from 68 adult cattle from different grazing tropical herds, was 38.2% and 8.8%, respectively. Ninety-eight percent of animals infected with M. digitatus presented lesions in their abomasum such as mucosal inflammation, hemorrhage, and ulcers; some of them had necrosis. The current reappearance of M. digitatus in a Mexican herd suggests the possibility of an underestimated prevalence of this nematode amongst grazing cattle.