Anthelmintic efficacy of an organic fraction from Guazuma ulmifolia leaves and evaluation of reactive oxidative stress on Haemonchus contortus.

This study describes the anthelmintic efficacy of an organic fraction (EtOAc-F) from Guazuma ulmifolia leaves and the evaluation of its reactive oxidative stress on Haemonchus contortus. The first step was to assess the anthelmintic effect of EtOAc-F at 0.0, 3.5, 7.0 and 14 mg kg of body weight (BW) in gerbil's (Meriones unguiculatus) artificially infected with H. contortus infective larvae (L3). The second step was to evaluate the preliminary toxicity after oral administration of the EtOAc-F in gerbils. Finally, the third step was to determine the relative expression of biomarkers such as glutathione (GPx), catalase (CAT), and superoxide dismutase (SOD) against H. contortus L3 post-exposition to EtOAc-F. Additionally, the less-polar compounds of EtOAc-F were identified by gas mass spectrophotometry (GC-MS). The highest anthelmintic efficacy (97.34%) of the organic fraction was found in the gerbils treated with the 14 mg/kg of BW. Histopathological analysis did not reveal changes in tissues. The relative expression reflects overexpression of GPx (p<0.05, fold change: 14.35) and over expression of SOD (p≤0.05, fold change: 0.18) in H. contortus L3 exposed to 97.44 mg/mL of EtOAc-F compared with negative control. The GC-MS analysis revealed the presence of 4-hydroxybenzaldehyde (1), leucoanthocyanidin derivative (2), coniferyl alcohol (3), ferulic acid methyl ester acetate (4), 2,3,4-trimethoxycinnamic acid (5) and epiyangambin (6) as major compounds. According to these results, the EtOAc-F from G. ulmifolia leaves exhibit anthelmintic effect and increased the stress biomarkers on H. contortus.

Haematological values in cattle reared in humid and subhumid tropics of Mexico.

The objective of this study was to determine some factors that influence the haematological values of cattle reared in the humid and subhumid tropics of Mexico. Whole blood samples were taken from 1355 crossbred cattle in the years 2017 to 2019. Haematocrit (HTC, %), total plasma protein (TPP, g/dL) and peripheral eosinophils count (PEOS, × 103/µL) were determined manually, and the main haematological variables were recorded with an automatic analyser. The statistical analysis considered as classification variables age, sex, season (cold, dry and rainy), year (2017, 2018 and 2019) and origin of the cattle. The mean of the haematological parameters was determined along with the confidence limits (CL) of the different categories of animals according to age. Calves younger than 1-year-old presented higher levels of HTC, red blood cell count (RBC), haemoglobin (HGB), red blood cell distribution width (RDW), platelet number (PLT), white blood cell count (WBC) and lymphocyte count (LYMF) than animals older than 2 years of age. However, they showed the lowest mean cell volume (MCV) and TPP values. In cows, the highest levels of PEOS, granulocytes (GRAN), MCV and medium cells (MID) were observed and the lowest HTC, RBC, RDW and WBC levels. Intervals were determined with the 1st quartile (Q1) or lower confidence interval (90% CI) as the minimum values and the 3rd quartile (Q3) or upper confidence interval (90% CI) as the maximum values. The haematological parameters of cattle reared in the Southeast of Mexico are significantly affected by age, sex and environmental conditions.

EVALUATION OF PELIBUEY LAMBS BORN TO MOTHERS PHENOTYPICALLY SEGREGATED ACCORDING TO RESISTANCE TO GASTROINTESTINAL NEMATODES IN THE HUMID TROPICS OF MEXICO.

The objective was to evaluate the effect of the resistance of ewes to gastrointestinal nematodes (GIN) on the parasitologic and productive performance of their lambs during preweaning and postweaning periods. Offspring from 44 Pelibuey ewes classified as resistant or susceptible to GIN were evaluated. During the preweaning period, birth weight (BW), live weight (LW) at 35 and 70 days of age, and live weight changes (LWC) were determined in 73 lambs. During the postweaning period, 20 lambs to susceptible or resistant ewes were evaluated. The lambs were challenged under natural GIN infections and artificial infection with L3 of Haemonchus contortus. The fecal egg count (FEC), hematocrit (HT), peripheral eosinophil count, plasma protein, LW, and LWC were determined. There were no differences among parameters evaluated during preweaning (P > 0.05). The type of lambing (single or multiple) affected the BW, LW, and LWC (P < 0.01). The sex (male) of the lamb had a positive effect on BW at 35 and 70 days of age (P < 0.05). After weaning, lambs of susceptible ewes showed higher FEC than resistant ewes (P < 0.05). The HT of lambs to resistant ewes was higher than those the susceptible group (P < 0.01). In conclusion, offspring of resistant ewes show low FEC compared with those susceptible during the postweaning stage.

In Vitro Anthelmintic Activity of a Hydroalcoholic Extract from Guazuma ulmifolia Leaves against Haemonchus contortus.

The purpose of the present study was to assess the ovicidal and larvicidal activity of a hydroalcoholic extract (HAE) and their fractions (aqueous, Aq-F and organic, EtOAc-F) from Guazuma ulmifolia leaves using Haemonchus contortus as a biological model. The egg hatching inhibition (EHI) and larval mortality against infective larvae (L3) tests were used to determine the anthelmintic effect of the treatments. The extract and fractions were tested at different concentrations against eggs and L3. Additionally, distilled water and methanol were used as negative controls and ivermectin as a positive control. The extract and fractions were subjected to HPLC analysis to identify the major compounds. The HAE displayed the highest ovicidal activity (100% EHI at 10 mg/mL). Fractionation of the HA extract allowed increasing the nematicidal effect in the EtOAc-F (100% EHI at 0.62 mg/mL and 85.35% mortality at 25 mg/mL). The phytochemical analysis of the extract and fractions revealed the presence of kaempferol, ethyl ferulate, ethyl coumarate, flavonol, luteolin, ferulic acid, luteolin rhamnoside, apigenin rutinoside, coumaric acid derivative, luteolin glucoside, and quercetin glucoside. These results suggest that G. ulmifolia leaves could be potential candidates for the control of H. contortus or other gastrointestinal parasitic nematodes.

Protein Supplementation as a Nutritional Strategy to Reduce Gastrointestinal Nematodiasis in Periparturient and Lactating Pelibuey Ewes in a Tropical Environment.

The study was carried out to evaluate the effect of energy and protein supplementation on parasitological and hematological response during peripartum and lactation of productive and non-productive Pelibuey ewes in a tropical environment. Forty-eight Pelibuey ewes aged 3-5 years and with a body weight of 31 ± 5 kg were used. Four groups of 12 ewes, including non-pregnant and productive ewes, were formed. A factorial treatment design was formulated, where two levels of energy (low, 9.6 MJ/kg, n = 24; and high, 10.1 MJ/kg, n = 24) and two levels of protein (high, 15% crude protein in diet, n = 24; and low, 8% crude protein in diet, n = 24) were studied. Fecal and blood samples were collected to determine the fecal egg count (FEC) of gastrointestinal nematodes (GIN), packed cell volume (PCV) and peripheral eosinophil (EOS) count. These variables were rearranged with respect to the lambing date in a retrospective study. The high dietary protein level had a significant effect on reducing the FEC and increasing the PCV of ewes during lactation, in comparison with animals fed with the low protein level. Differences in the study variables were attributed to physiological stage. Lactating ewes showed the highest FEC values (2709 ± 359 EPG), the lowest PCV values (21.9 ± 0.7%) and the lowest EOS (0.59 ± 0.6 Cells × 103 µL). It is concluded that high levels of dietary protein improve the hematological response and reduce the FEC in Pelibuey ewes under grazing conditions. The non-pregnant ewes maintained some resilience and resistance to GIN infection compared to productive ewes.

An Artemisia cina n-hexane extract reduces the Haemonchus contortus and Teladorsagia circumcincta fecal egg count in naturally infected periparturient goats.

The objective of this study was to evaluate an n-hexane extract of Artemisia cina (Acn-h) as a natural anthelmintic treatment for periparturient goats naturally infected with the nematodes Haemonchus contortus and Teladorsagia circumcincta. A total of 200 periparturient Alpine and Nubian goats were used. Deworming criteria were based on the following parameters: fecal egg account (epg), ocular mucosa color (OMC), and body condition (BC). A previous analysis using coprocultures of the flock revealed the presence of H. contortus (80%) and T. circumcincta (20%). The Acn-h contained two new compounds identified by mass spectrometry data as isoguaiacin and norisoguaiacin at 284.14 and 315 m/z. The lethal effects of Acn-h at 0.5, 1, 2, and 4 mg/mL on H. contortus adult stages were 31.6, 66.5, 81.3, and 86.9%, respectively (p < 0.05), showing similar efficacy at 2 and 4 mg/mL with albendazole (positive control group). Then, two experimental groups, with 100 goats in peripartum in each, were distributed randomly and treated at day 0 as follows: group 1 = 4 mg/kg of Acn-h as single oral dose, and group 2 = control group, treated with water (as a placebo). The epg, OMC, and BC parameters were recorded at 0 (periparturient period), 7 (birth period), and 23 (postpartum) days and analyzed using a completely randomized design with Duncan's test for comparison of means and analysis of variance. The following epg reductions were recorded in the Acn-h-treated group as follows: 20.1 ± 34.4 and 31.7 ± 38.2% at days 7 and 23 compared to the control group. During the whole experiment, no significant differences in OMC or BC were observed in relation to the control group, excepting at day 23 (p < 0.05) for BC in the group treated with A. cina. Thus, Acn-h can be a useful natural alternative tool for the control of the nematodes H. contortus and T. circumcincta in periparturient goat flocks.

In vitro anthelmintic activity of Pimienta dioica and Origanum vulgare essential oils on gastrointestinal nematodes from sheep and cattle.

The aim of the study was to evaluate the in vitro nematicide activity of the allspice (Pimienta dioica) and oregano (Origanum vulgare) essential oils (EOs) on eggs and larvae of gastrointestinal nematodes (GIN) from ruminant. The EOs were obtained by hydro-distillation of previously dehydrated plant material. In the in vitro evaluation, the egg hatching and larval mobility of Haemonchus contortus and Cooperia spp. of sheep and cattle, respectively, were determined. EOs at concentrations of 20.2-252.3 µg µL-1 with serial dilutions were used, also a negative control (distilled water + Tween 20) and Thiabendazol (77 µg µL-1) and Levamisole (120 µg µL-1) as a positive control to eggs and larvae, respectively, were placed on microplates. The results were analyzed with the PROBIT procedure. PPEO showed ovicidal effect in vitro test, with lethal doses 99 (LD99) of 0.56-4.19 µg µL-1, while oregano essential oil (OREO) was shown to be a larvicidal activity with doses of LD99 of 0.15-6.60  L-1. The results confirm that P. dioica and O. vulgare EOs have ovicidal and larvicidal capacity against GIN in vitro test.

High Predatory Capacity of a Novel Arthrobotrys oligospora Variety on the Ovine Gastrointestinal Nematode Haemonchus contortus (Rhabditomorpha: Trichostrongylidae).

With the worldwide development of anthelmintic resistance, new alternative approaches for controlling gastrointestinal nematodes in sheep are urgently required. In this work, we identified and characterized native nematode-trapping fungi. We collected seven isolates of fungi with the capacity to form adhesive, three-dimensional networks as the main mechanism to capture, kill, and consume nematodes. The nematode-trapping fungi were classified into two groups; the first group includes the R2-13 strain, showing faster growth, abundant aerial hyphae, scarce conidia production, bigger conidia, and it formed a clade with Arthrobotrys oligospora sensu stricto. The second comprises the A6, A12, A13, R2-1, R2-6, and R2-14 strains, showing a growth adhering to the culture medium, forming little aerial hyphae, smaller conidia, and these formed a sister clade to A. oligospora. Except for the R2-6 strain, conidia production was induced by light. In all the strains, the predatory capacity against the sheep gastrointestinal nematode Haemonchus contortus was greater than 58% compared with the control group. The A6 and A13 strains were the most active against the infective H. contortus third instar (L3) larvae, with an average capture capacity of 91%. Altogether, our results support evidence for a novel A. oligospora variety with high nematode-trapping activity and promissory in helminthic control.

Effect of an Arthrobotrys musiformis (Fungi: Orbiliales) culture filtrate on the population of gastrointestinal parasitic nematode eggs in faeces of grazing lambs.

This study assessed the anthelmintic activity of the oral administration of a free-spore culture filtrate of the nematophagous fungus (NF) Arthrobotrys musiformis (M-10) on gastrointestinal parasitic nematodes (GIN) in naturally infected lambs. The fungus was grown on potato-dextrose agar plates (PDA) and transferred to a fermented rice medium (FRM). After 40-day incubation the total amount of FRM with the growing fungi was transferred to a flask shaker with distilled water for a 24 h period. The fungus was centrifuged and filtered. Three groups of six naturally-infected lambs (>1000 epg) each were treated once as follows: Group 1) 63.8 mg/kg A. musiformis culture filtrate (CF) (per os); Group 2) Levamisole 7.5 mg/ml (intramuscularly), Group 3) 15 ml of distilled water (per os). Faecal samples were individually collected on days -2, 0, 2, 4, 6, 8 and 10 after treatment. For each experimental group, mean egg shedding was calculated and transformed (log 10 [epg + 1]). Means between the fungal filtrate group and the negative control were analysed using a T-Student Test. The faecal egg count reduction test (FECRT) was performed in groups treated with CF and Levamisole in relation to the control group (water) were 36.8-57.4% and 89-95.4%, respectively., although due to the difference between groups, no statistical significance was found (p > 0.05). The use of A. musiformis CF appears to be a good alternative treatment, although, more studies should be performed to establish the use of these fungal products as potential tools for GIN control.

Haemonchus contortus infection induces a variable immune response in resistant and susceptible Pelibuey sheep.

The immune response and phenotypic characteristics of Pelibuey lambs were analysed after the induction of a Haemonchus contortus trickle infection. Male lambs (n = 29; 20 kg live weight) were infected with 100 H. contortus infective larvae per kg of live weight on day 3, 5 and 7 of the experiment. The number of eggs per gram (epg), seven haematological parameters and the immunoglobulin A (IgA) level were analysed for 56 experimental days. In addition, histopathological samples from the fundic abomasal region and the relative expression of 10 immune-related genes from 15 infected and three non-infected lambs were analysed at day 0 and 49 of the experiment. The epg count and some haematological parameters (leucocytes, red blood cells, haemoglobin and total protein) with statistically significant differences (P < 0.01) were used to identify nine resistant and 20 susceptible lambs (1166 ±â€¯1071 and 3171 ±â€¯1463 epg, respectively). Moreover, acute infiltration of immune cells and parasitic granuloma formation were observed in susceptible lambs; the resistant group had moderate inflammatory cell infiltration. With respect to relative gene expression, resistant lambs showed upregulation (P < 0.001) of 10 genes, from 2.2 to 15.99 fold. Moreover, there was a strong indirect correlation (P < 0.05) between the epg count and interleukin 5 (IL5) gene expression. By contrast, there was an average 0.34 fold downregulation in nine of the immune-related genes (P ≤ 0.05) in susceptible lambs (the only exception was Fc fragment of IgE receptor Ia [FCER1A] upregulation). In addition, there was a direct correlation (P ≤ 0.05) between the epg count and the expression of IL8, which encodes an inflammatory chemokine. In conclusion, this study showed differential IL5 and IL8 gene expression during haemonchosis in resistant and susceptible Pelibuey lambs, respectively, together with a variable immune response based on histopathological and haematological parameters.

Prevalence and Factors Associated with Ectoparasite Infestations in Dogs from the State of Tabasco, Mexico.

This study was carried out to identify the ectoparasites that infest owned dogs in the state of Tabasco, Mexico. In total, 1,302 dogs were sampled in the 5 ecological regions of Tabasco. The dog owners were surveyed to identify the factors associated with infestations. Ectoparasites were identified using taxonomic keys. Eleven species of ectoparasites were observed. General prevalence was 26.65%. Rhipicephalus sanguineus and Ctenocephalides felis were the most prevalent and abundant ectoparasites. The most important factors associated with ectoparasite infestations in the studied dogs were living outdoors, being a non-purebred, having short hair, being dark-haired, and having a body condition <3. Ectoparasite studies such as the one presented herein generate important information to create control programs focused on decreasing infestations in companion animals and thus the likelihood of zoonotic transmission of pathogens.

Assessment of anthelmintic effectiveness to control Fasciola hepatica and paramphistome mixed infection in cattle in the humid tropics of Mexico.

The objective of this study was to evaluate the effectiveness of the main anthelmintics used for the control of trematodes in cattle in an endemic area in the humid Mexican tropics. A diagnostic study was carried out in nine cattle farms to determine the prevalence of flukes through faecal examination. Only three farms with more than 20 cows positive to trematodes were chosen to determine the effectiveness of commercial anthelmintics (triclabendazole, TCBZ; ivermectin + closantel, (IVM + CLOS); IVM + clorsulon, (CLORS); nitroxynil, NITROX). The prevalence of Fasciola hepatica was 27.1% and 29.6% of paramphistomes. The faecal egg count of trematodes ranged from 0.0 to 12.2 eggs per gram of faeces. The highest effectiveness against F. hepatica was 96.7%, and 92.7% against paramphistomes. NITROX was the most effective in the control of trematodes, while other products, such as IVM + CLORS and TCBZ obtained values lower than 90%, which puts sustainable trematode control at risk. The presence of trematodes was observed on most farms, although the prevalence per herd was highly variable, which indicates that the trematodes F. hepatica and paramphistomes are endemic to the region and a suitable management programme is suggested to control infections caused by these parasites.

Dirofilaria immitis proteins recognized by antibodies from individuals living with microfilaremic dogs.

INTRODUCTION: Dirofilaria immitis is a nematode that affects human health in several countries of the world. This study was conducted to examine whether serum samples from the owners of microfilaremic dogs present immunoreactivity to parasite proteins. METHODOLOGY: Eight serum samples from the owners of microfilaremic dogs were examined. Total proteins were extracted from adult worms and 12% SDS-PAGE was performed. The gel was electroblotted to a nitrocellulose membrane, and a Western blot (WB) was performed. Reactive bands of 22, 33, 39, 49, and 63 kDa in WB were excised from the gel and analyzed by mass spectrometry (MS). RESULTS: The MS results showed the presence of 10 different proteins of D. immitis recognized by the human serum samples. CONCLUSIONS: These results indicate that in endemic areas of D. immitis, owners of infected dogs recognize specific proteins of the parasite, suggesting a possible infection.

Immune response related to Pelibuey sheep naturally infected with gastrointestinal nematodes in a tropical region of Mexico.

We analysed the immune response involved in sheep naturally infected with gastrointestinal (GI) nematodes. Fifteen Pelibuey lambs were grazed in paddocks contaminated with GI nematodes for 13 weeks. To assess the infection, the number of eggs per gram (epg) and the percentage of packed cell volume (pcv) were evaluated. Blood and abomasal tissue samples were collected at week 8 post-infection to analyse the expression levels of IL-4, IL-5, IL-6, IL-8, IL-13, TGF-ß and FCεR1A genes. The nematode Haemonchus contortus was the main species identified. In addition, two groups of lambs were classified based on the x ± SE of epg and pcv values: G-1, with 151 ± 28 and 29 ± 0.33%, respectively, and G-2, with 475 ± 59.5 and 26 ± 0.38%, respectively. For G-1, upregulation of IL-4, IL-8, IL-13, TGF-ß and FCεR1A genes from 2.42- to 14.99-fold was observed in blood and abomasal tissue samples (p > .05), and IL-5, IL-8 and TGF-ß genes had significant gene expression levels in blood (p < .05). For G-2, moderate gene expression levels, ranging from 1.22- to 3.45-fold, were observed in abomasal tissue (p > .05), and the IL-5 gene presented significant gene expression in blood (p < .05). Strong positively correlated values (r) between pcv and IL-4, IL-8 and TGF-ß genes were observed in G-1. In contrast, significant negative correlations between epg and IL-4, IL-5 and FCεR1A genes indicate acute infection for G-2. Our results suggest that IL-4, IL-5, IL-13, TGF-ß and FCεR1A genes are important modulators of GI nematode infections of Pelibuey lambs.

Relationships among body condition score, FAMACHA© score and haematological parameters in Pelibuey ewes.

The objective of the present study was to determine the associations between the FAMACHA© score, body condition score (BCS) and several haematological parameters of Pelibuey ewes. In total, 52 clinically healthy Pelibuey ewes were evaluated in this study. A blood sample was collected from each animal to determine haematological variables. The BCS was evaluated on a scale of 1 to 5 and subsequently grouped into four categories: (1) < 1.5, (2) 2.0-2.5, (3) 3.0-3.5 and (4) > 4.0. The coloration of conjunctival mucosa was evaluated using the FAMACHA© system. The analysed haematological parameters were red blood cell (RBC) count, haemoglobin (HGB), haematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), red blood cell distribution width standard deviation (RDW-SD) and red blood cell distribution width coefficient of variation (RDW-CV). A descriptive statistical analysis was performed, and the Pearson correlation coefficients between variables were estimated. Haematological parameters were analysed with one-way analyses of variance, and the Tukey test was applied when significant differences were detected. The FAMACHA© score was only associated with MCHC (r = - 0.30, P < 0.05). The haematological parameters of RBC count, HCT, RDW-SD and RDW-CV statistically differed among BCS groups. The BCS was associated with RBC count, HCT, RDW-SD and RDW-CV, with r values ranging from 0.35 to 0.48. The haematological values were similar to those reported in ewes from Iran and Iraq. Hence, the optimization of the BCS calculation of Pelibuey ewes can generate association tables with haematological parameters to aid in the sanitary and feeding management of sheep.

Butlerius butleri (Nematoda: Diplogasteridae) Feeds on Haemonchus contortus (Nematoda: Trichostrongylidae) Infective Larvae and Free-Living Nematodes in Sheep Faecal Cultures Under Laboratory Conditions: Preliminary Report.

AIM: This study assessed the feeding activity of the predatory nematode Butlerius butleri on the infective larvae of the sheep parasitic nematode Haemonchus contortus (Hc-L3) and on two free-living nematodes Panagrellus redivivus (Pr) and Rhabditis sp. (R) in sterile sheep faecal cultures. MATERIALS AND METHODS: A bioassay was carried out in sheep faecal cultures, where predatory and prey nematodes were put together to assess the predatory behaviour on the different nematodes. Faecal cultures (n = 9) were set up in different treatments as follows: Treatments 1, 2 and 3 contained 20 specimens of B. butleri and 2000 Hc-L3, Pr and R, respectively; treatments 4, 5 and 6, contained only 2000 Hc-L3, Pr and R, respectively, as control groups. Treatment 7 contained 20 B. butleri as control. All treatments were incubated at room temperature for 30 days. The average number of nematodes recovered from each treatment was obtained and the reduction rate with respect to their corresponding control was estimated. Data were analysed using an ANOVA test using a completely randomised design in the SAS programme. RESULTS: Butlerius butleri reduced the Hc-L3, population by 91.8%, whereas its population increased 131.6 times. The interactions between B. butleri and Pr and R resulted in 80.9% and 62.5% nematode reduction and B. butleri increased its populations 268.3 and 82.7 times, respectively. This experiment was performed only once, so this can be considered a preliminary report at this time. CONCLUSION: This study provides evidence for the first time that B. butleri feeds on one of the most economically important nematodes of small ruminants, Hc-L3.

High energy levels in the diet reduce the parasitic effect of Haemonchus contortus in Pelibuey sheep.

Nutritional aspects modulate the parasitological and immune response in infected sheep. The aim of the present study was to evaluate the parasitological and humoral immune response of Pelibuey sheep experimentally infected with Haemonchus contortus that were fed diets with two different energy (n = 12) and protein (n = 12) levels. Twenty-four Pelibuey lambs infected with H. contortus were evaluated over 12 weeks. An additional six animals were considered as a control group. Fecal egg count (FEC) was determined, in addition to packed cell volume (PCV), total plasma protein (TPP), and immunoglobulin levels (IgA, IgG, and IgM) by indirect enzyme-linked immunosorbent assay (iELISA). Data were analyzed by repeated measures over time. The lambs that received a high-energy (HighE) diet had the lowest FEC (P < 0.01) regardless of whether they received a high (HighP) or low (LowP) level of protein. The effect of energy level was also observed over time: FEC values decreased and PCV and TPP values increased. Higher immunoglobulin levels were obtained for females (P < 0.05) than males yet, overall, the energy and protein levels of the diets did not affect the response of the immunoglobulins. Over time, however, an increase in IgG and IgM was observed, whereas the IgA level remained basal.

Excreted and secreted products (72/60 kDa) from Haemonchus placei larvae induce in vitro peripheral blood mononuclear cell proliferation and activate the expression of cytokines and FCεR1A receptor.

The aim of the present study was to assess the expression of cytokines and FCεR1A receptor stimulated by Haemonchus placei larval excretory and secretory (ES) products associated with the pathogenesis in calves. Bovine peripheral blood mononuclear cells (PBMC) were stimulated in in vitro assays with H. placei L4 ES product at 8, 12, 16 and 24 h. ES products were collected in in vitro assays at 48 h with molecular weight of 72/60 kDa and isoelectric point of 7.2 pI. Specific IgG for infected and control calves, positive and negative, were employed to recognise H. placei larval ES products by indirect ELISA, showing a mean of 1.8, 0.83 and 0.28 OD, respectively, (p ≤ 0.001). The quantification of relative gene expression was performed using a set of cytokines (IL-2, IFNγ, TGFß, IL-4, IL-5, IL-6, IL-8, IL-10 and IL-13), FCεR1A receptor and housekeeping (GAPDH, ß-actin and ß-2-microglobulin) by RT-qPCR. An early increased expression, 2.2- to 3.4-fold change, of IL-2 (p ≤ 0.001), IL-5 and TGFß (p ≥ 0.05) was determined, followed by TGFß (30.7 and 14.14), IL-8 (102.8 and 1504.4) and IL-10 (60.4 and 1.7) (p ≤ 0.05) after 12 and 16 h, respectively, and reducing the expression level at 24 h. In addition, IL-6, IL-13 and FCεR1A receptor also displayed mild expression level, 2.1 - to 7.60-fold change, at 24 h (p ≥ 0.05). We conclude that ES products of 72/60 kDa collected in vitro from H. placei larvae are recognised by infected hosts and have the ability to induce diverse immune factors to modulate the nematode damage.

Anthelmintic resistance in hair sheep farms in a sub-humid tropical climate, in the Huasteca Potosina, Mexico.

This is the first report about anthelmintic resistance (AR) in hair sheep farms determined in a sub-humid tropical climate, in an area known as Huasteca Potosina, Mexico. Faecal egg count reduction tests (FECRT) and egg hatch in vitro tests were conducted to identify the level of AR against benzimidazole (BZ) in parasitic gastrointestinal nematode (GIN) populations. An allele-specific polymerase chain reaction (AS-PCR) was performed to obtain a 250 bp band, indicating resistance, and a 550 bp band, indicating susceptibility to BZ. Macrocyclic lactones (ML) and imidazothiazole (IMZ) drugs were also tested with the FECRT. A PROBIT analysis was conducted using SAS to determine the 50% lethal doses (LD50) of the drugs according to the egg hatch in vitro test. Resistance to BZ and ML was found on all farms (0-70% effectiveness), whereas the susceptibility of nematodes to IMZ was detected with the FECRT (93-100% effectiveness). The LD50 was higher than the discriminating dose (0.1 µg ml-1) for BZ and confirmed AR to this anthelmintic; we also confirmed a high AR frequency with AS-PCR. Therefore, we suggest that strategic deworming should be performed to avoid the development of resistance to imidazothiazole.

Variation in phenotypic resistance to gastrointestinal nematodes in hair sheep in the humid tropics of Mexico.

The objective of the study was to evaluate phenotypic resistance against gastrointestinal nematodes in Blackbelly, Pelibuey and Katahdin ewes before pregnancy in the humid tropics of Mexico. Individual faecal and blood samples were taken in 59 Pelibuey, 69 Blackbelly and 73 Katahdin ewes. The egg count per gram of faeces (EPG) of gastrointestinal nematodes (GINs) was determined. The percentage of packed cell volume (PCV) and body condition score (BCS) of each animal were also recorded. The ewes were segregated as susceptible, intermediate or resistant based on the EPG using the quartile method. The data were analysed using the general linear method, and the means between breeds were compared by Tukey's test. The relationships between the EPG, PCV and BCS were evaluated by Spearman correlation. The Katahdin ewes showed the highest EPG counts (3613.6 ± 5649) compared to the Blackbelly and Pelibuey ewes (576.1 ± 1009 and 56.8 ± 187, respectively, P < 0.01). The PCV values between breeds were similar (P > 0.05). The susceptible ewes had the highest EPG counts and the lowest PCV percentage (5069 ± 6404 and 22.8% ± 8.1% respectively) compared to the resistant ewes (P < 0.01). A higher percentage of Katahdin ewes were susceptible compared to the other breeds (P < 0.05). The main GIN species were Haemonchus contortus, Trichostrongylus colubriformis and Cooperia curticei. In conclusion, Katahdin ewes showed susceptibility to GIN compared to Blackbelly and Pelibuey ewes before the pregnancy period in the humid tropics of Mexico.